减数分裂的研究进展

No abstract available

BACKGROUND During mammalian oocyte meiosis, accurate chromosome segregation critically depends on precise regulation of kinetochore-microtubule (K-MT) attachments, a process monitored by the spindle assembly checkpoint (SAC). While CENP-F has been well characterized as a kinetochore-associated protein that stabilizes K-MT connections during mitosis, its functional mechanisms during meiosis remain poorly understood. In particular, there is still controversy over whether farnesylation modification governs localization and functionality of CENP-F. Concurrently, clinical investigations face a knowledge gap regarding the genetic basis of oocyte maturation arrest, a prevalent phenotype observed in female infertility patients. OBJECTIVE This study aims to reveal the regulatory mechanism of CENP-F farnesylation modification on its meiotic function and explore the association between CENP-F gene mutations and female oocyte maturation disorders. STUDY DESIGN Previous studies have shown that CENP-F is essential for chromosome segregation during mitosis, but its functional mechanism during meiosis remains poorly understood. Oocyte microinjection, western blotting, co-immunoprecipitation (Co-IP), and immunofluorescence were used to explore the localization and function of CENP-F in oocytes. The role of CENP-F farnesylation in mouse oocytes was investigated using pharmacological (farnesyltransferase inhibitor treatment) and genetic (C3111S point mutation) methods. Subsequently, four patients with CENP-F mutations were identified in the whole-exome sequencing (WES) dataset consisting of 179 infertile patients with oocyte maturation disorders. Mouse oocyte and 293T cell models were used to verify the mechanism of patient-derived CENP-F mutations causing oocyte maturation disorders. RESULTS Microinjection of Cenp-f siRNA into mouse oocytes significantly reduced maturation rates (77.84±2.087% vs 34.26±4.748%, P<.01), with the majority arrested at metaphase I (MI) (17.69±2.207% vs 44.93±5.539%, P<.05). Time-course immunofluorescence analysis revealed dynamic CENP-F localization: initially dispersed across chromosome following nuclear envelope breakdown (NEBD), then progressively accumulating at kinetochores by MI. Co-IP assays confirmed a direct interaction between CENP-F and AURKB. Knockdown of AURKB would damage the kinetochore localization of CENP-F in oocytes. Farnesylation inhibition (via farnesyltransferase inhibitor or C3111S mutation) significantly decreased oocyte maturation rates (75.58±3.703% vs 46.18±1.282%, P<.01; 75.58±3.703% vs 44.04±2.541%, P<.01), concomitantly weakening interaction between CENP-F and AURKB (P<.01) and disrupting kinetochore localization. Genetic screening identified four CENP-F mutations in 179 infertile women with oocyte maturation arrest. Microinjection of patient-derived mutant CENP-F cRNAs into mouse oocytes significantly reduced maturation rates (77.00±2.411% vs 49.10±6.561%, P<.01; 77.00±2.411% vs35.43±1.035%, P<.01; 77.00±2.411% vs 55.43±1.288%, P<.05; 77.00±2.411% vs 40.00±4.187%, P<.01). Two of these mutations (K1708T/S1971fs) can reduce the farnesylation of CENP-F (P<.05/P<.01), damage its interaction with AURKB (P<0.05/P<0.01), and disrupt the kinetochore localization. Both CENP-F depletion and patient mutations induced constitutive SAC activation, and the treatment with SAC inhibitor partially rescued the meiotic arrest phenotype in oocytes (P<.05). CONCLUSION This study represents the first demonstration of a direct association between CENP-F genetic defects and human infertility, uncovering a novel farnesylation-dependent mechanism that governs meiotic progression, while simultaneously identifying CENP-F as a potential molecular marker for diagnosing oocyte maturation failure.

Androgen receptor (AR) signaling in Sertoli cells is known to be important for germ-cell progression through meiosis, but the extent to which androgens indirectly regulates specific meiosis stages is not known. Here, we combine synchronization of spermatogenesis, cytological analyses and single-cell RNAseq (scRNAseq) in the Sertoli cell androgen receptor knockout (SCARKO) mutant and control mice, and demonstrate that SCARKO mutant spermatocytes exhibited normal expression and localization of key protein markers of meiotic prophase events, indicating that initiation of meiotic prophase is not androgen dependent. However, spermatocytes from SCARKO testes failed to acquire competence for the meiotic division phase. ScRNAseq analysis of wild type and SCARKO mutant testes revealed a molecular transcriptomic block in an early meiotic prophase state (leptotene/zygotene) in mutant germ cells, and identified several misregulated genes in SCARKO Sertoli cells, many of which have been previously implicated in male infertility. Together, our coordinated cytological and single-cell RNAseq analyses identified germ-cell intrinsic and extrinsic genes responsive to Sertoli-cell androgen signaling that promotes cellular states permissive for the meiotic division phase.

Kinesin family member 3A (KIF3A) is a microtubule-oriented motor protein that belongs to the kinesin-2 family for regulating intracellular transport and microtubule movement. In this study, we characterized the critical roles of KIF3A during mouse oocyte meiosis. We found that KIF3A associated with microtubules during meiosis and depletion of KIF3A resulted in oocyte maturation defects. LC–MS data indicated that KIF3A associated with cell cycle regulation, cytoskeleton, mitochondrial function and intracellular transport-related molecules. Depletion of KIF3A activated the spindle assembly checkpoint, leading to metaphase I arrest of the first meiosis. In addition, KIF3A depletion caused aberrant spindle pole organization based on its association with KIFC1 to regulate expression and polar localization of NuMA and γ-tubulin; and KIF3A knockdown also reduced microtubule stability due to the altered microtubule deacetylation by histone deacetylase 6 (HDAC6). Exogenous Kif3a mRNA supplementation rescued the maturation defects caused by KIF3A depletion. Moreover, KIF3A was also essential for the distribution and function of mitochondria, Golgi apparatus and endoplasmic reticulum in oocytes. Conditional knockout of epithelial splicing regulatory protein 1 (ESRP1) disrupted the expression and localization of KIF3A in oocytes. Overall, our results suggest that KIF3A regulates cell cycle progression, spindle assembly and organelle distribution during mouse oocyte meiosis.

Meiosis produces haploid gametes that are essential for sexual reproduction in most eukaryotes, but the specific cell-cycle regulation that controls meiotic progression in plants remains to be fully characterized. KNUCKLES (KNU), a known transcriptional repressor in floral meristem regulation, also regulates anther development. Loss of KNU function leads to male sterility in Arabidopsis, but its detailed regulatory mechanism is unknown. Here, we find that KNU is specifically localized in meiocytes during anther development, and mutation of KNU disrupts meiotic progression and behavior, leading to apoptosis of microsporocytes. Transcriptome analysis shows that numerous genes related to meiosis are downregulated in knu-2 meiocytes. We demonstrate that KNU can directly repress the expression of two cell-cycle inhibitors, INTERACTOR/INHIBITOR OF CDK 1/KIP-RELATED PROTEIN 1 (ICK1/KRP1) and KRP3, and knockout of KRP1 or KRP3 in the null allele knu-2 background largely rescues the knu-2 defects in male meiosis and fertility. Consistent with these results, overexpression of KRP1 driven by the native KNU promoter results in meiotic defects and reduced expression of some meiosis-related genes, similar to the phenotypes of knu-2. Thus, our findings provide evidence that the transcription factor KNU regulates the expression of meiotic cell-cycle regulators and cohesins through suppression of KRP1/3, significantly broadening our understanding of plant meiosis.

The generation of a high-quality egg for reproduction requires faithful segregation of chromosome during oocyte meiosis. Here, we report that echinoderm microtubule-associated protein like 6 (EML6) is highly expressed in oocytes, and responsible for accurate segregation of homologous chromosomes in mice. Quantitative real-time RT-PCR and immunohistochemistry analyses revealed that EML6 was predominantly expressed by oocytes in the ovaries. Whole mount immunofluorescent staining showed that EML6 was colocalized with spindle microtubules in oocytes at various stages after meiotic resumption. This specialized localization was disrupted by nocodazole, the microtubule destabilizer, while enhanced by Taxol, a microtubule stabilizing reagent. In vivo knockdown of Eml6 expression by the specific siRNA resulted in chromosome misalignment and alteration in spindle dimension at both metaphase Ⅰ and Ⅱ stages, as well as the increased aneuploidy in the mature oocytes. Thus, these data suggest that EML family proteins participate in the control of oocyte meiotic division.

Abstract Pachynema progression contributes to the completion of prophase I. Nevertheless, the regulation of this significant meiotic process remains poorly understood. In this study, we identified a novel testis-specific protein HSF5, which regulates pachynema progression during male meiosis in a manner dependent on chromatin-binding. Deficiency of HSF5 results in meiotic arrest and male infertility, characterized as unconventional pachynema arrested at the mid-to-late stage, with extensive spermatocyte apoptosis. Our scRNA-seq data confirmed consistent expressional alterations of certain driver genes (Sycp1, Msh4, Meiob, etc.) crucial for pachynema progression in Hsf5−/− individuals. HSF5 was revealed to primarily bind to promoter regions of such key divers by CUT&Tag analysis. Also, our results demonstrated that HSF5 biologically interacted with SMARCA5, SMARCA4 and SMARCE1, and it could function as a transcription factor for pachynema progression during meiosis. Therefore, our study underscores the importance of the chromatin-associated HSF5 for the differentiation of spermatocytes, improving the protein regulatory network of the pachynema progression.

In mammals, the production of mature oocytes necessitates rigorous regulation of the discontinuous meiotic cell-cycle progression at both the transcriptional and post-transcriptional levels. However, the factors underlying this sophisticated but explicit process remain largely unclear. Here we characterize the function of N-acetyltransferase 10 (Nat10), a writer for N4-acetylcytidine (ac4C) on RNA molecules, in mouse oocyte development. We provide genetic evidence that Nat10 is essential for oocyte meiotic prophase I progression, oocyte growth and maturation by sculpting the maternal transcriptome through timely degradation of poly(A) tail mRNAs. This is achieved through the ac4C deposition on the key CCR4-NOT complex transcripts. Importantly, we devise a method for examining the poly(A) tail length (PAT), termed Hairpin Adaptor-poly(A) tail length (HA-PAT), which outperforms conventional methods in terms of cost, sensitivity, and efficiency. In summary, these findings provide genetic evidence that unveils the indispensable role of maternal Nat10 in oocyte development.

Meiosis occurs specifically in germ cells to produce sperm and oocytes that are competent for sexual reproduction. Multiple factors are required for successful meiotic entry, progression, and termination. Among them, trimethylation of histone H3 on lysine 4 (H3K4me3), a mark of active transcription, has been implicated in spermatogenesis by forming double-strand breaks (DSBs). However, the role of H3K4me in transcriptional regulation during meiosis remains poorly understood. Here, we reveal that mouse CXXC finger protein 1 (Cfp1), a component of the H3K4 methyltransferase Setd1a/b, is dynamically expressed in differentiating male germ cells and safeguards meiosis by controlling gene expression. Genetic ablation of mouse CFP1 in male germ cells caused complete infertility with failure in prophase I of the 1st meiosis. Mechanistically, CFP1 binds to genes essential for spermatogenesis, and its loss leads to a reduction in H3K4me3 levels and gene expression. Importantly, CFP1 is highly enriched within the promoter/TSS of target genes to elevate H3K4me3 levels and gene expression at the pachytene stage of meiotic prophase I. The most enriched genes were associated with meiosis and homologous recombination during the differentiation of spermatocytes to round spermatids. Therefore, our study establishes a mechanistic link between CFP1-mediated transcriptional control and meiotic progression and might provide an unprecedented genetic basis for understanding human sterility. Details of the role of a protein in the development of sperm cells in mice could lead to new understanding of sterility in men. An international research team led by Youngsok Choi and Kwonho Hong at Konkuk University, Seoul, South Korea, investigated the role of protein Cfp1, which they found to be required for sperm formation in mice. The protein is a component of an enzyme complex that transfers methyl groups (CH3) onto other proteins involved in controlling gene activity. The researchers identified key aspects of the mechanism by which Cfp1 controls the activity of genes essential for sperm formation to proceed normally. Absence of Cfp1 specifically interferes with the process of meiosis, which generates sperm cells containing only one copy of each chromosome instead of the two copies found in other cells.

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During meiotic prophase, the essential events of homolog pairing, synapsis, and recombination are coordinated with meiotic progression to promote fidelity and prevent aneuploidy. The conserved AAA+ ATPase PCH-2 coordinates these events to guarantee crossover assurance and accurate chromosome segregation. How PCH-2 accomplishes this coordination is poorly understood. Here, we provide evidence that PCH-2 decelerates pairing, synapsis and recombination in C. elegans by remodeling meiotic HORMADs. We propose that PCH-2 converts the closed versions of these proteins, which drive these meiotic prophase events, to unbuckled conformations, destabilizing interhomolog interactions and delaying meiotic progression. Further, we find that PCH-2 distributes this regulation among three essential meiotic HORMADs in C. elegans: PCH-2 acts through HTP-3 to regulate pairing and synapsis, HIM-3 to promote crossover assurance, and HTP-1 to control meiotic progression. In addition to identifying a molecular mechanism for how PCH-2 regulates interhomolog interactions, our results provide a possible explanation for the expansion of the meiotic HORMAD family as a conserved evolutionary feature of meiosis. Taken together, our work demonstrates that PCH-2’s remodeling of meiotic HORMADs has functional consequences for the rate and fidelity of homolog pairing, synapsis, recombination and meiotic progression, ensuring accurate meiotic chromosome segregation.

Background In mitotic cells, WAPL acts as a cohesin release factor to remove cohesin complexes from chromosome arms during prophase to allow the accurate chromosome segregation in anaphase. However, we have recently documented that Wapl exerts a unique meiotic function in the spindle assembly checkpoint (SAC) control through maintaining Bub3 stability during mouse oocyte meiosis I. Whether this noncanonical function is conserved among species is still unknown. Methods We applied RNAi-based gene silencing approach to deplete WAPL in porcine oocytes, validating the conserved roles of WAPL in the regulation of SAC activity during mammalian oocyte maturation. We also employed immunostaining, immunoblotting and image quantification analyses to test the WAPL depletion on the meiotic progression, spindle assembly, chromosome alignment and dynamics of SAC protein in porcine oocytes. Results We showed that depletion of WAPL resulted in the accelerated meiotic progression by displaying the precocious polar body extrusion and compromised spindle assembly and chromosome alignment. Notably, we observed that the protein level of BUB3 was substantially reduced in WAPL-depleted oocytes, especially at kinetochores. Conclusions Collectively, our data demonstrate that WAPL participates in the porcine oocyte meiotic progression through maintenance of BUB3 protein levels and SAC activity. This meiotic function of WAPL in oocytes is highly conserved between pigs and mice.

Several developmental stages of spermatogenesis are transcriptionally quiescent which presents major challenges associated with the regulation of gene expression. Here we identify that the zygotene to pachytene transition is not only associated with the resumption of transcription but also a wave of programmed mRNA degradation that is essential for meiotic progression. We explored whether terminal uridydyl transferase 4- (TUT4-) or TUT7-mediated 3′ mRNA uridylation contributes to this wave of mRNA degradation during pachynema. Indeed, both TUT4 and TUT7 are expressed throughout most of spermatogenesis, however, loss of either TUT4 or TUT7 does not have any major impact upon spermatogenesis. Combined TUT4 and TUT7 (TUT4/7) deficiency results in embryonic growth defects, while conditional gene targeting revealed an essential role for TUT4/7 in pachytene progression. Loss of TUT4/7 results in the reduction of miRNA, piRNA and mRNA 3′ uridylation. Although this reduction does not greatly alter miRNA or piRNA expression, TUT4/7-mediated uridylation is required for the clearance of many zygotene-expressed transcripts in pachytene cells. We find that TUT4/7-regulated transcripts in pachytene spermatocytes are characterized by having long 3′ UTRs with length-adjusted enrichment for AU-rich elements. We also observed these features in TUT4/7-regulated maternal transcripts whose dosage was recently shown to be essential for sculpting a functional maternal transcriptome and meiosis. Therefore, mRNA 3′ uridylation is a critical determinant of both male and female germline transcriptomes. In conclusion, we have identified a novel requirement for 3′ uridylation-programmed zygotene mRNA clearance in pachytene spermatocytes that is essential for male meiotic progression.

Plant spermatogenesis is a complex process that directly affects crop breeding. A rapid change in gene abundance occurs at early meiosis prophase, when gene regulation is selective. However, how these genes are regulated remains unknown. Here, we show that rice reproductive phasiRNAs are essential for the elimination of a specific set of RNAs during meiotic prophase I. These phasiRNAs cleave target mRNAs in a regulatory manner such that one phasiRNA can target more than one gene, and/or a single gene can be targeted by more than one phasiRNA to efficiently silence target genes. Our investigation of phasiRNA-knockdown and PHAS-edited transgenic plants demonstrates that phasiRNAs and their nucleotide variations are required for meiosis progression and fertility. This study highlights the importance of reproductive phasiRNAs for the reprogramming of gene expression during meiotic progression and establishes a basis for future studies on the roles of phasiRNAs with a goal of crop improvement. 21-nucleotide phased secondary siRNAs are expressed in the developing rice anthers during meiosis. Here, the authors show that phasiRNAs can cleave target mRNA and act to ensure the progression of meiosis and fertility in rice.

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Epigenetic modifiers are emerging as important regulators of the genome. However, how they regulate specific processes during meiosis is not well understood. Methylation of H3K79 by the histone methyltransferase Dot1 has been shown to be involved in the maintenance of genomic stability in various organisms. In S. cerevisiae, Dot1 modulates the meiotic checkpoint response triggered by synapsis and/or recombination defects by promoting Hop1-dependent Mek1 activation and Hop1 distribution along unsynapsed meiotic chromosomes, at least in part, by regulating Pch2 localization. However, how this protein regulates meiosis in metazoans is unknown. Here, we describe the effects of H3K79me depletion via analysis of dot-1.1 or zfp-1 mutants during meiosis in Caenorhabditis elegans. We observed decreased fertility and increased embryonic lethality in dot-1.1 mutants suggesting meiotic dysfunction. We show that DOT-1.1 plays a role in the regulation of pairing, synapsis and recombination in the worm. Furthermore, we demonstrate that DOT-1.1 is an important regulator of mechanisms surveilling chromosome synapsis during meiosis. In sum, our results reveal that regulation of H3K79me plays an important role in coordinating events during meiosis in C. elegans.

