Meiotic Double-strand Break Formation Research Articles

Genetic and physical interactions reveal overlapping and distinct contributions to meiotic double-strand break formation in C. elegans.

Double-strand breaks (DSBs) are the most deleterious lesions experienced by our genome. Yet, DSBs are intentionally induced during gamete formation to promote the exchange of genetic material between homologous chromosomes. While the conserved topoisomerase-like enzyme Spo11 catalyzes DSBs, additional regulatory proteins-referred to as "Spo11 accessory factors"- regulate the number, timing, and placement of DSBs during early meiotic prophase ensuring that SPO11 does not wreak havoc on the genome. Despite the importance of the accessory factors, they are poorly conserved at the sequence level suggesting that these factors may adopt unique functions in different species. In this work, we present a detailed analysis of the genetic and physical interactions between the DSB factors in the nematode Caenorhabditis elegans providing new insights into conserved and novel functions of these proteins. This work shows that HIM-5 is the determinant of X-chromosome-specific crossovers and that its retention in the nucleus is dependent on DSB-1, the sole accessory factor that interacts with SPO-11. We further provide evidence that HIM-5 coordinates the actions of the different accessory factors sub-groups, providing insights into how components on the DNA loops may interact with the chromosome axis.

Analysis of meiotic recombination in Drosophila simulans shows no evidence of an interchromosomal effect.

Chromosome inversions are of unique importance in the evolution of genomes and species because when heterozygous with a standard arrangement chromosome, they suppress meiotic crossovers within the inversion. In Drosophila species, heterozygous inversions also cause the interchromosomal effect, whereby the presence of a heterozygous inversion induces a dramatic increase in crossover frequencies in the remainder of the genome within a single meiosis. To date, the interchromosomal effect has been studied exclusively in species that also have high frequencies of inversions in wild populations. We took advantage of a recently developed approach for generating inversions in Drosophila simulans, a species that does not have inversions in wild populations, to ask if there is an interchromosomal effect. We used the existing chromosome 3R balancer and generated a new chromosome 2L balancer to assay for the interchromosomal effect genetically and cytologically. We found no evidence of an interchromosomal effect in D. simulans. To gain insights into the underlying mechanistic reasons, we qualitatively analyzed the relationship between meiotic double-stranded break (DSB) formation and synaptonemal complex (SC) assembly. We found that the SC is assembled prior to DSB formation as in D. melanogaster; however, we show that the SC is assembled prior to localization of the oocyte determination factor Orb, whereas in D. melanogaster, SC formation does not begin until the Orb is localized. Together, our data show no evidence that heterozygous inversions in D. simulans induce an interchromosomal effect and that there are differences in the developmental programming of the early stages of meiosis.

A Rad50-null mutation in mouse germ cells causes reduced DSB formation, abnormal DSB end resection and complete loss of germ cells.

The conserved MRE11-RAD50-NBS1/Xrs2 complex is crucial for DNA break metabolism and genome maintenance. Although hypomorphic Rad50 mutation mice showed normal meiosis, both null and hypomorphic rad50 mutation yeast displayed impaired meiosis recombination. However, the in vivo function of Rad50 in mammalian germ cells, particularly its in vivo role in the resection of meiotic double strand break (DSB) ends at the molecular level remains elusive. Here, we have established germ cell-specific Rad50 knockout mouse models to determine the role of Rad50 in mitosis and meiosis of mammalian germ cells. We find that Rad50-deficient spermatocytes exhibit defective meiotic recombination and abnormal synapsis. Mechanistically, using END-seq, we demonstrate reduced DSB formation and abnormal DSB end resection occurs in mutant spermatocytes. We further identify that deletion of Rad50 in gonocytes leads to complete loss of spermatogonial stem cells due to genotoxic stress. Taken together, our results reveal the essential role of Rad50 in mammalian germ cell meiosis and mitosis, and provide in vivo views of RAD50 function in meiotic DSB formation and end resection at the molecular level.

Chromatin remodeler CHD8 is required for spermatogonial proliferation and early meiotic progression.