During spermatogenesis, meiosis is accompanied by a robust alteration in gene expression and chromatin status. However, it remains elusive how the meiotic transcriptional program is established to ensure completion of meiotic prophase. Here, we identify a protein complex that consists of germ-cell-specific zinc-finger protein ZFP541 and its interactor KCTD19 as the key transcriptional regulators in mouse meiotic prophase progression. Our genetic study shows that ZFP541 and KCTD19 are co-expressed from pachytene onward and play an essential role in the completion of the meiotic prophase program in the testis. Furthermore, our ChIP-seq and transcriptome analyses identify that ZFP541 binds to and suppresses a broad range of genes whose function is associated with biological processes of transcriptional regulation and covalent chromatin modification. The present study demonstrates that a germ-cell specific complex that contains ZFP541 and KCTD19 promotes the progression of meiotic prophase towards completion in male mice, and triggers the reconstruction of the transcriptional network and chromatin organization leading to post-meiotic development. The authors add to our knowledge of the transcriptional regulation of the meiotic program in mice spermatocytes, showing ZFP541 regulates meiotic prophase and transition to the division phase by being the target for upstream factors MEIOSIN/STRA8.

CK2 (casein kinase 2) is a serine/threonine-selective protein kinase that has been involved in a variety of cellular processes such as DNA repair, cell cycle control and circadian rhythm regulation. However, its functional roles in oocyte meiosis have not been fully determined. We report that CK2 is essential for porcine oocyte meiotic maturation by regulating spindle assembly checkpoint (SAC). Immunostaining and immunoblotting analysis showed that CK2 was constantly expressed and located on the chromosomes during the entire oocyte meiotic maturation. Inhibition of CK2 activity by its selective inhibitor CX-4945 impaired the first polar body extrusion and arrested oocytes at M I stage, accompanied by the presence of BubR1 at kinetochores, indicative of activated SAC. In addition, we found that spindle/chromosome structure was disrupted in CK2-inhibited oocytes due to the weakened microtubule stability, which is a major cause resulting in the activation of SAC. Last, we found that the level DNA damage as assessed by γH2A.X staining was considerably elevated when CK2 was inhibited, suggesting that DNA damage might be another critical factor leading to the SAC activation and meiotic failure of oocytes. Our findings demonstrate that CK2 promotes the porcine oocyte maturation by ensuring normal spindle assembly and DNA damage repair.

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Oocyte meiotic progression and maternal-to-zygote transition are accompanied by dynamic epigenetic changes. The functional significance of these changes and the key epigenetic regulators involved are largely unknown. Here we show that Setdb1, a lysine methyltransferase, controls the global level of histone H3 lysine 9 di-methyl (H3K9me2) mark in growing oocytes. Conditional deletion of Setdb1 in developing oocytes leads to meiotic arrest at the germinal vesicle and meiosis I stages, resulting in substantially fewer mature eggs. Embryos derived from these eggs exhibit severe defects in cell cycle progression, progressive delays in preimplantation development, and degeneration before reaching the blastocyst stage. Rescue experiments by expressing wild-type or inactive Setdb1 in Setdb1-deficient oocytes suggest that the catalytic activity of Setdb1 is essential for meiotic progression and early embryogenesis. Mechanistically, up-regulation of Cdc14b, a dual-specificity phosphatase that inhibits meiotic progression, greatly contributes to the meiotic arrest phenotype. Setdb1 deficiency also leads to derepression of transposons and increased DNA damage in oocytes, which likely also contribute to meiotic defects. Thus, Setdb1 is a maternal-effect gene that controls meiotic progression and is essential for early embryogenesis. Our results uncover an important link between the epigenetic machinery and the major signaling pathway governing meiotic progression.

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Upon nitrogen starvation, Schizosaccharomyces pombe exit the mitotic cell cycle and become irreversibly committed to the completion of meiosis program. Meiotic cell divisions are coordinated with sporulation events to produce haploid spores. In the last few decades, experiments on fission yeast have revealed different molecular players involved in two meiotic cell divisions, meiosis I (MI) and meiosis II (MII). How the MI entry, MI-to-MII transition, and MII exit occur because of the dynamics of the regulatory network is not well understood. In this work, we developed a comprehensive mathematical model of the network that describes the temporal dynamics of meiotic progression. The model accounts for the phenotypes of several experimental data (single and multiple mutations). We demonstrate the control strategy involving multiple feedback loops to yield two successive division cycles. The differential regulation of anaphase-promoting complex/cyclosome (APC/C) coactivators and its inhibitors is crucial for the dynamics of both MI-to-MII transition and MII exit. This model generates mechanistic insights that help in further experiments and modeling.

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Abstract Enhancer of zeste homolog 2 (EZH2) has been extensively investigated to participate in diverse biological processes, including carcinogenesis, the cell cycle, X-chromosome inactivation, and early embryonic development. However, the functions of this protein during mammalian oocyte meiotic maturation remain largely unexplored. Here, combined with RNA-Seq, we provided evidence that EZH2 is essential for oocyte meiotic maturation in pigs. First, EZH2 protein expression increased with oocyte progression from GV to MII stage. Second, the siRNA-mediated depletion of EZH2 led to accelerated GVBD and early occurrence of the first polar body extrusion. Third, EZH2 knockdown resulted in defective spindle assembly, abnormal SAC activity, and unstable K-MT attachment, which was concomitant with the increased rate of aneuploidy. Finally, EZH2 silencing exacerbated oxidative stress by increasing ROS levels and disrupting the distribution of active mitochondria in porcine oocytes. Furthermore, parthenogenetic embryonic development was impaired following the depletion of EZH2 at GV stage. Taken together, we concluded that EZH2 is necessary for porcine oocyte meiotic progression through regulating spindle organization, maintaining chromosomal integrity, and mitochondrial function. Summary sentence EZH2, an important maternal factor, is demonstrated to have a non-classical function: participating in porcine oocyte meiosis through modulating spindle organization and maintaining chromosomal integrity. Graphical Abstract

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Meiotic progression in S. pombe is regulated by stage-specific gene expression and translation, changes in RNA stability, expression of anti-sense transcripts, and targeted proteolysis of regulatory proteins. We have used SILAC labeling to examine the relative levels of proteins in diploid S. pombe cells during meiosis. Among the 3,268 proteins quantified at all time points, the levels of 880 proteins changed at least 2-fold; the majority of proteins showed stepwise increases or decreases during the meiotic divisions, while some changed transiently. Overall, we observed reductions in proteins involved in anabolism and increases in proteins involved in catabolism. We also observed increases in the levels of proteins of the ESCRT-III complex and revealed a role for ESCRT-III components in chromosome segregation and spore formation. Correlation with studies of meiotic gene expression and ribosome occupancy reveals that many of the changes in steady-state protein levels are post-transcriptional.

Abstract Epigenetic reprogramming is widespread and highly active during gametogenesis, which is usually involving in the expression of critical genes. The expression of genes couple with transcription activation, and the transcriptional regulation by transcription factors predetermine protein translation for biological processes. In this study, we found that EHMT1-mediated re-establishment of H3K9me2 played crucial roles in the progression of meiosis in female germ cells. EHMT1 deficient female mice were nearly infertile due to the arrest of zygotene in embryonic germ cells, which was caused by downregulated expression of key meiotic genes. Furthermore, we identified transcription suppressor, particularly ST18, for meiotic genes by combining RNA-seq, Cut&Tag seq analysis, and luciferase reporter assays. We uncovered that H3K9me2 mediated ST18 expression homeostasis and played critical roles in regulating the timed expression of key meiotic genes. Overall, we revealed that EHMT1-mediated H3K9me2 re-establishment facilitated the expression of key meiotic genes for female meiosis progression.

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What is the function of ASZ1 in regulating meiosis, particularly in piRNA-independent pathways? ASZ1 sustains meiotic progression by dual epigenetic mechanisms: DNA methylation and histone modifications, with deficiency leading to meiosis arrest and azoospermia. Meiosis is an evolutionarily conserved process precisely regulated by thousands of genes through epigenetic pathways, including DNA methylation, histone modifications and PIWI/piRNA pathway. ASZ1 is a piRNA protein involved in the post-transcriptional regulation of genes critical for meiosis. However, the cross-link between ASZ1 and piRNA-independent pathways during meiosis remains poorly understood. Whole-exome sequencing was performed for 626 individuals with NOA from the Reproductive and Genetic Hospital of CITIC-Xiangya between 2014 and 2022 to explore the underlying genetic factors. After a stringent variant screening strategy, two novel homozygous ASZ1 splicing variants were identified in two patients from two unrelated consanguineous families. Moreover, we constructed ASZ1-mutant mice mimicking patient mutations and combined molecular biology tools and multi-omics sequencing to reveal the role of ASZ1 in meiosis. Genetic analysis was conducted using whole-exome sequencing and Sanger sequencing of two patient cohorts with candidate homozygous ASZ1 splicing variants. Two patients with ASZ1 variants and ASZ1-mutant mice were analyzed. Methods included Molecular biology experiments such as western blotting, immunofluorescence, IP assay, and multi-omics sequencing like RNA-seq, Small RNA-seq, Whole Genome Bisulfite Sequencing analysis, RIP analysis, and Mass spectrometry analysis to explore ASZ1’s function in meiosis and epigenetic regulation. We identified two novel pathogenic homozygous ASZ1 variants in two azoospermia-affected individuals due to meiosis arrest. Mutant mice mimicking the patients’ mutation recapitulated the azoospermia phenotype, manifesting as meiotic zygotene arrest and aberrant “winding-shaped” expression of the lateral element of the synaptonemal complex in spermatocytes. Transcriptional and DNA methylation analyses demonstrate that ASZ1 deficiency gave rise to an increase in methylation of meiosis-related genes and a decrease in methylation of repeats, including transposable elements (TEs). RIP-seq revealed that ASZ1 impacts the methylation of TEs by directly binding to piRNA precursors. Mechanistically, ASZ1 cooperates with histone demethylation protein LSD1 to regulate the expression of meiosis-related genes through impacting H3K4me1/2 modification. The study is limited by the small sample size of human patients and the need for further validation of ASZ1’s interactions with other epigenetic regulators. Our findings uncover an essential role of ASZ1 in sustaining the progression of meiotic prophase I through dual mechanisms involving DNA methylation and histone modification, which provide prospective molecular targets for the clinical diagnosis and therapy of infertility. No

Objective Oocyte quality is critical for the stable transmission of genetic information and affects early embryonic development. But the precise mechanisms governing oocyte meiotic progression remains largely unclear. Transcription regulation through chromatin compaction and decompaction is regulated through various chromatin-remodeling complexes such as nucleosome remodeling and histone deacetylation (NuRD) complex. GATAD2B is known to be a component of the NuRD complex but whether GATAD2B regulates chromatin modification in mouse oocyte meiosis remains unclear. We hope to explore the role of GATAD2B in mouse oocyte meiosis. Methods In this study, we initially utilized western blot and immunofluorescence to delineate the expression and subcellular localization of GATAD2B during oocyte meiotic maturation. To further elucidate the role of GATAD2B in regulating oocyte meiotic division, we employed the method of microinjection of Gatad2b-specific siRNA to knock down the protein expression of GATAD2B. Subsequently, dynamic changes in oocyte meiotic division were captured in real-time using live-cell imaging with Geri. Additionally, spindle staining, DNA staining, spread analysis, and reanalysis of RNA-seq data were performed to investigate the mechanisms through which GATAD2B regulates oocyte meiotic division. Results GATAD2B was stably expressed during oocyte meiosis and was significantly increased during the metaphase II (MII) stage. To further explore the effect of GATAD2B on oocyte meiotic maturation, we observed increased abnormal spindle, severe chromosome misalignment and metaphase I (MI) block in GATAD2B knocked-down (GATAD2B-KD) oocytes. Interestingly, the distribution of microtubule organizing center was abnormal and aneuploidy was significantly increased in GATAD2B-KD oocytes. In addition, some deacetylation-related genes were significantly downregulated and acetylated proteins accumulated abnormally in GATAD2B-KD oocytes. Conclusion These findings implicate GATAD2B as a novel regulator of histone deacetylation during oocyte maturation and provide evidence that such deacetylation is required for proper spindle assembly.

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Regulation of the transcriptome to promote meiosis is important for sperm development and fertility. However, how chromatin remodeling directs the transcriptome during meiosis in male germ cells is largely unknown. Here, we demonstrate that the ISWI family ATP-dependent chromatin remodeling factor SMARCA5 (SNF2H) plays a critical role in regulating meiotic prophase progression during spermatogenesis. Males with germ cell-specific depletion of SMARCA5 are infertile and unable to form sperm. Loss of Smarca5 results in failure of meiotic progression with abnormal spermatocytes beginning at the pachytene stage and an aberrant global increase in chromatin accessibility, especially at genes important for meiotic prophase.

An important aim of population genetics is to elucidate the processes affecting genetic diversity across regions of the genome and across species. Canonical population genetic models of sexually reproducing species define the rate of meiotic recombination in terms of the frequency of recombination events per site per sexual generation. This paper studies the interplay of several factors with respect to their effects on neutral genetic diversity in a facultatively sexual, diploid, unicellular species such as yeast. The relevant factors are the prevalence of meiosis versus mitosis, the recombination rate, and the selection and dominance coefficients at loci under positive selection. We assume that many generations of mitotic cell divisions are interspersed with episodes of sexual reproduction, in which all individuals in the population undergo meiosis, followed by random matings among the resulting gametes. Our findings reveal that a single hard selective sweep can reduce neutral nucleotide site diversity across the entire genome, provided that the frequency of meiotic events is sufficiently low, and that the effects of a selective sweep on levels of neutral diversity at sites linked to the target of selection can be markedly different from those predicted by standard models of sweeps based on obligate sexuality. Species that reproduce by facultative sex are thus likely to exhibit unusual patterns of genetic diversity. Author summary In this study, we explored how different sexual strategies influence genetic diversity. Specifically, we looked at how the balance between sexual reproduction (which involves meiosis) and asexual reproduction (which involves mitosis) affect genetic variation in a species like yeast. Our research focused on the effects of recombination rates and the role of selective sweeps — when a beneficial genetic variant spreads rapidly through a population — on genetic diversity. We found that, in species with facultative sex, a selective sweep can dramatically reduce genetic diversity across the genome, but only if sexual reproduction occurs infrequently. We compared our findings to existing models and also developed a new mathematical framework for understanding the effects of sweeps when sexual reproduction is rare. The fact that facultative sex can reduce genetic diversity over the entire genome is likely to complicate inferences about population size and evolutionary dynamics in species with mixed reproductive strategies.

During meiotic recombination, homologue-templated repair of programmed DNA double-strand breaks (DSBs) produces relatively few crossovers and many difficult-to-detect non-crossovers. By intercrossing two diverged mouse subspecies over five generations and deep-sequencing 119 offspring, we detect thousands of crossover and non-crossover events genome-wide with unprecedented power and spatial resolution. We find that both crossovers and non-crossovers are strongly depleted at DSB hotspots where the DSB-positioning protein PRDM9 fails to bind to the unbroken homologous chromosome, revealing that PRDM9 also functions to promote homologue-templated repair. Our results show that complex non-crossovers are much rarer in mice than humans, consistent with complex events arising from accumulated non-programmed DNA damage. Unexpectedly, we also find that GC-biased gene conversion is restricted to non-crossover tracts containing only one mismatch. These results demonstrate that local genetic diversity profoundly alters meiotic repair pathway decisions via at least two distinct mechanisms, impacting genome evolution and Prdm9-related hybrid infertility. During meiotic recombination, genetic information is transferred or exchanged between parental chromosome copies. Using a large hybrid mouse pedigree, the authors generated high-resolution maps of these transfer/exchange events and discovered new properties governing their processing and resolution.

Meiosis prepares the genome for sexual reproduction and is a driver of genetic diversity. Essential for both aspects are rapid chromosome movements enabling the pairing of homologous chromosomes, thereby promoting their recombination and ensuring their correct segregation afterward. However, it is unknown how meiocytes reorganize their nuclear architecture when chromosomes are tethered to the nuclear lamina in somatic cells. Here, we reveal the meiosis-specific Skp1-Cul1-F-box-protein (SCF) E3 ubiquitin ligase SCFRMF as a key regulator of this reorganization in Arabidopsis. We find that the nuclear lamina proteins of the CRWN family are degraded in an SCFRMF-dependent manner in this process. Thus, the SCFRMF functions to release the somatic constraints on chromosome mobility through disrupting the nuclear lamina, allowing homologous chromosomes to pair.

Rationale: Meiotic homologous recombination is a critical event in gametogenesis, which is tightly regulated to ensure the generation of crossovers on homologous chromosomes. This process is crucial for ensuring the accurate segregation of genetic material and maintaining genetic diversity within species, ultimately contributing to reproductive success. Nevertheless, comprehensive mechanisms of post-translational modification (PTM) regulating homologous recombination during meiosis require further investigation. The aim of this study is to investigate the regulatory mechanisms and physiological functions of NAE1-mediated neddylation during meiosis of mammalian spermatogenesis and its consequential role in infertility. Methods: The dynamic localization of NAE1 at various sub-stages during spermatogenesis was determined using immunofluorescence staining and seminiferous tubule staging. We explore the role of NAE1-mediated neddylation by utilizing germ cell-specific Nae1-knockout mice. The impact on homologous synapsis and recombination during the meiosis prophase I were verified through chromosome spread fluorescence staining. We used 10 × Genomics single cell transcriptomics and ubiquitinomics to analysis the causes of spermatogenesis arrest and spermatogenic apoptosis. Results: NAE1 exhibited high nuclear expression within spermatocytes from the pachytene stage onwards. Nae1-SKO male mice showed a late-pachytene arrest in spermatocytes, resulting in infertility. In NAE1-deficient spermatocytes, there is an increase in apoptosis. Nae1 deletion led to double-strand break (DSB) repair failure with normal autosomes synapsis. From a mechanistic perspective, we verified excessive recombination intermediate stabilization and failed crossover formation, which ultimately resulted in impaired meiotic recombination. Further analysis showed that ubiquitination regulation coordinated with NAE1-mediated neddylation was implicated in meiotic recombination. Conclusion: NAE1-mediated neddylation regulates ubiquitination during meiosis and is involved in the stabilization of recombination proteins related to crossover differentiation. We provide cytological evidence for the neddylation-ubiquitination system (NUS) in mammalian meiotic recombination during spermatogenesis.

Does the rate of recombination in maternal and paternal genomes of euploid embryos influence the likelihood of pregnancy and a successful live birth in IVF? Family, cycle, and embryo specific recombination rates have no significant correlation with pregnancy outcomes following the transfer of euploid embryos in putative fertile couples. Recombination during meiosis is essential for accurate chromosome segregation and promoting genetic diversity. Previous studies of recombination in human embryos have primarily focused on the relationship with chromosomal aneuploidy, but little is known about how recombination rates on parental genomes in euploid embryos affect pregnancy outcomes following IVF and embryo transfer. Around 50% of euploid embryos have negative pregnancy outcome with no identifiable cause. Given the importance of meiotic recombination in gamete formation and viability of resulting embryos, investigating how meiotic recombination takes place in putative fertile couples with euploid embryos is an important element that is currently lacking. This retrospective multi-centre cohort study analyzes recombination rates in maternal and paternal genomes in 82 couples undergoing preimplantation genetic testing for monogenic disorders. Single nucleotide polymorphism (SNP) array was used to assess recombination events across 165 euploid embryos from a total of 94 IVF cycles performed between 2018 and 2023. Embryological and clinical pregnancy outcomes, including live birth and miscarriage rates, were evaluated following single euploid embryo transfer. An algorithm based on haplotyping using a minimum of two references to phase haplotypes was employed to identify parental haplotype switches using SNP array data.Recombination points were assigned when at least two haplotype switches were identified at the same genomic position by an automated rolling-window approach. Recombination rates were normalized per chromosomal megabase (Mb) and family, cycle, and embryo specific rates were analyzed in relation to pregnancy outcomes and embryo morphology using statistical models. On average 33 (SD:8.84) maternal and 19.8 (SD:5.14) paternal recombination events were detected per embryo. Kendall’s rank correlation revealed no statistical correlation between the number of recombination events (per embryo and per individual chromosome) and maternal age (p-value=0.9; average maternal age:34.5; SD:3.27). Per family analysis revealed an average variation in recombination rates of 7.33 (0-16.9) and 4.29 (0-12.02) in maternal and paternal genomes, respectively. An inter-cycle variation was also observed with an average of 4.9 recombination events for both parental genomes. A per chromosome analysis showed smaller chromosomes have a significantly higher number of recombination events per Mb (Wilcoxon rank; p-value<2.2e-16) while also being more prone to absence of recombination in both parental genomes, with a greater effect for maternal chromosomes likely due to the higher recombination rates. No correlation was evidenced between recombination and blastocyst morphology assigned according to Gardner’s criteria (Kruskal-Wall’s Rank p-value=0.1). Recombination rates in parental genomes of 165 transferred embryos showed no association with pregnancy outcomes, assessed in terms of implantation (maternal p-value=0.8; paternal p-value=0.3), miscarriage (maternal p-value=0.5; paternal p-value=0.8) and live birth (maternal p-value=0.6; paternal p-value=0.4). The sample size is limited, potentially reducing the power to detect smaller effect sizes on pregnancy outcomes. Additionally, this study population consists of putatively fertile individuals undergoing IVF for monogenic disease risk, making the findings on recombination in embryos not directly generalizable to infertile individuals seeking treatment for reproductive failure. This first study examines how recombination patterns influence pregnancy outcomes, exploring their potential role in oocyte and embryo health. While euploid embryos in this fertile cohort showed minimal recombination variability, ongoing research aims to determine its impact on meiotic integrity and reproductive success in infertility cycles. No

Genetic breeding involves the recombination and selection of various valuable genes. Meiotic crossover (CO) promotes the generation of new allelic combinations on chromosomes, which is essential for breeding elite varieties (Wijnker and de Jong, 2008). An increase in CO promotes genetic diversity, whereas a decrease can rapidly stabilize excellent traits (Mercier et al., 2015). Furthermore, the complete elimination of CO facilitates heterotic fixation during apomixis (Wang et al., 2019). However, the number and distribution of COs are tightly restrained in crops, severely hindering agricultural breeding (Crismani et al., 2012). To date, many meiotic genes involved in CO formation have been identified in different organisms. Unfortunately, null mutations in these genes usually cause infertility, thus preventing direct use of the mutants in crop breeding.

Abstract Meiotic crossovers promote correct chromosome segregation and the shuffling of genetic diversity. However, the measurement of crossovers remains challenging, impeding our ability to decipher the molecular mechanisms that are necessary for their formation and regulation. Here we demonstrate a novel repurposing of the single-nucleus Assay for Transposase Accessible Chromatin with sequencing (snATAC-seq) as a simple and high-throughput method to identify and characterize meiotic crossovers from haploid testis nuclei. We first validate the feasibility of obtaining genome-wide coverage from snATAC-seq by using ATAC-seq on bulk haploid mouse testis nuclei, ensuring adequate variant detection for haplotyping. Subsequently, we adapt droplet-based snATAC-seq for crossover detection, revealing >25 000 crossovers in F1 hybrid mice. Comparison between the wild type and a hyper-recombinogenic Fancm-deficient mutant mouse model confirmed an increase in crossover rates in this genotype, however with a distribution which was unchanged. We also find that regions with the highest rate of crossover formation are enriched for PRDM9. Our findings demonstrate the utility of snATAC-seq as a robust and scalable tool for high-throughput crossover detection, offering insights into meiotic crossover dynamics and elucidating the underlying molecular mechanisms. It is possible that the research presented here with snATAC-seq of haploid post-meiotic nuclei could be extended into fertility-related diagnostics.

BackgroundKomagataella phaffii is a yeast widely used in the pharmaceutical and biotechnology industries, and is one of the two species that were previously called Pichia pastoris. However, almost all laboratory work on K. phaffii has utilized strains derived from a single natural isolate, CBS7435. There is little information about the sequence diversity of K. phaffii or the genetic properties of this species.ResultsWe sequenced the genomes of all the known isolates of K. phaffii. We made a genetic cross between derivatives of two isolates that differ at 44,000 single nucleotide polymorphism sites, and used this cross to analyze the rate and landscape of meiotic recombination. We conducted tetrad analysis by making use of the property that K. phaffii haploids do not mate in rich media, which enabled us to isolate and sequence the four types of haploid cell that are present in the colony that forms when a tetra-type ascus germinates.ConclusionsWe found that only four distinct natural isolates of K. phaffii exist in public yeast culture collections. The meiotic recombination rate in K. phaffii is approximately 3.5 times lower than in Saccharomyces cerevisiae, with an average of 25 crossovers per meiosis. Recombination is suppressed, and genetic diversity among natural isolates is low, in a region around centromeres that is much larger than the centromeres themselves. Our work lays a foundation for future quantitative trait locus analysis in K. phaffii.

No abstract available

Meiotic recombination is a conserved mechanism that plays an important role in genetic diversity and genome evolution. By shuffling parental alleles to produce novel haplotype, recombination increases the efficiency of selection at linked loci and thus facilitates population adaptation. During meiosis, DNA double-strand breaks initiates homologous recombination at specific loci called hotspots. In many mammals, their localization corresponds to specific DNA sequences bound by the zinc finger (ZnF) array of PRDM9. These hotspots are also marked with specific epigenetic modifications (H3K4me3 and H3K36me3) that are catalyzed by PRDM9 itself. A remarkable property of PRDM9 is the high diversity and fast evolution of its ZnF domain. The consequence is the fast evolution of recombination map for species having a PRDM9 for hotspot localisation and thus a potentially higher evolvability.PRDM9 appears in the last common ancestor of metazoans but its partial or complete loss have been surprisingly reported in many taxa. Species lacking a full-length PRDM9 have evolutionary stable hotspots that are located near promoter-like regions, that are evicted by PRDM9. The involvement of PRDM9 in meiotic recombination has not been yet explored outside vertebrate species, one reason being that invertebrate model species (drosophila, nematode) have no PRDM9. Here, we propose to fill these gaps by investigating PRDM9 function in meiotic recombination in closely-related species of freshwater snails, including Biomphalaria glabrata and Biomphalaria pfeifferi and Bulinus truncatus, for which we recently identified full-length PRDM9 conservation by exploring their recently published genome assemblies.This project will address the following questions: Where are meiotic hotspots located along the genome of freshwater snails? Is PRDM9 essential for meiotic progress and fertility? Can we find evidence of PRDM9 function in the localisation of hotspots?We will use complementary approaches such as population genomics, genotyping of PRDM9 zinc finger array, molecular mapping of histone modifications and DNA breaks in snail gonads (ChIPseq approaches), histology and genetic manipulation of snail. Preliminary results about PRDM9 ZnF diversity showed hypervariable amino-acid at positions contacting DNA and diversity in number and order of ZnF between individuals and populations. This indicates a probable involvement of PRDM9 in specifying recombination hotspot. The epigenetic profiling is currently being optimized and will be applied to our snails lines already selected for different Prdm9 alleles.This project should help to identify the molecular bases behind the adaptation of freshwater snails to their biotic and abiotic environments.

Wheat is one of the most important crops worldwide, and understanding its genome organisation is crucial for geneticists and breeders. In this study, we examined the dynamic roles of telomeric and subtelomeric regions in wheat, focusing on their influence on homologous chromosome pairing during meiosis, the process that produces gametes. We analysed various Triticum species and modern cultivars, uncovering a complex “barcode” at chromosome ends that rules homologous recognition. Phylogenetic analysis of the ZIP4-5B gene highlighted the evolutionary relationships among wheat species, emphasising the contribution of wild relatives to genetic diversity, especially in terminal chromosomal regions. Our findings suggest that telomeric regions, although traditionally seen as conserved, display significant variability and structural complexity influenced by genetic background and chromosomal context. We observed a strong link between telomere position and variant accumulation, with subtelomeric regions acting as hot spots for instability and chromatin remodelling. G-quadruplex (G4) structures were found to be distributed unevenly, with their density affected by telomere distance and genomic context. Subtelomeric regions were identified as key sites for genetic improvement, harbouring rapidly evolving sequences and transposable elements that may impact meiotic pairing accuracy. Our results indicate that telomeres and subtelomeres serve as dynamic genomic centres, encoding chromosomal identity and regulating homologous pairing through a balance of sequence diversity and structural motifs. This research enhances our understanding of wheat genome stability and provides insights for breeding strategies aimed at increasing genetic diversity. Telomeric and subtelomeric regions in wheat play a dynamic and critical role in meiotic chromosome pairing by acting as genomic “barcodes” that guide homologous recognition. These regions, once considered stable, show high variability and structural complexity influenced by genetic and evolutionary factors, especially through contributions from wild wheat relatives. The findings highlight subtelomeres as hotspots for genetic improvement and genome evolution, offering valuable insights for enhancing wheat breeding and genetic diversity.

Meiosis in males is a critical process that ensures complete spermatogenesis and genetic diversity. However, the key regulators involved in this process and the underlying molecular mechanisms remain unclear. Here, we report an essential role of the m6A methyltransferase METTL16 in meiotic sex chromosome inactivation (MSCI), double‐strand break (DSB) formation, homologous recombination and SYCP1 deposition during male meiosis. METTL16 depletion results in a significantly upregulated transcriptome on sex chromosomes in pachytene spermatocytes and leads to reduced DSB formation and recombination, and increased SYCP1 depositioin during the first wave of spermatogenesis. Mechanistically, in pachytene spermatocytes, METTL16 interacts with MDC1/SCML2 to coordinate DNA damage response (DDR) and XY body epigenetic modifications that establish and maintain MSCI, and in early meiotic prophase I, METTL16 regulates DSB formation and recombination by regulating protein levels of meiosis‐related genes. Furthermore, multi‐omics analyses reveal that METTL16 interacts with translational factors and controls m6A levels in the RNAs of meiosis‐related genes (e.g., Ubr2) to regulate the expression of critical meiotic regulators. Collectively, this study identified METTL16 as a key regulator of male meiosis and demonstrated that it modulates meiosis by interacting with MSCI‐related factors and regulating m6A levels and translational efficiency (TE) of meiosis‐related genes.

Meiotic crossover (CO) formation between homologous chromosomes ensures their subsequent proper segregation and generates genetic diversity among offspring. In maize, however, the mechanisms that modulate crossover formation remain poorly characterized. Here, we found that both maize BREAST CANCER SUSCEPTIBILITY PROTEIN 2 (BRCA2) and AAA-ATPase FIDGETIN-LIKE-1 (FIGL1) act as positive factors of crossover formation by controlling the assembly or/and stability of two conserved DNA recombinases RAD51 and DMC1 filaments. Our results revealed that ZmBRCA2 not only is involved in repair of DNA double-strand breaks (DSBs), but also regulates crossover formation in a dosage-dependent manner. In addition, ZmFIGL1 interacts with RAD51 and DMC1, and Zmfigl1 mutants had a significantly reduced number of RAD51/DMC1 foci and COs. Further, simultaneous loss of ZmFIGL1 and ZmBRCA2 abolished RAD51/DMC1 foci and exacerbated meiotic defects compared with the single mutant Zmbrca2 or Zmfigl1. Together, our data demonstrate that ZmBRCA2 and ZmFIGL1 act coordinately to regulate the dynamics of RAD51/DMC1-dependent DSB repair to promote crossover formation in maize. This conclusion is surprisingly different from the antagonistic roles of BRCA2 and FIGL1 in Arabidopsis, implying that although key factors that control crossover formation are evolutionarily conserved, specific characteristics have been adopted in diverse plant species. This article is protected by copyright. All rights reserved.

Background The 1RS arm of wheat-rye 1BL.1RS translocations contains several subtelomeric tandem repeat families. To study the effect of the difference in the composition of these tandem repeats on the meiotic recombination of 1RS arms can help to enrich the genetic diversity of 1BL.1RS translocation chromosomes. Results Five wheat-rye 1BL.1RS translocation cultivars/lines were used to build two cross combinations including group 1 (20T401 × Zhou 8425B, 20T401 × Lovrin 10 and 20T401 × Chuannong 17) and group 2 (20T360-2 × Zhou 8425B, 20T360-2 × Lovrin 10 and 20T360-2 × Chuannong 17). Oligonucleotide (oligo) probes Oligo-s120.3, Oligo-TR72, and Oligo-119.2-2 produced the same signal pattern on the 1RS arms in lines 20T401 and 20T360-2, and another signal pattern in the three cultivars Zhou 8425B, Lovrin 10 and Chuannong 17. The Oligo-pSc200 signal disappeared from the 1RS arms of the line 20T401, and the signal intensity of this probe on the 1RS arms of the line 20T360-2 was weaker than that of the three cultivars. The five cultivars/lines had the same signal pattern of the probe Oligo-pSc250. The recombination rate of 1RS arms in group 1 was significantly lower than that in group 2. In the progenies from group 1, unequal meiotic recombination in the subtelomeric pSc119.2 and pSc250 tandem repeat regions, and a 1BL.1RS with inversion of 1RS segment between the pSc200 and the nucleolar organizer region were found. Conclusions This study provides a visual tool to detect the meiotic recombination of 1RS arms. The meiotic recombination rate of 1RS arms was affected by the variation of pSc200 tandem repeat, indicating the similar composition of subtelomeric tandem repeats on these arms could increase their recombination rate. These results indicate that the 1RS subtelomeric structure will affect its recombination, and thus the localization of genes on 1RS by means of meiotic recombination might also be affected.

Abstract Recombination is essential for physical attachments and genetic diversity. The Han Chinese population is the largest ethnic group worldwide, therefore, the construction of a genetic map regarding recombination for the population is essential. In this study, 164 and 240 couples who underwent preimplantation genetic testing for monogenic diseases or segmental rearrangement were included in the analysis. Blastocysts and probands from couples who underwent preimplantation genetic testing for monogenic diseases by single nucleotide polymorphism array were included for recombination analysis. The location of recombination was determined from haplotype phase transitions in parent-offspring pairs at loci where the parents were heterozygous. The genetic map for Chinese in vitro fertilization embryos was constructed by the expectation–maximization algorithm with chip-level data. Our results confirmed that homologous recombination occurred more often in maternal chromosomes, and the age effect was more significant in maternal homologous recombination. A total of 6,494 homologous recombination hotspots (32.3%) were identified in genes of Online Mendelian Inheritance in Man. A uniform association between homologous recombination and aneuploidy was not established. In addition, carriers with identified breakpoints of reciprocal translocations were analyzed, and locations of breakpoints were found partly overlapped with homologous recombination hotspots, implying a possible similar mechanism behind both events. This study highlights the significance of constructing a recombination map, which may improve the accuracy of haplotype analysis for preimplantation genetic testing for monogenic diseases. Overlapping locations of translocation and recombination are worthy of further investigation.

Abstract Meiotic recombination ensures accurate chromosome segregation and genetic diversity during gametogenesis, and its disruption leads to infertility. The dual histone methylation writer–reader system, in which PRDM9 deposits H3K4me3 and H3K36me3 marks at nucleosomes to define recombination hotspots and ZCWPW1 acts as a reader recognizing these marks, is essential for meiotic recombination. However, the regulatory mechanisms of this system remain unclear. Here, we showed that deficiency of ZCWPW2 causes recombination defects in humans and mice, including impaired homologous chromosome synapsis and defective DNA double-strand break repair. CUT&Tag analysis revealed that ZCWPW2 exhibits increased enrichment at dual H3K4me3 and H3K36me3 sites in the presence of PRDM9, while binding to promoter regions independently of PRDM9 to regulate meiotic transcription. Mass spectrometry further showed that ZCWPW2 forms a complex with ZCWPW1 and interacts with recombination-associated proteins in a ZCWPW1-dependent manner. Mechanistically, we demonstrate that the ZCWPW1–ZCWPW2 complex enhances the functions of key lactylation regulators LDHA and EP300, thereby promoting lactylation of recombination-associated proteins and stabilizing their abundance. Collectively, we identify ZCWPW2 as a previously unrecognized but essential factor in meiotic recombination, elucidate the molecular mechanism of the PRDM9/ZCWPW1/ZCWPW2 system in regulating recombination, and uncover a critical role for lactylation in meiosis.

Meiotic recombination ensures genetic diversity and accurate chromosome segregation by mediating reciprocal DNA exchange between homologous chromosomes. In this process, the meiosis-specific recombinase DMC1 plays a pivotal role in homology search and pairing, but the molecular mechanisms underlying its function remain unclear. Using single-molecule imaging, we demonstrate that the human DMC1-ssDNA presynaptic complex employs a diffusion-based mechanism to search for homologous DNA. Although this diffusing complex generates a migrating DNA "bubble," it cannot align with the homologous sequence in the absence of free DMC1 protein. Strikingly, the meiosis-specific cofactor complex HOP2-MND1 compensates for the lack of free DMC1 and enables homology recognition. Notably, HOP2-MND1 achieves this by codiffusing with the presynaptic complex, acting to clamp the ssDNA-dsDNA junctions and maintain an expanded DNA bubble conducive to sequence alignment. Our findings identify DMC1 together with HOP2-MND1 as a functional homology search unit and provide mechanistic insights into how auxiliary factors regulate DMC1-driven strand exchange during meiotic recombination.

Meiotic recombination through chromosomal crossing-over is a fundamental feature of sex and an important driver of genomic diversity. It ensures proper disjunction, allows increased selection responses, and prevents mutation accumulation; however, it is also mutagenic and can break up favourable haplotypes. This cost/benefit dynamic is likely to vary depending on mechanistic and evolutionary contexts, and indeed, recombination rates show huge variation in nature. Identifying the genetic architecture of this variation is key to understanding its causes and consequences. Here, we investigate individual recombination rate variation in wild house sparrows (Passer domesticus). We integrate genomic and pedigree data to identify autosomal crossover counts (ACC) and intra-chromosomal allelic shuffling (r̅intra) in 13,056 gametes. Females had 1.37 times higher ACC, and 1.55 times higher r̅intra than males. ACC and r̅intra were heritable in females and males (ACC h2 = 0.23 and 0.11; r̅intra h2 = 0.12 and 0.14), but cross-sex additive genetic correlations were low (rA = 0.29 and 0.32 for ACC and r̅intra). Conditional bivariate analyses showed that all measures remained heritable after accounting for genetic values in the opposite sex, indicating that sex-specific ACC and r̅intra can evolve somewhat independently. Genome-wide models showed that ACC and r̅intra are polygenic and driven by many small-effect loci, many of which are likely to act in trans as global recombination modifiers. Our findings show that recombination rates of females and males can have different evolutionary potential in wild birds, providing a compelling mechanism for the evolution of sexual dimorphism in recombination.

The allopolyploid okra (Abelmoschus esculentus) unveiled telomeric repeats flanking distal gene-rich regions and short interstitial TTTAGGG telomeric repeats, possibly representing hallmarks of chromosomal speciation. Ribosomal RNA (rRNA) genes organize into 5S clusters, distinct from the 18S-5.8S-28S units, indicating an S-type rRNA gene arrangement. The assembly, in line with cytogenetic and cytometry observations, identifies 65 chromosomes and a 1.45 Gb genome size estimate in a haploid sibling. The lack of aberrant meiotic configurations implies limited to no recombination among sub-genomes. k-mer distribution analysis reveals 75% has a diploid nature and 15% heterozygosity. The configurations of Benchmarking Universal Single-Copy Ortholog (BUSCO), k-mer, and repeat clustering point to the presence of at least two sub-genomes one with 30 and the other with 35 chromosomes, indicating the allopolyploid nature of the okra genome. Over 130 000 putative genes, derived from mapped IsoSeq data and transcriptome data from public okra accessions, exhibit a low genetic diversity of one single nucleotide polymorphisms per 2.1 kbp. The genes are predominantly located at the distal chromosome ends, declining toward central scaffold domains. Long terminal repeat retrotransposons prevail in central domains, consistent with the observed pericentromeric heterochromatin and distal euchromatin. Disparities in paralogous gene counts suggest potential sub-genome differentiation implying possible sub-genome dominance. Amino acid query sequences of putative genes facilitated phenol biosynthesis pathway annotation. Comparison with manually curated reference KEGG pathways from related Malvaceae species reveals the genetic basis for putative enzyme coding genes that likely enable metabolic reactions involved in the biosynthesis of dietary and therapeutic compounds in okra.

Meiosis is essential for evolution and genetic diversity in almost all sexual eukaryotic organisms. The mechanisms of meiotic recombination, such as synapsis, have been extensively investigated. However, it is still unclear whether signals from the cytoplasm or even outside of the cell can regulate the meiosis process. Cilia are microtubule-based structures that protrude from cell surface and function as signaling hubs to sense extracellular signals. Here, we reported an unexpected and critical role of cilia during meiotic recombination. During gametogenesis of zebrafish, cilia were specifically present in the prophase stages of both spermatocytes and primary oocytes. By developing a novel germ cell-specific CRISPR/Cas9 system, we demonstrated that germ cell-specific depletion of ciliary genes resulted in compromised double strand break repair, reduced crossover formation, and increased germ cell apoptosis. Our study reveals a previously undiscovered role for cilia during meiosis and suggests that extracellular signals may regulate meiotic recombination via this particular organelle.

How do the number and location of meiotic crossovers contribute to the formation of aneuploidies observed in preimplantation human embryos? Normalized across chromosomes, trisomies possess 35% fewer crossovers on average compared to disomies, while the genomic distribution of crossovers is also substantially altered. Meiotic recombination is a crucial source of genetic diversity and is also critical for ensuring the accuracy of chromosome segregation. Understanding the landscape of meiotic recombination, its variation across individuals, and the processes by which it goes awry are long-standing goals in human genetics. Current approaches for inferring the landscape of recombination either rely on population genetic patterns of linkage disequilibrium—capturing a time-averaged view—or direct detection of crossovers in gametes or multi-generation pedigrees, limiting the scale and availability of relevant datasets. Moreover, most of these methods are designed for discovering recombination using data from normal, disomic chromosomes. We present a method for mapping sex-specific recombination landscapes from low-coverage (<0.1×) data from preimplantation genetic testing for aneuploidy (PGT-A) of embryos with arbitrary ploidy configurations. To overcome the sparsity of these data, our method exploits its inherent relatedness structure, knowledge of haplotypes from external population reference panels, as well as the frequent occurrence of chromosome loss in embryos, whereby the remaining chromosome is phased by default. We benchmarked our method by simulating crossovers between known haplotypes. Encouraged by the performance on simulated data we extended our study to retrospective analysis utilizing de-identified PGT-A data obtained between April 2021 and August 2022 at the CReATe Fertility Centre (Toronto, Canada). The data include 20,160 embryos (2,559 IVF patients) with an average depth of coverage of ∼0.05×, facilitating the mapping of crossovers at an average resolution of ∼150 kbp. Our benchmarking results demonstrate high sensitivity and specificity across all ancestries at a coverage of 0.05x per homolog (AUC = 0.989), with AUC declining by 0.014 and 0.053 for when coverage is reduced to 0.025x and 0.013x, respectively. Extending our analysis to real PGT-A data, we observed that our inferred sex-specific landscapes of meiotic crossovers on disomic chromosomes were strongly correlated with published genetic maps from studies based on high-coverage sequencing of parent-offspring trios (r = 0.86 for female map; r = 0.53 for male map), broadly supporting the accuracy of our method. Notably, the total length of the female genetic map was reduced by 35% for trisomies compared to disomies, consistent with the hypothesized role of reduced crossovers and exchangeless chromosomes in the origins of female meiotic aneuploidy. In addition, the genomic distribution of crossovers is also altered in a chromosome-specific manner. Examples include a reduction in crossovers near the centromere of trisomies versus disomies of chromosome 16, as well as an enrichment of crossovers on the q-arm of trisomies versus disomies of chromosome 22. Together, our results provide a detailed corroboration of the hypothesis that aberrant meiotic recombination contributes to the origins of aneuploidies. The accuracy of our method is influenced by genomic heterogeneity in depth of coverage, rates of heterozygosity, and mismatches between the ancestry of the reference panel and the tested sequence. Moreover, technical errors such as spurious alignment and genotyping could hinder analysis in repetitive genomic regions. Together, our study helps clarify the dual function of meiotic recombination in generating genetic diversity while ensuring meiotic fidelity. Our method for patient-specific mapping of meiotic recombination phenotypes may offer clues about how dysregulation of this process contributes to infertility. NIH r R35GM13374

Meiosis is an essential component of the sexual life cycle in eukaryotes. The independent assortment of chromosomes in meiosis increases genetic diversity at the level of whole chromosomes and meiotic recombination increases genetic diversity within chromosomes. The resulting variability fuels evolution. Interestingly, global mapping of recombination in diverse taxa revealed dramatic changes in its frequency distribution between closely related species, subspecies, and even isolated populations of the same species. New insight into mechanisms for these evolutionarily rapid changes has come from analyses of environmentally induced plasticity of recombination in fission yeast. Many different DNA sites, and where identified their binding/activator proteins, control the positioning of recombination at hotspots. Each different class of hotspots functions as an independently controlled rheostat that modulates rates of recombination over a broad dynamic range in response to changing conditions. Together, this independent modulation can rapidly and dramatically alter the global frequency distribution of recombination. This process likely contributes substantially to (i.e., can largely explain) evolutionarily rapid, Prdm9-independent changes in the recombination landscape. Moreover, the precise control mechanisms allow cells to dynamically favor or disfavor newly arising combinations of linked alleles in response to changing extracellular and intracellular conditions, which has striking implications for the impacts of meiotic recombination on evolution.

Simple Summary The selection of varieties more resilient to disease and climate change requires generating new genetic diversity for breeding. The main mechanism for reshuffling genetic information is through the recombination of chromosomes during meiosis. We showed in oilseed rape (Brassica napus, AACC, 2n = 4x = 38), which is a natural hybrid formed from a cross between turnip (B. rapa, AA, 2n = 2x = 20) and cabbage (B. oleracea, CC, 2n = 2x = 18), that there is significantly more crossovers occurring along the entire A chromosomes in allotriploid AAC (crossbetween B. napus and B. rapa) than in diploid AA or allotetraploid AACC hybrids. We demonstrated that these allotriploid AAC hybrids are highly efficient to introduce new variability within oilseed rape varieties, notably by enabling the introduction of small genomic regions carrying genes controlling agronomically interesting traits. Abstract Meiotic recombination is the main tool used by breeders to generate biodiversity, allowing genetic reshuffling at each generation. It enables the accumulation of favorable alleles while purging deleterious mutations. However, this mechanism is highly regulated with the formation of one to rarely more than three crossovers, which are not randomly distributed. In this study, we showed that it is possible to modify these controls in oilseed rape (Brassica napus, AACC, 2n = 4x = 38) and that it is linked to AAC allotriploidy and not to polyploidy per se. To that purpose, we compared the frequency and the distribution of crossovers along A chromosomes from hybrids carrying exactly the same A nucleotide sequence, but presenting three different ploidy levels: AA, AAC and AACC. Genetic maps established with 202 SNPs anchored on reference genomes revealed that the crossover rate is 3.6-fold higher in the AAC allotriploid hybrids compared to AA and AACC hybrids. Using a higher SNP density, we demonstrated that smaller and numerous introgressions of B. rapa were present in AAC hybrids compared to AACC allotetraploid hybrids, with 7.6 Mb vs. 16.9 Mb on average and 21 B. rapa regions per plant vs. nine regions, respectively. Therefore, this boost of recombination is highly efficient to reduce the size of QTL carried in cold regions of the oilseed rape genome, as exemplified here for a QTL conferring blackleg resistance.

Abstract During meiosis, homologous chromosomes pair and recombine, which can result in reciprocal crossovers that increase genetic diversity. Crossovers are unevenly distributed along eukaryote chromosomes and show repression in heterochromatin and the centromeres. Within the chromosome arms, crossovers are often concentrated in hotspots, which are typically in the kilobase range. The uneven distribution of crossovers along chromosomes, together with their low number per meiosis, creates a limitation during crop breeding, where recombination can be beneficial. Therefore, targeting crossovers to specific genome locations has the potential to accelerate crop improvement. In plants, meiotic crossovers are initiated by DNA double-strand breaks that are catalyzed by SPO11 complexes, which consist of 2 catalytic (SPO11-1 and SPO11-2) and 2 noncatalytic subunits (MTOPVIB). We used the model plant Arabidopsis thaliana to coexpress an MTOPVIB-dCas9 fusion protein with guide RNAs specific to the 3a crossover hotspot. We observed that this was insufficient to significantly change meiotic crossover frequency or pattern within 3a. We discuss the implications of our findings for targeting meiotic recombination within plant genomes.

Meiotic recombination is a critical process for sexually reproducing organisms. This exchange of genetic information between homologous chromosomes during meiosis is important not only because it generates genetic diversity, but also because it is often required for proper chromosome segregation. Consequently, the frequency and distribution of crossovers are tightly controlled to ensure fertility and offspring viability. However, in many systems it has been shown that environmental factors can alter the frequency of crossover events. Two studies in flies and yeast point to nutritional status affecting the frequency of crossing over. However, this question remains unexplored in mammals. Here we test how crossover frequency varies in response to diet in Mus musculus males. We use immunohistochemistry to estimate crossover frequency in multiple genotypes under two diet treatments. Our results indicate that while crossover frequency was unaffected by diet in some strains, other strains were sensitive even to small composition changes between two common laboratory chows. Therefore, recombination is both resistant and sensitive to certain dietary changes in a strain-dependent manner and, hence, this response is genetically determined. Our study is the first to report a nutrition effect on genome-wide levels of recombination. Moreover, our work highlights the importance of controlling diet in recombination studies and may point to diet as a potential source of variability among studies, which is relevant for reproducibility.

Human recombination and mutation mapped Genetic recombination is an essential process in generating genetic diversity. Recombination occurs both through the shuffling of maternal and paternal chromosomes and through mutations generated by resolution of the physical breaks necessary for this process. Halldorsson et al. sequenced the full genomes of parents and offspring to create a map of human recombination and estimate the relationship with de novo mutations. Interestingly, transcribed regions of the genome were less likely to have crossovers, suggesting that there may be selection to reduce changes in genetic sequences via recombination or mutation in these regions. Science, this issue p. eaau1043 A high-resolution human genetic map from whole genomes identifies patterns of recombination and de novo mutation. INTRODUCTION Diversity in the sequence of the human genome, arising from recombinations and mutations, is fundamental to human evolution and human diversity. Meiotic recombination is initiated from double-strand breaks (DSBs). DSBs occur more frequently in regions of the genome termed hotspots, and a small subset eventually gives rise to crossovers, a reciprocal exchange of large pieces between homologous chromosomes. The majority of DSBs do not lead to crossovers but end as localized transfers of short segments between homologous chromosomes or sister chromatids, observable as gene conversions when the segment includes a heterozygous marker. Crossovers co-occurring with distal gene conversions are known as complex crossovers. RATIONALE Current meiotic recombination maps either have limited resolution or the events cannot be resolved to an individual level. The detection of recombination and de novo mutations (DNMs) requires genetic data on a proband and its parents, and a fine resolution of these events is possible only with whole-genome sequence data. Whole-genome sequencing and DNA microarray data allowed us to identify crossovers and DNMs in families at a high resolution. We resolved crossovers at an individual level, allowing us to examine variation in crossover patterns between individuals, analyzing which crossovers are complex and how crossover patterns are influenced by age, sex, sequence variants, and epigenomic factors. It is known that the mutation rate is increased near crossovers, but the rate of DNMs near crossovers has been characterized only indirectly or at a small scale. RESULTS We show that a number of epigenomic factors influence crossover location, shifting crossovers from exons to enhancers. Complex crossovers are more common in females than males, and the rate of complex crossovers increases with maternal age. Maternal age also correlates with an increase in the recombination rate in general and a shift in the location of crossovers toward later-replicating regions and regions of lower GC content. Both sexes show an ~50-fold increase in DNMs within 1 kb of crossovers, but the types of DNMs differ considerably between the sexes. Females, but not males, also exhibit an increase in the mutation rate up to 40 kb from crossovers, particularly at complex crossovers. We found 47 variants at 35 loci affecting the recombination rate and/or the location of crossover, 24 of which are coding or splice region variants. Whereas some of the variants affect both the recombination rate and several measures of crossover location in both sexes, other variants affect only one of these measures in one of the sexes. Many of these variants are in genes that encode the synaptonemal complex. CONCLUSION Our genome-wide recombination map provides a resolution of 682 base pairs. We show that crossovers have a direct mutagenic effect and demonstrate that DNMs and crossovers accumulate in the same regions with advancing maternal age. Furthermore, our results illustrate extensive genetic control of meiotic recombinations and highlight genes linked to the formation of the synaptonemal complex as determinants of crossovers. Our search for crossovers in parents and their offspring. Histone modifications influence crossover location. The DNM rate is higher within 1 kb from a crossover in both sexes, but the type of mutations differs between the sexes. The DNM rate is also higher up to 40 kb from crossovers in females with enrichment of G→C mutations. We used crossovers from many individuals to construct genetic maps and performed genome-wide association studies (GWAS) on the recombination rate and attributes of crossover locations to search for genes that control crossover characteristics. Genetic diversity arises from recombination and de novo mutation (DNM). Using a combination of microarray genotype and whole-genome sequence data on parent-child pairs, we identified 4,531,535 crossover recombinations and 200,435 DNMs. The resulting genetic map has a resolution of 682 base pairs. Crossovers exhibit a mutagenic effect, with overrepresentation of DNMs within 1 kilobase of crossovers in males and females. In females, a higher mutation rate is observed up to 40 kilobases from crossovers, particularly for complex crossovers, which increase with maternal age. We identified 35 loci associated with the recombination rate or the location of crossovers, demonstrating extensive genetic control of meiotic recombination, and our results highlight genes linked to the formation of the synaptonemal complex as determinants of crossovers.

Changes in environmental temperature influence cellular processes and their dynamics, and thus affect the life cycle of organisms that are unable to control their cell/body temperature. Meiotic recombination is the cellular process essential for producing healthy haploid gametes by providing physical links (chiasmata) between homologous chromosomes to guide their accurate segregation. Additionally, meiotic recombination—initiated by programmed DNA double-strand breaks (DSBs)—can generate genetic diversity and, therefore, is a driving force of evolution. Environmental temperature influencing meiotic recombination outcome thus may be a crucial determinant of reproductive success and genetic diversity. Indeed, meiotic recombination frequency in fungi, plants and invertebrates changes with temperature. In most organisms, these temperature-induced changes in meiotic recombination seem to be mediated through the meiosis-specific chromosome axis organization, the synaptonemal complex in particular. The fission yeast Schizosaccharomyces pombe does not possess a synaptonemal complex. Thus, we tested how environmental temperature modulates meiotic recombination frequency in the absence of a fully-fledged synaptonemal complex. We show that intragenic recombination (gene conversion) positively correlates with temperature within a certain range, especially at meiotic recombination hotspots. In contrast, crossover recombination, which manifests itself as chiasmata, is less affected. Based on our observations, we suggest that, in addition to changes in DSB frequency, DSB processing could be another temperature-sensitive step causing temperature-induced recombination rate alterations.

Genome wide screening of pooled pollen samples from a single interspecific F1 hybrid obtained from a cross between tomato, Solanum lycopersicum and its wild relative, Solanum pimpinellifolium using linked read sequencing of the haploid nuclei, allowed profiling of the crossover (CO) and gene conversion (GC) landscape. We observed a striking overlap between cold regions of CO in the male gametes and our previously established F6 recombinant inbred lines (RILs) population. COs were overrepresented in non-coding regions in the gene promoter and 5'UTR regions of genes. Poly-A/T and AT rich motifs were found enriched in 1 kb promoter regions flanking the CO sites. Non-crossover associated allelic and ectopic GCs were detected in most chromosomes, confirming that besides CO, GC represents also a source for genetic diversity and genome plasticity in tomato. Furthermore, we identified processed break junctions pointing at the involvement of both homology directed and non-homology directed repair pathways, suggesting a recombination machinery in tomato that is more complex than currently anticipated.

Centromeric DNA and proteins evolve rapidly despite conserved function in mediating kinetochore-microtubule attachments during cell division. This paradox is explained by selfish DNA sequences preferentially binding centromeric proteins to disrupt attachments and bias their segregation into the egg (drive) during female meiosis. Adaptive centromeric protein evolution is predicted to prevent preferential binding to these sequences and suppress drive. Here, we test this prediction by defining the impact of adaptive evolution of the DNA-binding histone fold domain of CENP-T, a major link between centromeric DNA and microtubules. We reversed adaptive changes by creating chimeric variants of mouse CENP-T with the histone fold domain from closely related species, expressed exogenously in mouse oocytes or in a transgenic mouse model. We show that adaptive evolution of mouse CENP-T reduced centromere binding, which supports robust female gametogenesis. However, this innovation is independent of the centromeric DNA sequence, as shown by comparing the binding of divergent CENP-T variants to distinct centromere satellite arrays in mouse oocytes and in somatic cells from other species. Overall, our findings support a model in which selfish sequences drive to fixation, disrupting attachments of all centromeres to the spindle. DNA sequence-specific innovations are not needed to mitigate fitness costs in this model, so centromeric proteins adapt by modulating their binding to all centromeres in the aftermath of drive.

Genome duplication, which leads to polyploidy, poses challenges to the meiotic segregation of the now-multiple homologous chromosome copies. Genome scan data showed previously that adaptation to polyploid meiosis in autotetraploid Arabidopsis arenosa is likely multigenic, involving genes encoding interacting proteins. But what does this really mean? Functional follow-up studies to genome scans for multigenic traits remain rare in most systems, and thus many mysteries remain about the “functional architecture” of polygenic adaptations. Do different genes all contribute subtle and additive progression towards a fitness optimum, or are there more complex interactions? We previously showed that derived alleles of genes encoding two interacting meiotic axis proteins (ASY1 and ASY3) have additive functional consequences for meiotic adaptation. Here we study derived versus ancestral alleles of the meiotic cohesin subunit REC8, which has roles in chromatin condensation, recruiting the axes, and other critical functions in meiosis. We use genetic and cytological approaches to assess the functional effects of REC8 diploid versus tetraploid alleles, as well as their interaction with ancestral versus derived alleles of ASY1 and ASY3. We show that homozygotes for derived (tetraploid) REC8 alleles have significantly fewer unpaired univalents, a common problem in neotetraploids. Interactions with ASY1 and ASY3 are complex, with the genes in some cases affecting distinct traits, and additive or even antagonistic effects on others. These findings suggest that the road to meiotic adaptation in A. arenosa was perhaps neither straight nor smooth.

The human-directed, global selection for glyphosate resistance in weeds has revealed a fascinating diversity of evolved resistance mechanisms, including herbicide sequestration in the vacuole, a rapid cell death response, nucleotide polymorphisms in the herbicide target (5-enolpyruvylshikimate-3-phosphate synthase, EPSPS) and increased gene copy number of EPSPS. For this latter mechanism, two distinct molecular genetic mechanisms have been observed, a tandem duplication mechanism and a large extrachromosomal circular DNA (eccDNA) that is tethered to the chromosomes and passed to gametes at meiosis. These divergent mechanisms have a range of consequences for the spread, fitness, and inheritance of resistance traits, and, particularly in the case of the eccDNA, demonstrate how evolved herbicide resistance can generate new insights into plant adaptation to contemporary environmental stress.

Hybridization is increasingly recognized as an important force impacting adaptation and evolution in many lineages of fungi. During hybridization, divergent genomes and alleles are brought together into the same cell, potentiating adaptation by increasing genomic plasticity. Here, we review hybridization in fungi by focusing on two fungal pathogens of animals. Hybridization is common between the basidiomycete yeast species Cryptococcus neoformans × Cryptococcus deneoformans, and hybrid genotypes are frequently found in both environmental and clinical settings. The two species show 10–15% nucleotide divergence at the genome level, and their hybrids are highly heterozygous. Though largely sterile and unable to mate, these hybrids can propagate asexually and generate diverse genotypes by nondisjunction, aberrant meiosis, mitotic recombination, and gene conversion. Under stress conditions, the rate of such genetic changes can increase, leading to rapid adaptation. Conversely, in hybrids formed between lineages of the chytridiomycete frog pathogen Batrachochytrium dendrobatidis (Bd), the parental genotypes are considerably less diverged (0.2% divergent). Bd hybrids are formed from crosses between lineages that rarely undergo sex. A common theme in both species is that hybrids show genome plasticity via aneuploidy or loss of heterozygosity and leverage these mechanisms as a rapid way to generate genotypic/phenotypic diversity. Some hybrids show greater fitness and survival in both virulence and virulence-associated phenotypes than parental lineages under certain conditions. These studies showcase how experimentation in model species such as Cryptococcus can be a powerful tool in elucidating the genotypic and phenotypic consequences of hybridization.

Symbiosis between dinoflagellates of the genus Symbiodinium and reef-building corals forms the trophic foundation of the world’s coral reef ecosystems. Here we present the first draft genome of Symbiodinium goreaui (Clade C, type C1: 1.03 Gbp), one of the most ubiquitous endosymbionts associated with corals, and an improved draft genome of Symbiodinium kawagutii (Clade F, strain CS-156: 1.05 Gbp) to further elucidate genomic signatures of this symbiosis. Comparative analysis of four available Symbiodinium genomes against other dinoflagellate genomes led to the identification of 2460 nuclear gene families (containing 5% of Symbiodinium genes) that show evidence of positive selection, including genes involved in photosynthesis, transmembrane ion transport, synthesis and modification of amino acids and glycoproteins, and stress response. Further, we identify extensive sets of genes for meiosis and response to light stress. These draft genomes provide a foundational resource for advancing our understanding of Symbiodinium biology and the coral-algal symbiosis. Huanle Liu et al. report draft genomes of two Symbiodinium species, one from the most dominant type of symbionts in reef-building corals. They find evidence of positive selection in genes related to stress response, meiosis and other traits required for forming successful symbiotic relationships.

Killer meiotic drivers are genetic parasites that destroy ‘sibling’ gametes lacking the driver allele. The fitness costs of drive can lead to selection of unlinked suppressors. This suppression could involve evolutionary tradeoffs that compromise gametogenesis and contribute to infertility. Schizosaccharomyces pombe, an organism containing numerous gamete (spore)-killing wtf drivers, offers a tractable system to test this hypothesis. Here, we demonstrate that in scenarios analogous to outcrossing, wtf drivers generate a fitness landscape in which atypical spores, such as aneuploids and diploids, are advantageous. In this context, wtf drivers can decrease the fitness costs of mutations that disrupt meiotic fidelity and, in some circumstances, can even make such mutations beneficial. Moreover, we find that S. pombe isolates vary greatly in their ability to make haploid spores, with some isolates generating up to 46% aneuploid or diploid spores. This work empirically demonstrates the potential for meiotic drivers to shape the evolution of gametogenesis.

Inverted meiosis, in which sister chromatids segregate before homologous chromosomes, is a common aberration of conventional meiosis (in which sister chromatids segregate after homologous chromosomes) and is routinely observed in certain species. This raises an evolutionary mystery: what is the adaptive advantage of the more common, conventional order of segregation in meiosis? I use a population genetic model to show that asexual mutants arising from inverted meiosis are relatively immune from the deleterious effects of loss of complementation (heterozygosity), unlike the asexual mutants arising from conventional meiosis, in which loss of complementation can outweigh the two‐fold cost of meiosis. Hence, asexual reproduction can replace sexual reproduction with inverted meiosis, but not with conventional meiosis. The results are in line with analogous considerations on other alternative types of reproduction and support the idea that amphimixis is stable in spite of the two‐fold cost of meiosis because loss of complementation in mutant asexuals outweigh the two‐fold cost.

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Species harbor extensive structural variation underpinning recent adaptive evolution. However, the causality between genomic features and the induction of new rearrangements is poorly established. Here, we analyze a global set of telomere-to-telomere genome assemblies of a fungal pathogen of wheat to establish a nucleotide-level map of structural variation. We show that the recent emergence of pesticide resistance has been disproportionally driven by rearrangements. We use machine learning to train a model on structural variation events based on 30 chromosomal sequence features. We show that base composition and gene density are the major determinants of structural variation. Retrotransposons explain most inversion, indel and duplication events. We apply our model to Arabidopsis thaliana and show that our approach extends to more complex genomes. Finally, we analyze complete genomes of haploid offspring in a four-generation pedigree. Meiotic crossover locations are enriched for new rearrangements consistent with crossovers being mutational hotspots. The model trained on species-wide structural variation accurately predicts the position of >74% of newly generated variants along the pedigree. The predictive power highlights causality between specific sequence features and the induction of chromosomal rearrangements. Our work demonstrates that training sequence-derived models can accurately identify regions of intrinsic DNA instability in eukaryotic genomes. Structural variation in genomes of the same species is frequent but what drives the rearrangements remains unclear. Machine-learning of rearrangement patterns among telomere-to-telomere assemblies can accurately identify regions of intrinsic DNA instability in a eukaryotic pathogen.

The adaptive value of sexual reproduction is still debated. A short‐term disadvantage of asexual reproduction is loss of heterozygosity, which leads to the unmasking of recessive deleterious mutations. The cost of this loss of complementation is predicted to be higher than the twofold cost of meiosis for most types of asexual reproduction. Automixis with terminal fusion of sister nuclei is especially vulnerable to the effect of loss of complementation. It is found, however, in some taxa including oribatid mites, the most prominent group of ancient asexuals. Here, I show that automixis with terminal fusion is stable if it is associated with inverted meiosis and that this appears to be the case in nature, notably in oribatid mites. The existence of automixis with terminal fusion, and its co‐occurrence with inverted meiosis, therefore, is consistent with the hypothesis that loss of complementation is important in the evolution of sexual reproduction.

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Male meiosis in two species of Saldula, S. orthothila and S. saltatoria was studied. The diploid chromosome numbers were found to be 2n=32+m+XY and 2n=32+2m+XO respectively. The existence of a Y-chromosome was recorded for the first time in this primitive heteropteran genus. The meiotic behaviour of the sex chromosomes was found to be similar to that of sex chromosomes in more advanced Heteroptera. Autosomal bivalents during late meiotic prophase and metaphase I consisted of two parallelly aligned chromosomes. No traces of chiasmata could be found. The role of Y-chromosome in the evolution of sex chromosome systems in Heteroptera, the mechanisms behind the regular segregation of achiasmatic chromosomes, the adaptive significance of achiasmatic meiosis and the restriction of achiasmatic meiosis to the heterogametic sex are discussed.

Selfish centromere DNA sequences bias their transmission to the egg in female meiosis. Evolutionary theory suggests that centromere proteins evolve to suppress costs of this "centromere drive." In hybrid mouse models with genetically different maternal and paternal centromeres, selfish centromere DNA exploits a kinetochore pathway to recruit microtubule-destabilizing proteins that act as drive effectors. We show that such functional differences are suppressed by a parallel pathway for effector recruitment by heterochromatin, which is similar between centromeres in this system. Disrupting the kinetochore pathway with a divergent allele of CENP-C reduces functional differences between centromeres, whereas disrupting heterochromatin by CENP-B deletion amplifies the differences. Molecular evolution analyses using Murinae genomes identify adaptive evolution in proteins in both pathways. We propose that centromere proteins have recurrently evolved to minimize the kinetochore pathway, which is exploited by selfish DNA, relative to the heterochromatin pathway that equalizes centromeres, while maintaining essential functions.

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Hotspots of rapid genome evolution hold clues about human adaptation. Here, we present a comparative analysis of nine whole-genome sequenced primates to identify high-confidence targets of positive selection. We find strong statistical evidence for positive selection acting on 331 protein-coding genes (3%), pinpointing 934 adaptively evolving codons (0.014%). Our stringent procedure and quality control of alignments and evolutionary inferences reveal substantial artefacts (20% of initial predictions) that have inflated previous estimates of positive selection, the large majority relating to transcript definitions (61%) or gene models (38%). Our final set of 331 positively selected genes (PSG) are strongly enriched for innate and adaptive immune functions, secreted and cell membrane proteins (e.g. pattern recognition, complement, cytokine pathways, defensins, immune receptors, MHC, Siglecs). We also find evidence for positive selection in reproduction, chromosome segregation and meiosis (e.g. centromere-associated CENPO, CENPT), apolipoproteins, smell/taste receptors, and proteins interacting with mitochondrial-encoded molecules. Focusing on the virus-host interaction, we retrieve most evolutionary conflicts known to influence antiviral activity (e.g. TRIM5, MAVS, SAMHD1, tetherin) and predict 70 novel cases through integration with virus-host interaction data (virus-human PPIs, immune cell expression, infection screens). Protein structure analysis identifies positive selection in the interaction interfaces between viruses and their human cellular receptors (CD4 – HIV; CD46 [MCP] – measles, adenoviruses; CD55 [DAF] – picornaviruses). Finally, the primate PSG consistently show high sequence variation in human exomes, suggesting ongoing evolution. Our curated dataset of positively selected genes and positions, available at http://www.cmbi.umcn.nl/∼rvdlee/positive_selection/, is a rich source for studying the genetics underlying human (antiviral) phenotypes.

Dinoflagellates of the Symbiodiniaceae family encompass diverse symbionts that are critical to corals and other species living in coral reefs. It is well known that sexual reproduction enhances adaptive evolution in changing environments. Although genes related to meiotic functions were reported in Symbiodiniaceae, cytological evidence of meiosis and fertilisation are however yet to be observed in these taxa. Using transcriptome and genome data from 21 Symbiodiniaceae isolates, we studied genes that encode proteins associated with distinct stages of meiosis and syngamy. We report the absence of genes that encode main components of the synaptonemal complex (SC), a protein structure that mediates homologous chromosomal pairing and class I crossovers. This result suggests an independent loss of canonical SCs in the alveolates, that also includes the SC-lacking ciliates. We hypothesise that this loss was due in part to permanently condensed chromosomes and repeat-rich sequences in Symbiodiniaceae (and other dinoflagellates) which favoured the SC-independent class II crossover pathway. Our results reveal novel insights into evolution of the meiotic molecular machinery in the ecologically important Symbiodiniaceae and in other eukaryotes.

Centromere drive model describes an evolutionary process initiated by centromeric repeats expansion, which leads to the recruitment of excess kinetochore proteins and consequent preferential segregation of an expanded centromere to the egg during female asymmetric meiosis. In response to these selfish centromeres, the histone protein CenH3, which recruits kinetochore components, adaptively evolves to restore chromosomal parity and counter the detrimental effects of centromere drive. Holocentric chromosomes, whose kinetochores are assembled along entire chromosomes, have been hypothesized to prevent expanded centromeres from acquiring a selective advantage and initiating centromere drive. In such a case, CenH3 would be subjected to less frequent or no adaptive evolution. Using codon substitution models, we analyzed 36 CenH3 sequences from 35 species of the holocentric family Cyperaceae. We found 10 positively selected codons in the CenH3 gene [six codons in the N-terminus and four in the histone fold domain (HFD)] and six branches of its phylogeny along which the positive selection occurred. One of the positively selected codons was found in the centromere targeting domain (CATD) that directly interacts with DNA and its mutations may be important in centromere drive suppression. The frequency of these positive selection events was comparable to the frequency of positive selection in monocentric clades with asymmetric female meiosis. Taken together, these results suggest that preventing centromere drive is not the primary adaptive role of holocentric chromosomes, and their ability to suppress it likely depends on their kinetochore structure in meiosis.

The transgenerational inheritance of stress-induced epigenetic modifications is still controversial. Despite several examples of defense “priming” and induced genetic rearrangements, the involvement and persistence of transgenerational epigenetic modifications is not known to be general. Here I argue that non-transmission of epigenetic marks through meiosis may be regarded as an epigenetic modification in itself, and that we should understand the implications for plant evolution in the context of both selection for and selection against transgenerational epigenetic memory. Recent data suggest that both epigenetic inheritance and resetting are mechanistically directed and targeted. Stress-induced epigenetic modifications may buffer against DNA sequence-based evolution to maintain plasticity, or may form part of plasticity’s adaptive potential. To date we have tended to concentrate on the question of whether and for how long epigenetic memory persists. I argue that we should now re-direct our question to investigate the differences between where it persists and where it does not, to understand the higher order evolutionary methods in play and their contribution.

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Intermixing of genomes through meiotic reassortment and recombination of homologous chromosomes is a unifying theme of sexual reproduction in eukaryotic organisms and is considered crucial for their adaptive evolution. Previous studies of the budding yeast species Saccharomycodes ludwigii suggested that meiotic crossing over might be absent from its sexual life cycle, which is predominated by fertilization within the meiotic tetrad. We demonstrate that recombination is extremely suppressed during meiosis in Sd. ludwigii. DNA double-strand break formation by the conserved transesterase Spo11, processing and repair involving interhomolog interactions are required for normal meiosis but do not lead to crossing over. Although the species has retained an intact meiotic gene repertoire, genetic and population analyses suggest the exceptionally rare occurrence of meiotic crossovers in its genome. A strong AT bias of spontaneous mutations and the absence of recombination are likely responsible for its unusually low genomic GC level. Sd. ludwigii has followed a unique evolutionary trajectory that possibly derives fitness benefits from the combination of frequent mating between products of the same meiotic event with the extreme suppression of meiotic recombination. This life style ensures preservation of heterozygosity throughout its genome and may enable the species to adapt to its environment and survive with only minimal levels of rare meiotic recombination. We propose Sd. ludwigii as an excellent natural forum for the study of genome evolution and recombination rates.

ABSTRACT Evolution has provided a vast diversity of yeasts that play fundamental roles in nature and society. This diversity is not limited to genotypically homogeneous species with natural interspecies hybrids and allodiploids that blur species boundaries frequently isolated. Thus, life cycle and the nature of breeding systems have profound effects on genome variation, shaping heterozygosity, genotype diversity and ploidy level. The apparent enrichment of hybrids in industry-related environments suggests that hybridization provides an adaptive route against stressors and creates interest in developing new hybrids for biotechnological uses. For example, in the Saccharomyces genus where regulatory circuits controlling cell identity, mating competence and meiosis commitment have been extensively studied, this body of knowledge is being used to combine interesting traits into synthetic F1 hybrids, to bypass F1 hybrid sterility and to dissect complex phenotypes by bulk segregant analysis. Although these aspects are less known in other industrially promising yeasts, advances in whole-genome sequencing and analysis are changing this and new insights are being gained, especially in the food-associated genera Zygosaccharomyces and Kluyveromyces. We discuss this new knowledge and highlight how deciphering cell identity circuits in these lineages will contribute significantly to identify the genetic determinants underpinning complex phenotypes and open new avenues for breeding programmes.

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The regulation of transposable elements (TEs) requires overlapping epigenetic modifications that must be reinforced every cell division and generation. In plants, this is achieved by multiple pathways including small RNAs, DNA methylation, and repressive histone marks that act together to control TE expression and activity throughout the entire life cycle. However, transient TE activation is observed during reproductive transitions as a result of epigenome reprogramming, thus providing windows of opportunity for TE proliferation and epigenetic novelty. Ultimately, these events may originate complex TE-driven transcriptional networks or cell-to-cell communication strategies via mobile small RNAs. In this review, we discuss recent findings and current understanding of TE regulation during sexual plant reproduction, and its implications for fertility, early seed development, and epigenetic inheritance.

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Pericentric heterochromatin (PCH) is a particular form of constitutive heterochromatin that is localized to both sides of centromeres and that forms silent compartments enriched in repressive marks. These genomic regions contain species-specific repetitive satellite DNA that differs in terms of nucleotide sequences and repeat lengths. In spite of this sequence diversity, PCH is involved in many biological phenomena that are conserved among species, including centromere function, the preservation of genome integrity, the suppression of spurious recombination during meiosis, and the organization of genomic silent compartments in the nucleus. PCH organization and maintenance of its repressive state is tightly regulated by a plethora of factors, including enzymes (e.g., DNA methyltransferases, histone deacetylases, and histone methyltransferases), DNA and histone methylation binding factors (e.g., MECP2 and HP1), chromatin remodeling proteins (e.g., ATRX and DAXX), and non-coding RNAs. This evidence helps us to understand how PCH organization is crucial for genome integrity. It then follows that alterations to the molecular signature of PCH might contribute to the onset of many genetic pathologies and to cancer progression. Here, we describe the most recent updates on the molecular mechanisms known to underlie PCH organization and function.

Germline development is a key step in sexual reproduction. Sexual plant reproduction begins with the formation of haploid spores by meiosis of megaspore mother cells (MMCs). Although many evidences, directly or indirectly, show that epigenetics plays an important role in MMC specification, how it controls the commitment of the MMC to downstream stages of germline development is still unclear. Electrophoretic mobility shift assay (EMSA), western blot, immunofluorescence, and chromatin immunoprecipitation coupled with quantitative PCR analyses were performed. Genetic interactions between BZR1 transcription factor family and the SWR1-SDG2-ER pathway in the control of female germline development were further studied. The present findings showed in Arabidopsis that two epigenetic factors, the chromatin remodeling complex SWI2/SNF2-RELATED 1 (SWR1) and a writer for H3K4me3 histone modification SET DOMAIN GROUP 2 (SDG2), genetically interact with the ERECTA (ER) receptor kinase signaling pathway and regulate female germline development by restricting the MMC cell fate to a single cell in the ovule primordium and ensure that only that single cell undergoes meiosis and subsequent megaspore degeneration. We also showed that SWR1-SDG2-ER signaling module regulates female germline development by promoting the protein accumulation of BZR1 transcription factor family on the promoters of primary miRNA processing factors, HYPONASTIC LEAVES 1 (HYL1), DICER-LIKE 1 (DCL1), and SERRATE (SE) to activate their expression. Our study elucidated a Gene Regulation Network that provides new insights for understanding how epigenetic factors and receptor kinase signaling pathways function in concert to control female germline development in Arabidopsis.

The Wassilewskija strain of Arabidopsis has four genes encoding the tryptophan enzyme phosphoribosylanthranilate isomerase (PAI) located at three unlinked sites. These four PAI genes are methylated over their regions of DNA homology. When PAI copy number is reduced by deletion of two tandemly arrayed genes (MePAI1-PAI4), a mutant with fluorescent, tryptophan-deficient phenotypes results, because the two remaining methylated PAI genes (MePAI2 and MePAI3) supply insufficient PAI activity. These two methylated genes can be inherited through meiosis, even when they are segregated away from each other in crosses to a strain with unmethylated PAI genes. However, the mutant phenotypes conferred by the methylated PAI genes are unstable, and mutant plants yield occasional revertant somatic sectors and progeny. Revertant lines display coordinately reduced methylation of both PAI2 and PAI3, implying that this hypomethylation acts in a concerted manner across the genome rather than at individual sites.

Eukaryotic centromeres contain the kinetochore, which connects chromosomes to the spindle allowing segregation. During meiosis, centromeres are suppressed for inter-homolog crossover, as recombination in these regions can cause chromosome missegregation and aneuploidy. Plant centromeres are surrounded by transposon-dense pericentromeric heterochromatin that is epigenetically silenced by histone 3 lysine 9 dimethylation (H3K9me2), and DNA methylation in CG and non-CG sequence contexts. However, the role of these chromatin modifications in control of meiotic recombination in the pericentromeres is not fully understood. Here, we show that disruption of Arabidopsis thaliana H3K9me2 and non-CG DNA methylation pathways, for example, via mutation of the H3K9 methyltransferase genes KYP/SUVH4 SUVH5 SUVH6, or the CHG DNA methyltransferase gene CMT3, increases meiotic recombination in proximity to the centromeres. Using immunocytological detection of MLH1 foci and genotyping by sequencing of recombinant plants, we observe that H3K9me2 and non-CG DNA methylation pathway mutants show increased pericentromeric crossovers. Increased pericentromeric recombination in H3K9me2/non-CG mutants occurs in hybrid and inbred backgrounds and likely involves contributions from both the interfering and noninterfering crossover repair pathways. We also show that meiotic DNA double-strand breaks (DSBs) increase in H3K9me2/non-CG mutants within the pericentromeres, via purification and sequencing of SPO11-1-oligonucleotides. Therefore, H3K9me2 and non-CG DNA methylation exert a repressive effect on both meiotic DSB and crossover formation in plant pericentromeric heterochromatin. Our results may account for selection of enhancer trap Dissociation (Ds) transposons into the CMT3 gene by recombination with proximal transposon launch-pads.

Epigenetic mechanisms, such as DNA methylation, RNA interference, posttranslational histone modifications and rearrangements of chromatin structure play an important role during genome reprogramming in both animals and plants. The correct epigenetic pattern of eu- and heterochromatin marks allows for maintaining chromatin in an active or transcriptionally silenced state. In the life cycle of angiosperms, epigenetic mechanisms participate in genome reprogramming during: 1) differentiation of sporophyte cells into spore mother cells (SMC) that undergo meiosis, 2) development of female and male gametophytes, within which the gametes differentiate and 3) after double fertilization during the embryo and endosperm development. SMC speciation and control of meiosis, followed by reprogramming of the sperm cells and egg cell genome, are non-cell-autonomous and require RdDM pathway. These processes involve companion cells, which produce „mobile” siRNAs signal molecules. Epigenetic control of gene expression through siRNAs also participates in maintenance of gametes and embryo genome integrity and in the parental imprinting.

During meiosis, crossovers between homologous chromosomes generate genetic diversity but are limited in number, widely spaced by interference, and biased toward gene-rich euchromatin while suppressed in pericentromeric heterochromatin. This constrained crossover patterning restricts the genetic variation available for plant breeding. Recent studies have identified key crossover regulators-including the anti-crossover helicases FANCM and RECQ4, the pro-crossover factor HEI10, and heterochromatin-organizing proteins-that can modulate crossover frequency and positioning, although the effects on fertility are species- and context-dependent. Manipulating these pathways offers a strategy to increase crossovers along chromosomes, including recombination-suppressed regions, thereby unlocking hidden genetic variation. Genetic and epigenetic control of crossover formation is emerging as a powerful tool to accelerate crop improvement and enhance genetic gain.

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Centromeres are unique chromosomal loci that form the anchorage point for the mitotic spindle during mitosis and meiosis. Their position and function are specified by a unique chromatin domain featuring the histone H3 variant CENP-A. While typically formed on centromeric satellite arrays, CENP-A nucleosomes are maintained and assembled by a strong self-templated feedback mechanism that can propagate centromeres even at non-canonical sites. Central to the epigenetic chromatin-based transmission of centromeres is the stable inheritance of CENP-A nucleosomes. While long-lived at centromeres, CENP-A can turn over rapidly at non-centromeric sites and even erode from centromeres in non-dividing cells. Recently, SUMO modification of the centromere complex has come to the forefront as a mediator of centromere complex stability, including CENP-A chromatin. We review evidence from different models and discuss the emerging view that limited SUMOylation appears to play a constructive role in centromere complex formation, while polySUMOylation drives complex turnover. The deSUMOylase SENP6/Ulp2 and the proteins segregase p97/Cdc48 constitute the dominant opposing forces that balance CENP-A chromatin stability. This balance may be key to ensuring proper kinetochore strength at the centromere while preventing ectopic centromere formation.

Comparative analysis in Arabidopsis ecotypes, hybrids, and tetraploids revealed the presence of natural variation in early reproductive development that depends on the function of epigenetic pathways. In angiosperms, the transition to the female gametophytic phase relies on the specification of premeiotic gamete precursors from sporophytic cells in the ovule. In Arabidopsis thaliana, a single diploid cell is specified as the premeiotic female gamete precursor. Here, we show that ecotypes of Arabidopsis exhibit differences in megasporogenesis leading to phenotypes reminiscent of defects in dominant mutations that epigenetically affect the specification of female gamete precursors. Intraspecific hybridization and polyploidy exacerbate these defects, which segregate quantitatively in F2 populations derived from ecotypic hybrids, suggesting that multiple loci control cell specification at the onset of female meiosis. This variation in cell differentiation is influenced by the activity of ARGONAUTE9 (AGO9) and RNA-DEPENDENT RNA POLYMERASE6 (RDR6), two genes involved in epigenetic silencing that control the specification of female gamete precursors. The pattern of transcriptional regulation and localization of AGO9 varies among ecotypes, and abnormal gamete precursors in ovules defective for RDR6 share identity with ectopic gamete precursors found in selected ecotypes. Our results indicate that differences in the epigenetic control of cell specification lead to natural phenotypic variation during megasporogenesis. We propose that this mechanism could be implicated in the emergence and evolution of the reproductive alternatives that prevail in flowering plants.

Oocyte quality and number are key determinants of reproductive life span and success. These variables are shaped in part by the elimination of oocytes that experience problems during the early stages of meiosis. Meiotic prophase-I marks an extended period of genome vulnerability in which epigenetic reprogramming unleashes retroelements and hundreds of DNA double-strand breaks (DSBs) are inflicted to initiate the programmed recombination required for accurate chromosome segregation at the first meiotic division. Expression of LINE-1 retroelements perturbs several aspects of meiotic prophase and is associated with oocyte death during the early stages of meiotic prophase I. Defects in chromosome synapsis and recombination also trigger oocyte loss, but typically at a later stage, as cells transition into quiescence and form primordial follicles. Interrelated pathways that signal defects in DSB repair and chromosome synapsis mediate this late oocyte attrition. Here, I review our current understanding of early and late oocyte attrition based on studies in mouse and describe how these processes appear to be both distinct and overlapping and how they help balance the quality and size of oocyte reserves to maximize fecundity.

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Most sexually reproducing eukaryotes use a specialized cell division called meiosis to halve the complement of chromosomes in their gametes. During meiotic prophase I, homologous chromosomes (homologs) recombine by reciprocally exchanging DNA to form cross-overs (COs) that are required for accurate chromosome segregation. COs also reshuffle parental genomes to create genetic diversity among progeny. Molecular genetic studies have identified hundreds of genes involved in meiotic recombination, which have been well summarized in several reviews. Here, we highlight recent advances in understanding endogenous mechanisms that regulate the frequency and distribution of meiotic COs, also called CO patterning. Specifically, we focus on genome-wide regulation, epigenetic control, transcription regulation, and post-transcription processes. Additionally, we highlight open questions that still need further investigation in this field.

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Long non-coding RNAs (lncRNAs) contribute to cell fate decisions by modulating genome expression and stability. In the fission yeast Schizosaccharomyces pombe, the transition from mitosis to meiosis results in a marked remodeling of gene expression profiles, which ultimately ensures gamete production and inheritance of genetic information to the offspring. This key developmental process involves a set of dedicated lncRNAs that shape cell cycle-dependent transcriptomes through a variety of mechanisms, including epigenetic modifications and the modulation of transcription, post-transcriptional and post-translational regulations, and that contribute to meiosis-specific chromosomal events. In this review, we summarize the biology of these lncRNAs, from their identification to mechanism of action, and discuss their regulatory role in the control of gametogenesis.

Polycomb group proteins (PcG) are major epigenetic regulators, essential for establishing heritable expression patterns of developmental control genes. The mouse PcG family member M33/Cbx2 (Chromobox homolog protein 2) is a component of the Polycomb-Repressive Complex 1 (PRC1). Targeted deletion of Cbx2/M33 in mice results in homeotic transformations of the axial skeleton, growth retardation and male-to-female sex reversal. In this study, we tested whether Cbx2 is involved in the control of chromatin remodeling processes during meiosis. Our analysis revealed sex reversal in 28.6% of XY−/− embryos, in which a hypoplastic testis and a contralateral ovary were observed in close proximity to the kidney, while the remaining male mutant fetuses exhibited bilateral testicular hypoplasia. Notably, germ cells recovered from Cbx2(XY−/−) testes on day 18.5 of fetal development exhibited premature meiosis onset with synaptonemal complex formation suggesting a role for Cbx2 in the control of meiotic entry in male germ cells. Mutant females exhibited small ovaries with significant germ cell loss and a high proportion of oocytes with abnormal synapsis and non-homologous interactions at the pachytene stage as well as formation of univalents at diplotene. These defects were associated with failure to resolve DNA double strand breaks marked by persistent γH2AX and Rad51 foci at the late pachytene stage. Importantly, two factors required for meiotic silencing of asynapsed chromatin, ubiquitinated histone H2A (ubH2A) and the chromatin remodeling protein BRCA1, co-localized with fully synapsed chromosome axes in the majority of Cbx2(−/−) oocytes. These results provide novel evidence that Cbx2 plays a critical and previously unrecognized role in germ cell viability, meiosis onset and homologous chromosome synapsis in the mammalian germline.

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It was recently reported that the sizes of many mRNAs change when budding yeast cells exit mitosis and enter the meiotic differentiation pathway. These differences were attributed to length variations of their untranslated regions. The function of UTRs in protein translation is well established. However, the mechanism controlling the expression of distinct transcript isoforms during mitotic growth and meiotic development is unknown. In this study, we order developmentally regulated transcript isoforms according to their expression at specific stages during meiosis and gametogenesis, as compared to vegetative growth and starvation. We employ regulatory motif prediction, in vivo protein-DNA binding assays, genetic analyses and monitoring of epigenetic amino acid modification patterns to identify a novel role for Rpd3 and Ume6, two components of a histone deacetylase complex already known to repress early meiosis-specific genes in dividing cells, in mitotic repression of meiosis-specific transcript isoforms. Our findings classify developmental stage-specific early, middle and late meiotic transcript isoforms, and they point to a novel HDAC-dependent control mechanism for flexible transcript architecture during cell growth and differentiation. Since Rpd3 is highly conserved and ubiquitously expressed in many tissues, our results are likely relevant for development and disease in higher eukaryotes.

Polycomb repressive complex 2 (PRC2) catalyses the repressive epigenetic modification, histone 3 lysine 27 tri-methylation (H3K27me3) and functions as a key epigenetic regulator during embryonic development. PRC2 is known to regulate the development of a range of tissues by transcriptional silencing of genes that control cell differentiation, but its roles in female germline and ovarian development remain unknown. Using a mouse model with hypomorphic EED function that reduced H3K27me3 in somatic and germ cells, we found that PRC2 was required for survival, with more than 95% of female animals dying before birth. Although surviving adult EED hypomorphic females appeared morphologically similar to controls and were fertile, Eedhypo/hypo adult ovaries were abnormal, with altered morphology characterised by abnormal follicles. Early Eedhypo/hypo and control fetal ovaries were morphologically similar, and germ cells entered meiosis normally. Immunofluorescent analyses of somatic and germline markers indicated that ovarian development in Eedhypo/hypo ovaries was similar to heterozygous and wild type controls. However, TUNEL analyses revealed higher rates of apoptosis in the ovarian surface epithelium and transcriptional analyses revealed changes in genes regulating epithelial and steroidogenic cell differentiation, possibly foreshadowing the defects observed in adult ovaries of hypomorphic females. While it was possible to analyse early-mid fetal ovarian development, postnatal stages were inaccessible due to the high level of lethality during late fetal stages. Despite this limitation, the data we were able to obtain reveal a novel role for EED in the ovary that is likely to alter ovarian development and ovarian function in adult animals.

Summary p53 alterations occur during culture of pluripotent stem cells (PSCs), but the significance of these events on epigenetic control of PSC fate determination remains poorly understood. Wdr5 deletion in p53-null (DKO) mouse ESCs (mESCs) leads to impaired self-renewal, defective retinal neuroectoderm differentiation, and de-repression of germ cell/meiosis (GCM)-specific genes. Re-introduction of a WDR5 mutant with defective H3K4 methylation activity into DKO ESCs restored self-renewal and suppressed GCM gene expression but failed to induce retinal neuroectoderm differentiation. Mechanistically, mutant WDR5 targets chromatin that is largely devoid of H3K4me3 and regulates gene expression in p53-null mESCs. Furthermore, MAX and WDR5 co-target lineage-specifying chromatin and regulate chromatin accessibility of GCM-related genes. Importantly, MAX and WDR5 are core subunits of a non-canonical polycomb repressor complex 1 responsible for gene silencing. This function, together with canonical, pro-transcriptional WDR5-dependent MLL complex H3K4 methyltransferase activity, highlight how WDR5 mediates crosstalk between transcription and repression during mESC fate choice.

During germ cell development, epigenetic modifications undergo extensive remodeling. Abnormal epigenetic modifications usually result in germ cell loss and reproductive defect. Prmt5 (Protein arginine methyltransferase 5) encodes a protein arginine methyltransferase which has been demonstrated to play important roles in germ cell development during embryonic stages. In the present study, we found that Prmt5 was also abundantly expressed in male germ cells after birth. Inactivation of this gene by crossing with Stra8-Cre transgenic mice resulted in germ cell loss during spermatogenesis. Further study revealed that the germ cell development was grossly normal before P10. However, most of the germ cells in Prmt5Δ/f; Stra8-Cre mice were blocked at meiotic stage. The expression of meiosis associated genes was reduced in Prmt5Δ/f; Stra8-Cre testes compared to control testes at P10. γH2AX was detected in sex body of control germ cells at P12, whereas multiple foci were observed in Prmt5-deficient germ cells. Further study revealed that H4R3me2s was virtually absent in germ cells after Prmt5 inactivation. The results of this study indicate that Prmt5 also plays important roles in germ cell development during spermatogenesis.

No abstract available

The epidermal growth factor receptor (EGFR) pathway is directly involved in oocyte meiotic resumption induced by a gonadotropic stimulus. Here, we used an EGFR inhibitor (AG1478) to inhibit spontaneous meiosis resumption in bovine oocytes (EGFR- group) during 8 hr prematuration and assessed the competence of such oocytes for embryonic development, apoptosis and gene expression in comparison with Control group which was not prematured. Data are presented as mean ± SEM. Blastocysts rate on day 7 (40.81%, averaged) and hatching rate on day 9 (77.35%, averaged) were unaffected by treatment (p > 0.05). Similarly, treatment did not affect (p > 0.05) the total cell number on day 7 (119.05, averaged) and on day 9 (189.5, averaged). Apoptosis was reduced (p < 0.05) in EGFR- group day 7-embryos compared to Control group (3.7% ± 1.0 vs. 5.2% ± 0.8). Abundance of several transcripts was upregulated (p < 0.05) in EGFR- group, including genes related to embryo development and quality (NANOG and RPLP0), epigenetic regulation (H2AFZ), apoptosis (BID) and stress response (GPX4 and HIF1A). Taken together, the results presented here demonstrated a reduction in the apoptosis index and upregulation of NANOG, H2AFZ and RPLP0 mRNA levels, which are related to embryonic development. Our data suggest that temporary meiosis blockage with EGFR inhibitor during prematuration culture of bovine oocytes may be an interesting strategy to improve embryo quality.

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The synaptonemal complex (SC) forms between homologous chromosomes during meiosis. In Arabidopsis thaliana, its central region (CR) is composed of the transverse filament protein ZYP1 and the central element proteins SCEP1 and SCEP2. Here we identify SCEP3 as a CR protein that is evolutionarily conserved across plant species. SCEP3 spatiotemporally overlaps with other CR proteins and localizes to the SC CR. The loss of SCEP3 prevents SC assembly, abolishes crossover (CO) assurance and interference, and eliminates sex-specific differences in CO rates (heterochiasmy) through increased CO in females. SCEP3 is required for a subset of COs in SC-deficient mutants, such as zyp1. Although SCEP3 physically interacts with ZYP1, it loads independently of other CR proteins. We propose that SCEP3 may associate with certain recombination intermediates, stabilizing them and/or recruiting additional factors, such as ZYP1, to a subset of these intermediates, thereby promoting and interlinking SC assembly and CO formation. This study identifies SCEP3 as a conserved central element of the plant synaptonemal complex. SCEP3 is critical for synapsis, heterochiasmy and crossover interference. SCEP3 also promotes a subset of crossover events.

Several proteins collaborate to promote the crossover recombination events critical for accurate chromosome segregation during meiosis. How these “ZMM” factors (Zip2, Zip3, Zip4, Spo16, Mer3 and MutSγ) collaboratively function remains incompletely understood. We previously reported that Zip3’s abundance and activity rely on the synaptonemal complex (SC) component Zip1, and specifically on Zip1’s N-terminal residues associated with crossovers and coupling SC assembly to the crossover pathway. Here, we demonstrate that Zip3 co-immunoprecipitates Zip1 from meiotic cells independent of recombination initiation and other ZMMs, and that Zip3’s interaction with Zip1 relies on Zip1’s N terminal residues. Co-expression and pull-down experiments in bacterial cells demonstrate that Zip1 and Zip3 interact directly. Experiments to identify Zip3 regions required for the Zip1 interaction unexpectedly revealed an incorrectly annotated translational start; we also determined that Zip3’s N-terminal structured region is necessary and sufficient for the interaction, and a predicted coil downstream of Zip3’s RING domain is essential for specific activities attributed to Zip1’s N-terminal tip such as proximity labeling of Zip3 by Zip2 and the coupling of crossover recombination to SC assembly. Finally, we discovered that interaction with Zip1 protects Zip3 not only from proteasome-mediated degradation but also from post-translational modification when another ZMM is absent. We propose that direct interaction with Zip1’s N terminus orients Zip3 within a nascent ZMM ensemble in a manner that facilitates crossover formation and the coupling of crossover intermediates to SC assembly, and furthermore ensures Zip3 remains both abundant and unmodified until all requisite ZMMs have joined the group.

In Drosophila melanogaster females, as in most organisms, the segregation of homologous chromosomes in meiosis depends on the formation of crossovers between them. In most cases, crossovers require the synaptonemal complex (SC), a conserved multi-protein structure that forms between homologous chromosomes in early meiosis. Recent studies leveraging partial-loss-of-function alleles suggest that the SC plays a more direct role in crossover formation. One SC protein that is involved in crossover formation is SYP-4 in nematodes, which we found is a likely ortholog of the D. melanogaster SC protein Corolla. To create a hypomorphic allele of corolla in D. melanogaster, we used CRISPR/Cas9 to replace it with its D. mauritiana ortholog, yielding corollamau. Since SC protein sequences are rapidly diverging while maintaining the SC’s structure, we hypothesized that this replacement would enable SC assembly but show defects in crossover formation. Indeed, at 25 °C corollamau homozygous females exhibited defects in SC maintenance and crossover formation, resulting in moderate levels of chromosome missegregation. At 18 °C, SC maintenance was rescued, and recombination rates were improved, although they remained significantly lower than observed in wild type. Unexpectedly, in homozygotes we also observed unique polycomplexes composed of the SC proteins Corolla and Corona but lacking the transverse filament protein C(3)G. Overall, we report a novel hypomorphic allele of corolla that suggests Corolla regulates crossover formation. Further, the unique polycomplexes found in mutant flies may provide new insights into SC architecture and protein-protein interactions.

The shuffling of genetic material facilitated by meiotic crossovers is a critical driver of genetic variation. Therefore, the number and positions of crossover events must be carefully controlled. In Arabidopsis, an obligate crossover and repression of nearby crossovers on each chromosome pair are abolished in mutants that lack the synaptonemal complex (SC), a conserved protein scaffold. We use mathematical modelling and quantitative super-resolution microscopy to explore and mechanistically explain meiotic crossover pattering in Arabidopsis lines with full, incomplete or abolished synapsis. For zyp1 mutants, which lack an SC, we develop a coarsening model in which crossover precursors globally compete for a limited pool of the pro-crossover factor HEI10, with dynamic HEI10 exchange mediated through the nucleoplasm. We demonstrate that this model is capable of quantitatively reproducing and predicting zyp1 experimental crossover patterning and HEI10 foci intensity data. Additionally, we find that a model combining both SC- and nucleoplasm-mediated coarsening can explain crossover patterning in wild-type Arabidopsis and in pch2 mutants, which display partial synapsis. Together, our results reveal that regulation of crossover patterning in wild-type Arabidopsis and SC defective mutants likely act through the same underlying coarsening mechanism, differing only in the spatial compartments through which the pro-crossover factor diffuses.

Significance Meiotic recombination promotes genetic diversity by shuffling parental chromosomes. As observed by the very first geneticists, crossovers inhibit the formation of another crossover nearby, an elusive phenomenon called crossover interference. Another intriguing observation is heterochiasmy, the marked difference in male and female crossover rates observed in many species. Here, we show that the synaptonemal complex, a structure that zips homologous chromosomes together during meiosis, is essential for crossover interference in Arabidopsis. This suggests that a signal that inhibits crossover formation nearby a first crossover propagates along this specific structure. Furthermore, in the absence of the synaptonemal complex, crossover frequencies become identical in both sexes, suggesting that heterochiasmy is due to variation of crossover interference imposed by the synaptonemal complex. Meiotic crossovers (COs) have intriguing patterning properties, including CO interference, the tendency of COs to be well-spaced along chromosomes, and heterochiasmy, the marked difference in male and female CO rates. During meiosis, transverse filaments transiently associate the axes of homologous chromosomes, a process called synapsis that is essential for CO formation in many eukaryotes. Here, we describe the spatial organization of the transverse filaments in Arabidopsis (ZYP1) and show it to be evolutionary conserved. We show that in the absence of ZYP1 (zyp1a zyp1b null mutants), chromosomes associate in pairs but do not synapse. Unexpectedly, in absence of ZYP1, CO formation is not prevented but increased. Furthermore, genome-wide analysis of recombination revealed that CO interference is abolished, with the frequent observation of close COs. In addition, heterochiasmy was erased, with identical CO rates in males and females. This shows that the tripartite synaptonemal complex is dispensable for CO formation and has a key role in regulating their number and distribution, imposing CO interference and heterochiasmy.

Meiotic crossovers are limited in number and are prevented from occurring close to each other by crossover interference. In many species, crossover number is subject to sexual dimorphism, and a lower crossover number is associated with shorter chromosome axes lengths. How this patterning is imposed remains poorly understood. Here, we show that overexpression of the Arabidopsis pro-crossover protein HEI10 increases crossovers but maintains some interference and sexual dimorphism. Disrupting the synaptonemal complex by mutating ZYP1 also leads to an increase in crossovers but, in contrast, abolishes interference and disrupts the link between chromosome axis length and crossovers. Crucially, combining HEI10 overexpression and zyp1 mutation leads to a massive and unprecedented increase in crossovers. These observations support and can be predicted by, a recently proposed model in which HEI10 diffusion along the synaptonemal complex drives a coarsening process leading to well-spaced crossover-promoting foci, providing a mechanism for crossover patterning. During meiosis, the number and distribution of crossovers (COs) are tightly controlled, but the mechanistic basis of this control is unclear. Here, by combining experimental data and mathematical modeling, the study advocates a CO patterning model via coarsening through the diffusion of HEI10 along the synaptonemal complex.

During the formation of ova and sperm, homologous chromosomes get physically attached through the synaptonemal complex and exchange DNA at crossover sites by a process known as meiotic recombination. Chromosomes that do not recombine or have anomalous crossover distributions often separate poorly during the subsequent cell division and end up in abnormal numbers in ova or sperm, which can lead to miscarriage or developmental defects. Crossover numbers and distribution along the synaptonemal complex can be visualized by immunofluorescent microscopy. However, manual analysis of large numbers of cells is very time-consuming and a major bottleneck for recombination studies. Some image analysis tools have been created to overcome this situation, but they are not readily available, do not provide synaptonemal complex data, or do not tackle common experimental difficulties, such as overlapping chromosomes. To overcome these limitations, we have created and validated an open-source ImageJ macro routine that facilitates and speeds up the crossover and synaptonemal complex analyses in mouse chromosome spreads, as well as in other vertebrate species. It is free, easy to use and fulfills the recommendations for enhancing rigor and reproducibility in biomedical studies.

Meiotic recombination is triggered by programmed double-strand breaks (DSBs), a subset of these being repaired as crossovers, promoted by eight evolutionarily conserved proteins, named ZMM. Crossover formation is functionally linked to synaptonemal complex (SC) assembly between homologous chromosomes, but the underlying mechanism is unknown. Here we show that Ecm11, a SC central element protein, localizes on both DSB sites and sites that attach chromatin loops to the chromosome axis, which are the starting points of SC formation, in a way that strictly requires the ZMM protein Zip4. Furthermore, Zip4 directly interacts with Ecm11 and point mutants that specifically abolish this interaction lose Ecm11 binding to chromosomes and exhibit defective SC assembly. This can be partially rescued by artificially tethering interaction-defective Ecm11 to Zip4. Mechanistically, this direct connection ensuring SC assembly from CO sites could be a way for the meiotic cell to shut down further DSB formation once enough recombination sites have been selected for crossovers, thereby preventing excess crossovers. Finally, the mammalian ortholog of Zip4, TEX11, also interacts with the SC central element TEX12, suggesting a general mechanism.

Abstract Chromosome axis-associated HORMA domain proteins (HORMADs), e.g. ASY1 in Arabidopsis, are crucial for meiotic recombination. ASY1, as other HORMADs, is assembled on the axis at early meiosis and depleted when homologous chromosomes synapse. Puzzlingly, both processes are catalyzed by AAA+ ATPase PCH2 together with its cofactor COMET. Here, we show that the ASY1 remodeling complex is temporally and spatially differently assembled. While PCH2 and COMET appear to directly interact in the cytoplasm in early meiosis, PCH2 is recruited by the transverse filament protein ZYP1 and brought to the ASY1-bound COMET assuring the timely removal of ASY1 during chromosome synapsis. Since we found that the PCH2 homolog TRIP13 also binds to the ZYP1 homolog SYCP1 in mouse, we postulate that this mechanism is conserved among eukaryotes. Deleting the PCH2 binding site of ZYP1 led to a failure of ASY1 removal. Interestingly, the placement of one obligatory crossover per homologous chromosome pair, compromised by ZYP1 depletion, is largely restored in this separation-of-function zyp1 allele suggesting that crossover assurance is promoted by synapsis. In contrast, this zyp1 allele, similar to the zyp1 null mutant, showed elevated type I crossover numbers indicating that PCH2-mediated eviction of ASY1 from the axis restricts crossover formation.

In this Outlook, Grey and de Massy discuss a study by Pyatnitskaya et al. in this issue of Genes & Development that highlights the central role of the Saccharomyces cerevisiae ZMM protein Zip4 in how crossover formation and synapsis initiation are linked.

The synaptonemal complex (SC) is essential for accurate homologous chromosome pairing, recombination, and segregation during meiosis. Although several core SC components have been identified in plants, the molecular mechanism coordinating their assembly remains poorly understood. Here, through a refined temporal transcriptomic analysis of another development in the allotetraploid Brassica napus, this study identifies ZYP1-SCEP1/2 Linker (ZSL) as a central element scaffold that bridges the transverse filament protein ZYP1 with the heterodimeric central element proteins SCEP1/2. It is shown that ZYP1 loading onto chromosomes occurs independently of ZSL and SCEP1/2, whereas ZYP1 is required for their recruitment. Loss of ZSL impedes SCEP1/2 recruitment but not vice versa, and zsl mutants completely lack continuous SC central region assembly, leading to synapsis failure and chromosome mis-segregation. This study further demonstrates that ZSL directly interacts with both ZYP1 and SCEP1/2. These findings define a hierarchical assembly cascade of ZYP1 → ZSL → SCEP1/2 during SC formation. Furthermore, analysis of HEI10 foci and genome-wide crossover (CO) mapping in zsl mutants reveals an ≈100% increase in both male and female COs, accompanied by a loss of interference and elimination of sex-specific CO differences. Together, the results establish ZSL as a key molecular adaptor coordinating SC central region assembly and CO patterning, providing new mechanistic insight into meiotic fidelity and genome stability in polyploid species.

The synaptonemal complex (SC) is a proteinaceous structure that mediates homolog engagement and genetic recombination during meiosis. Zip-Mer-Msh (ZMM) proteins promote crossover (CO) formation and initiate SC formation. In SC elongation, the SUMOylated SC component Ecm11 and its interacting protein Gmc2 facilitate the polymerization of Zip1, a SC-central region component in budding yeast. Through physical recombination, cytological, and genetic analyses, we here demonstrate that ecm11 and gmc2 mutants exhibit chromosome-specific defects in meiotic recombination. CO frequencies were reduced on a short chromosome (chromosome III), whereas CO and non-crossover (NCO) frequencies were increased on a long chromosome (chromosome VII). Further, persistent double-strand breaks (DSBs) occurred in unsynapsed chromosome regions during the late prophase, suggesting the presence of a negative regulation of DSB formation. The Ecm11-Gmc2 (EG) complex could participate in joint molecule (JM) processing and/or double-Holliday junction resolution for CO-designated recombination of the ZMM-dependent pathway. However, absence of the EG complex ameliorated the JM-processing defect in zmm mutants, suggesting a role of these proteins in suppression of ZMM-independent recombination. Therefore, the EG complex fosters ZMM-dependent processing and resolution of JMs while suppressing ZMM-independent JM processing and late DSB formation. Hence, EG-mediated SC central regions, which display properties similar to those of liquid crystals, may function as a compartment for sequestering recombination proteins in and out of the process to ensure meiosis specificity during recombination.

During sexual reproduction the parental homologous chromosomes find each other (pair) and align along their lengths by integrating local sequence homology with large-scale contiguity, thereby allowing for precise exchange of genetic information. The Synaptonemal Complex (SC) is a conserved zipper-like structure that assembles between the homologous chromosomes. This phase-separated interface brings chromosomes together and regulates exchanges between them. However, the molecular mechanisms by which the SC carries out these functions remain poorly understood. Here we isolated and characterized two mutations in the dimerization interface in the middle of the SC zipper in C. elegans. The mutations perturb both chromosome alignment and the regulation of genetic exchanges. Underlying the chromosome-scale phenotypes are distinct alterations to the way SC subunits interact with one another. We propose that the SC brings homologous chromosomes together through two biophysical activities: obligate dimerization that prevents assembly on unpaired chromosomes; and a tendency to phase-separate that extends pairing interactions along the entire length of the chromosomes.

Errors during meiosis are the leading cause of birth defects and miscarriages in humans. Thus, the coordinated control of meiotic events is critical for the faithful inheritance of the genome with each generation... Crossovers (COs) between homologous chromosomes are critical for meiotic chromosome segregation and form in the context of the synaptonemal complex (SC), a meiosis-specific structure that assembles between aligned homologs. During Caenorhabditis elegans meiosis, central region components of the SC (SYP proteins) are essential to repair double-strand DNA breaks (DSBs) as COs. Here, we investigate the relationships between the SYP proteins and conserved pro-CO factors by examining the immunolocalization of these proteins in meiotic mutants where SYP proteins are absent, reduced, or mislocalized. Although COs do not form in syp null mutants, pro-CO factors COSA-1, MSH-5, and ZHP-3 nevertheless colocalize at DSB-dependent sites during late prophase, reflecting an inherent affinity of these factors for DSB repair sites. In contrast, in mutants where SYP proteins are present but form aggregates or display abnormal synapsis, pro-CO factors consistently track with SYP-1 localization. Further, pro-CO factors usually localize to a single site per SYP-1 structure, even in SYP aggregates or in mutants where the SC forms between sister chromatids, suggesting that CO regulation occurs within these aberrant SC structures. Moreover, we find that the meiotic cohesin REC-8 is required to ensure that SC formation occurs between homologs and not sister chromatids. Taken together, our findings support a model in which SYP proteins promote CO formation by promoting the localization of pro-CO factors to recombination events within an SC compartment, thereby ensuring that pro-CO factors identify a recombination event within an SC structure and that CO maturation occurs only between properly aligned homologous chromosomes.

The centromere effect (CE) is a meiotic phenomenon that ensures meiotic crossover suppression in pericentromeric regions. Despite being a critical safeguard against nondisjunction, the mechanisms behind the CE remain unknown. Previous studies found that different regions of the Drosophila pericentromere, encompassing proximal euchromatin, beta and alpha heterochromatin, undergo varying levels of crossover suppression, raising the question of whether distinct mechanisms establish the CE in different regions. We asked whether different pericentromeric regions respond differently to mutations that impair features that may play a role in the CE. In flies with a mutation that affects the synaptonemal complex (SC), a structure is hypothesized to have roles in recombination and crossover patterning, we observed a redistribution of pericentromeric crossovers from proximal euchromatin towards beta heterochromatin but not alpha heterochromatin, indicating a role for the SC in suppressing crossovers in beta heterochromatin. In flies mutant for mei-218 or rec, which encode components of a critical pro-crossover complex, there was a more extreme redistribution of pericentromeric crossovers towards both beta and alpha heterochromatin, suggesting an important role for these meiotic recombination factors in suppressing heterochromatic crossovers. We mapped crossovers in flies mutant for Su(var)3-9, which encodes histone H3-lysine-9 methyltransferase. Although we expected strong alleviation of crossover suppression in heterochromatin, no changes in pericentromeric crossover distribution were observed in this mutant, indicating that this vital heterochromatin factor is dispensable for preventing crossovers in heterochromatin. Thus, in D. melanogaster the meiotic machinery seems to play a more significant role in suppressing centromere-proximal crossovers than chromatin state.

Meiotic recombination plays an important role in ensuring proper chromosome segregation during meiosis I through the creation of chiasmata that connect homologous chromosomes. Recombination plays an additional role in evolution by creating new allelic combinations. Organisms display species-specific crossover patterns, but how these patterns are established is poorly understood. Drosophila mauritiana displays a different meiotic recombination pattern compared to Drosophila melanogaster, with D. mauritiana experiencing a reduced centromere effect, the suppression of recombination emanating from the centromeres. To evaluate the contribution of the synaptonemal complex (SC) C(3)G protein to these recombination rate differences, the D. melanogaster allele was replaced with D. mauritiana c(3)G coding sequence. We found that the D. mauritiana C(3)G could interact with the D. melanogaster SC machinery to build full length tripartite SC and chromosomes segregated accurately, indicating sufficient crossovers were generated. However, the placement of crossovers was altered, displaying an increase in frequency in the centromere-proximal euchromatin indicating a decrease in the centromere effect, similar to that observed in D. mauritiana females. Recovery of chromatids with more than one crossover was also increased, likely due to the larger chromosome span now available for crossovers. As replacement of a single gene mediated a strong shift of one species’ crossover pattern towards another species, it indicates a small number of discrete factors may have major influence on species-specific crossover patterning. Additionally, it demonstrates the SC, a structure known to be required for crossover formation in many species, is likely one of these discrete factors.

During meiosis, the maternal and paternal homologous chromosomes must align along their entire length and recombine to achieve faithful segregation in the gametes. Meiotic recombination is accomplished through the formation of DNA double-strand breaks, a subset of which can mature into crossovers to link the parental homologous chromosomes and promote their segregation. Breast and ovarian cancer susceptibility protein BRCA1 and its heterodimeric partner BARD1 play a pivotal role in DNA repair in mitotic cells; however, their functions in gametogenesis are less well understood. Here we show that localization of BRC-1 and BRD-1 (Caenorhabditis elegans orthologues of BRCA1 and BARD1) is dynamic during meiotic prophase I; they ultimately becoming concentrated at regions surrounding the presumptive crossover sites, co-localizing with the pro-crossover factors COSA-1, MSH-5 and ZHP-3. The synaptonemal complex and PLK-2 activity are essential for recruitment of BRC-1 to chromosomes and its subsequent redistribution towards the short arm of the bivalent. BRC-1 and BRD-1 form in vivo complexes with the synaptonemal complex component SYP-3 and the crossover-promoting factor MSH-5. Furthermore, BRC-1 is essential for efficient stage-specific recruitment/stabilization of the RAD-51 recombinase to DNA damage sites when synapsis is impaired and upon induction of exogenous damage. Taken together, our data provide new insights into the localization and meiotic function of the BRC-1–BRD-1 complex and highlight its essential role in DNA double-strand break repair during gametogenesis.

Synaptonemal complex (SC) is a structurally conserved, supramolecular assembly that forms at the interface of aligned chromosome axes during meiosis, where it provides a physical context for crossover recombination intermediates. In yeast, the SC is composed of Zip1 transverse filaments and central element proteins Ecm11 and Gmc2. Here, we identify a biochemically stable constitutive complex between Ecm11 and Gmc2, which is mediated by their α-helical coiled-coil regions formed of amino acids 230-302 and 59-188, respectively. We find that the Ecm11-Gmc2 is a 2 : 2 hetero-oligomer, which has an architecture and dimensions similar to the mammalian SC central element complex SYCE2-TEX12. Through targeted mutagenesis in yeast, we show that 2 : 2 Ecm11-Gmc2 complex formation is essential for SC assembly in vivo. Further, we identify key additional residues, particularly in Ecm11, that are dispensable for heterocomplex formation in vitro but critical for stability of the complex in vivo.

Meiosis, a specialised form of cell division, is essential for sexual reproduction, which requires the proper formation of synaptonemal complex (SC) and homologous recombination (HR). However, the regulatory mechanisms underlying these processes remain incompletely understood. Here, we demonstrate that SOX30 is a key transcriptional regulator of male meiotic synapsis and recombination. In Sox30-knockout mice, zygotene spermatocytes accumulate with synapsis defects. SOX30 deficiency disrupts the SC central element components SYCE1, SYCE2, and TEX12 distribution. Furthermore, disrupted γ-H2AX distribution reveals impaired DNA double-strand break repair and the persistence of recombination proteins RAD51 and RPA2 in late spermatocytes confirms defective homologous recombination repair (HRR) which results in reduced crossover formation in Sox30-knockout mice spermatocytes. Mechanistically, SOX30 directly binds to SYCE1/SYCE2 promoters to modulate their transcription, thereby regulating SC assembly and HRR. Restoring SOX30 expression effectively rescues meiotic defects. Importantly, transcriptome co-expression analysis in non-obstructive azoospermia (NOA) testes identifies SOX30 as a central regulator of NOA transcriptional networks. Collectively, these findings underscore SOX30's crucial role in meiotic synapsis and recombination, highlighting its therapeutic potential for NOA.

Accurate chromosome segregation during meiosis relies on the prior establishment of at least one crossover recombination event between homologous chromosomes, which is often associated with the meiosis-specific MutSγ complex. The recombination intermediates that give rise to MutSγ interhomolog crossovers are embedded within a hallmark meiotic prophase structure called the synaptonemal complex (SC), but the mechanisms that coordinate the processes of SC assembly (synapsis) and crossover recombination remain poorly understood. Among known central region building blocks of the budding yeast SC, the Zip1 protein is unique for its SC-independent role in promoting MutSγ crossovers. Here we report that adjacent regions within Zip1’s unstructured N terminus encompass its crossover and SC assembly functions. We previously showed that deletion of Zip1 residues 21-163 abolishes tripartite SC assembly and prevents the robust SUMOylation of the SC central element component, Ecm11, but allows excess MutSγ crossover recombination. We find the reciprocal phenotype when Zip1 residues 2-9 or 10-14 are deleted; in these mutants SC assembles and Ecm11 is hyperSUMOylated, but MutSγ crossovers are strongly diminished. Interestingly, Zip1 residues 2-9 or 2-14 are required for the normal localization of Zip3, a putative E3 SUMO ligase and pro-MutSγ crossover factor, to Zip1 polycomplex structures and to recombination initiation sites. By contrast, deletion of Zip1 residues 15-20 does not detectably prevent Zip3’s localization at Zip1 polycomplex and supports some MutSγ crossing over but prevents normal SC assembly and robust Ecm11 SUMOylation. These results highlight distinct N terminal regions that are differentially critical for Zip1’s roles in crossover recombination and SC assembly; we speculate that the adjacency of these regions enables Zip1 to serve as a liaison, facilitating crosstalk between the two processes by bringing crossover recombination and synapsis factors in close proximity to one another.

To ensure the accurate segregation of homologous chromosomes and enhance the genetic diversity in the progeny, meiosis depends on the formation of crossovers between homologous chromosomes. The number and distribution of these crossovers must be precisely regulated through crossover assurance and interference to prevent chromosome missegregation and genomic instability. Here we show that the regulation of crossovers depends on a disordered domain within the synaptonemal complex, which is highly conserved. This domain is located at the C-terminus of the central element protein SYP-4 in C. elegans. While not necessary for synapsis, the C-terminus of SYP-4 is crucial for both crossover assurance and interference. Although the SYP-4 C-terminus contains many potential phosphorylation sites, we found that phosphorylation is not the primary regulator of crossover events. Instead, we discovered that nine conserved phenylalanines recruit a pro-crossover factor predicted to be an E3 ligase and regulate the physical properties of the synaptonemal complex. We propose that this conserved and disordered domain plays a crucial role in maintaining the synaptonemal complex in an activated state to promote crossing-over. This activation allows the synaptonemal complex to regulate the number and distribution of crossovers along chromosomes, thereby protecting the genome for future generations.

No abstract available

Asymmetric disassembly of the synaptonemal complex (SC) is crucial for proper meiotic chromosome segregation. However, the signaling mechanisms that directly regulate this process are poorly understood. Here we show that the mammalian Rho GEF homolog, ECT-2, functions through the conserved RAS/ERK MAP kinase signaling pathway in the C. elegans germline to regulate the disassembly of SC proteins. We find that SYP-2, a SC central region component, is a potential target for MPK-1-mediated phosphorylation and that constitutively phosphorylated SYP-2 impairs the disassembly of SC proteins from chromosomal domains referred to as the long arms of the bivalents. Inactivation of MAP kinase at late pachytene is critical for timely disassembly of the SC proteins from the long arms, and is dependent on the crossover (CO) promoting factors ZHP-3/RNF212/Zip3 and COSA-1/CNTD1. We propose that the conserved MAP kinase pathway coordinates CO designation with the disassembly of SC proteins to ensure accurate chromosome segregation. DOI: http://dx.doi.org/10.7554/eLife.12039.001

Although Lepidopteran females build a synaptonemal complex (SC) in pachytene, homologs do not crossover, necessitating an alternative method of homolog conjunction. In Bombyx mori oocytes, the SC breaks down at the end of pachytene, and homolog associations are maintained by a large oocyte-specific structure, which we call the bivalent bridge (BB), connecting paired homologs. The BB is derived from at least some components of the SC lateral elements (LEs). It contains the HORMAD protein HOP1 and the LE protein SYCP2 and is formed by the fusion of the two LE derivatives. As diplotene progresses, the BB increases in width and acquires a layered structure with a thick band of HOP1 separating two layers of SYCP2. The HOP1 interacting protein, PCH2, joins the BB in mid-diplotene, and by late-diplotene, it lies in the middle of the HOP1 filament. This structure is maintained through metaphase I. SYCP2 and PCH2 are lost at anaphase I, and the BB no longer connects the separating homologs. However, a key component of the BB, HOP1, remains at the metaphase I plate. These changes in organization of the BB occur simultaneously with the movement of the kinetochore protein, DSN1, from within the BB at mid-diplotene to the edge of the homologs facing the poles by metaphase I. We view these data in context of models in which SC components and regulators can be repurposed to achieve different functions, a fascinating example of evolution achieving homolog conjunction in an alternative way with recycling of SC proteins.

Interhomolog crossovers promote proper chromosome segregation during meiosis and are formed by the regulated repair of programmed double-strand breaks. This regulation requires components of the synaptonemal complex (SC), a proteinaceous structure formed between homologous chromosomes. In yeast, SC formation requires the “ZMM” genes, which encode a functionally diverse set of proteins, including the transverse filament protein, Zip1. In wild-type meiosis, Zmm proteins promote the biased resolution of recombination intermediates into crossovers that are distributed throughout the genome by interference. In contrast, noncrossovers are formed primarily through synthesis-dependent strand annealing mediated by the Sgs1 helicase. This work identifies a conserved region on the C terminus of Zip1 (called Zip1 4S), whose phosphorylation is required for the ZMM pathway of crossover formation. Zip1 4S phosphorylation is promoted both by double-strand breaks (DSBs) and the meiosis-specific kinase, MEK1/MRE4, demonstrating a role for MEK1 in the regulation of interhomolog crossover formation, as well as interhomolog bias. Failure to phosphorylate Zip1 4S results in meiotic prophase arrest, specifically in the absence of SGS1. This gain of function meiotic arrest phenotype is suppressed by spo11Δ, suggesting that it is due to unrepaired breaks triggering the meiotic recombination checkpoint. Epistasis experiments combining deletions of individual ZMM genes with sgs1-md zip1-4A indicate that Zip1 4S phosphorylation functions prior to the other ZMMs. These results suggest that phosphorylation of Zip1 at DSBs commits those breaks to repair via the ZMM pathway and provides a mechanism by which the crossover/noncrossover decision can be dynamically regulated during yeast meiosis.

Exchanges between DNA molecules during meiosis are tightly controlled. This work shows that the synaptonemal complex is instrumental in regulating meiotic exchanges both between the parental chromosomes and between the sister chromatids. During meiosis, programmed double-strand DNA breaks are repaired to form exchanges between the parental chromosomes called crossovers. Chromosomes lacking a crossover fail to segregate accurately into the gametes, leading to aneuploidy. In addition to engaging the homolog, crossover formation requires the promotion of exchanges, rather than non-exchanges, as repair products. However, the mechanism underlying this meiosis-specific preference is not fully understood. Here, we study the regulation of meiotic sister chromatid exchanges in Caenorhabditis elegans by direct visualization. We find that a conserved chromosomal interface that promotes exchanges between the parental chromosomes, the synaptonemal complex, can also promote exchanges between the sister chromatids. In both cases, exchanges depend on the recruitment of the same set of pro-exchange factors to repair sites. Surprisingly, although the synaptonemal complex usually assembles between the two DNA molecules undergoing an exchange, its activity does not rely on a specific chromosome conformation. This suggests that the synaptonemal complex regulates exchanges—both crossovers and sister exchanges—by establishing a nuclear domain conducive to nearby recruitment of exchange-promoting factors.

About 10% of reproductive-aged couples suffer from infertility. However, the genetic causes of human infertility cases are largely unknown. Meiosis produces haploid gametes for fertilization and errors in meiosis are associated with human infertility in both males and females. Successful meiosis relies on the assembly of the synaptonemal complex (SC) between paired homologous chromosomes during the meiotic prophase. The SC is ultrastructurally and functionally conserved, promoting inter-homologous recombination and crossover formation, thus critical for accurate meiotic chromosome segregation. With whole-genome/exome sequencing and mouse models, a list of mutations in SC coding genes has been linked to human infertility. Here we summarize those findings. We also analyzed SC gene variants present in the general population and presented complex interaction networks associated with SC components. Whether a combination of genetic variations and environmental factors causes human infertility demands further investigations.

DNA double-strand breaks (DSBs) are deleterious lesions, which must be repaired precisely to maintain genomic stability. During meiosis, programmed DSBs are repaired via homologous recombination (HR) while repair using the nonhomologous end joining (NHEJ) pathway is inhibited, thereby ensuring crossover formation and accurate chromosome segregation.1,2 How DSB repair pathway choice is implemented during meiosis is unknown. In C. elegans, meiotic DSB repair takes place in the context of the fully formed, highly dynamic zipper-like structure present between homologous chromosomes called the synaptonemal complex (SC).3-9 The SC consists of a pair of lateral elements bridged by a central region composed of the SYP proteins in C. elegans. How the structural components of the SC are regulated to maintain the architectural integrity of the assembled SC around DSB repair sites remained unclear. Here, we show that SYP-4, a central region component of the SC, is phosphorylated at Serine 447 in a manner dependent on DSBs and the ATM/ATR DNA damage response kinases. We show that this SYP-4 phosphorylation is critical for preserving the SC structure following exogenous (γ-IR-induced) DSB formation and for promoting normal DSB repair progression and crossover patterning following SPO-11-dependent and exogenous DSBs. We propose a model in which ATM/ATR-dependent phosphorylation of SYP-4 at the S447 site plays important roles both in maintaining the architectural integrity of the SC following DSB formation and in warding off repair via the NHEJ repair pathway, thereby preventing aneuploidy.

Abstract Meiosis is a specialized cell division that gives rise to genetically distinct gametic cells. Meiosis relies on the tightly controlled formation of DNA double-strand breaks (DSBs) and their repair via homologous recombination for correct chromosome segregation. Like all forms of DNA damage, meiotic DSBs are potentially harmful and their formation activates an elaborate response to inhibit excessive DNA break formation and ensure successful repair. Previous studies established the protein kinase ATM as a DSB sensor and meiotic regulator in several organisms. Here we show that Arabidopsis ATM acts at multiple steps during DSB formation and processing, as well as crossover (CO) formation and synaptonemal complex (SC) organization, all vital for the successful completion of meiosis. We developed a single-molecule approach to quantify meiotic breaks and determined that ATM is essential to limit the number of meiotic DSBs. Local and genome-wide recombination screens showed that ATM restricts the number of interference-insensitive COs, while super-resolution STED nanoscopy of meiotic chromosomes revealed that the kinase affects chromatin loop size and SC length and width. Our study extends our understanding of how ATM functions during plant meiosis and establishes it as an integral factor of the meiotic program.

No abstract available

No abstract available

Homologous recombination (HR) is an essential meiotic process that contributes to the genetic variation of offspring and ensures accurate chromosome segregation. Recombination is facilitated by the formation and repair of programmed DNA double-strand breaks. These DNA breaks are repaired via recombination between maternal and paternal homologous chromosomes and a subset result in the formation of crossovers. HR and crossover formation is facilitated by synapsis of homologous chromosomes by a proteinaceous scaffold structure known as the synaptonemal complex (SC). Recent studies in yeast and worms have indicated that polo-like kinases (PLKs) regulate several events during meiosis, including DNA recombination and SC dynamics. Mammals express four active PLKs (PLK1–4), and our previous work assessing localization and kinase function in mouse spermatocytes suggested that PLK1 coordinates nuclear events during meiotic prophase. Therefore, we conditionally mutated Plk1 in early prophase spermatocytes and assessed stages of HR, crossover formation, and SC processes. Plk1 mutation resulted in increased RPA foci and reduced RAD51/DMC1 foci during zygonema, and an increase of both class I and class II crossover events. Furthermore, the disassembly of SC lateral elements was aberrant. Our results highlight the importance of PLK1 in regulating HR and SC disassembly during spermatogenesis.

The synaptonemal complex (SC) is a zipper-like protein structure that forms between paired homologous chromosomes during meiosis. Hurlock et al. discover two novel SC components, SYP-5 and SYP-6, in C. elegans and investigate their contributions in both limiting and promoting crossover formation.

During meiosis, homologous chromosomes exchange genetic material. This exchange or meiotic recombination is mediated by a proteinaceous scaffold known as the Synaptonemal complex (SC). Any defects in its formation produce failures in meiotic recombination, chromosome segregation and meiosis completion. It has been proposed that DNA repair events that will be resolved by crossover between homologous chromosomes are predetermined by the SC. Hence, structural analysis of the organization of the DNA in the SC could shed light on the process of crossover interference. In this work, we employed an ultrastructural DNA staining technique on mouse testis and followed nuclei of pachytene cells. We observed structures organized similarly to the SCs stained with conventional techniques. These structures, presumably the DNA in the SCs, are delineating the edges of both lateral elements and no staining was observed between them. DNA in the LEs resembles two parallel tracks. However, a bubble-like staining pattern in certain regions of the SC was observed. Furthermore, this staining pattern is found in SCs formed between non-homologous chromosomes, in SCs formed between sister chromatids and in SCs without lateral elements, suggesting that this particular organization of the DNA is determined by the synapsis of the chromosomes despite their lack of homology or the presence of partially formed SCs.

During meiosis, pairing between homologous chromosomes is stabilized by the assembly of a protein lattice known as the synaptonemal complex (SC). The SC ensures the formation of crossovers between homologous chromosomes and also regulates their distribution. However, how the SC regulates crossover formation remains elusive. We isolated an unusual mutation in C. elegans that disrupts crossover interference but not the assembly of the SC. This mutation alters the unique C-terminal domain of an essential SC protein, SYP-4, a likely ortholog of the vertebrate SC protein SIX6OS1. To characterize the structure of the SC in wild-type and mutant animals, we use three-dimensional STochastic Optical Reconstruction Microscopy (3D-STORM) to interrogate the molecular architecture of the SC in intact germline tissue from C. elegans. The approach enabled us to define positions of protein epitopes with respect to the 3D architecture of this complex. Using a probabilistic mapping approach to analyze super-resolution image data, we detect a marked structural transition in wild-type animals that coincides with crossover designation. We also found that our syp-4 mutant subtly perturbs SC architecture. Our findings add to growing evidence that the SC is an active material whose molecular organization contributes to chromosome-wide crossover regulation.

No abstract available