Meiosis is a key step during germ cell differentiation, accompanied by the activation of thousands of genes through germline-specific chromatin reorganization. The chromatin remodeling mechanisms underpinning early meiotic stages remain poorly understood. Here we focus on the function of one of the major autism genes, CHD8, in spermatogenesis, based on the epidemiological association between autism and low fertility rates. Specific ablation of Chd8 in germ cells results in gradual depletion of undifferentiated spermatogonia and the failure of meiotic double-strand break (DSB) formation, leading to meiotic prophase I arrest and cell death. Transcriptional analyses demonstrate that CHD8 is required for extensive activation of spermatogenic genes in spermatogonia, necessary for spermatogonial proliferation and meiosis. CHD8 directly binds and regulates genes crucial for meiosis, including H3K4me3 histone methyltransferase genes, meiotic cohesin genes, HORMA domain-containing genes, synaptonemal complex genes, and DNA damage response genes. We infer that CHD8 contributes to meiotic DSB formation and subsequent meiotic progression through combined regulation of these meiosis-related genes. Our study uncovers an essential role of CHD8 in the proliferation of undifferentiated spermatogonia and the successful progression of meiotic prophase I.

Meiotic DNA double-strand break-independent role of protein phosphatase 4 in Hop1 assembly to promote meiotic chromosome axis formation in budding yeast.

Dynamic changes in chromosomal structure that occur during meiotic prophase play an important role in the progression of meiosis. Among them, meiosis-specific chromosomal axis-loop structures are important as a scaffold for integrated control between the meiotic recombination reaction and the associated checkpoint system to ensure accurate chromosome segregation. However, the molecular mechanism of the initial step of chromosome axis-loop construction is not well understood. Here, we showed that, in budding yeast, protein phosphatase 4 (PP4) that primarily counteracts Mec1/Tel1 phosphorylation is required to promote the assembly of a chromosomal axis component Hop1 and Red1 onto meiotic chromatin via interaction with Hop1. PP4, on the other hand, less affects Rec8 assembly. Notably, unlike the previously known function of PP4, this PP4 function in Hop1/Red1 assembly was independent of meiotic DSB-dependent Tel1/Mec1 kinase activities. The defect in Hop1/Red1 assembly in the absence of PP4 function was not suppressed by dysfunction of Pch2, which removes Hop1 protein from the chromosome axis, suggesting that PP4 is required for the initial step of chromatin loading of Hop1 rather than stabilization of Hop1 on axes. These results indicate phosphorylation/dephosphorylation-mediated regulation of Hop1 recruitment onto chromatin during chromosome axis construction before meiotic double-strand break formation.

OsPRD2 is essential for double-strand break formation, but not spindle assembly during rice meiosis.

Meiotic recombination starts with the programmed formation of double-strand breaks (DSB) in DNA, which are catalyzed by SPO11, a type II topoisomerase that is evolutionarily conserved, and several other accessary proteins. Homologs of MEIOSIS INHIBITOR 4 (MEI4/REC24/PRD2) are proteins that are also essential for the generation of meiotic DSBs in budding yeast, mice and Arabidopsis thaliana. In Arabidopsis, the protein ARABIDOPSIS THALIANA PUTATIVE RECOMBINATION INITIATION DEFECTS 2/MULTIPOLAR SPINDLE 1 (AtPRD2/MPS1) has been shown to have additional roles in spindle assembly, indicating a functional diversification. Here we characterize the role of the rice MEI4/PRD2 homolog in meiosis. The osprd2 mutant was completely male and female sterile. In male meiocytes of osprd2, no γH2AX foci were detected and twenty-four univalents were produced at diakinesis, suggesting that OsPRD2 is essential for DSB generation. OsPRD2 showed a dynamic localization during meiosis. For instance, OsPRD2 foci first appeared as discrete signals across chromosome at leptotene, and then became confined to the centromeres during zygotene, suggesting that they might be involved in assembly of the spindle. However we did not observe any obvious aberrant morphologies in neither the organization of the bipolar spindle nor in the orientation of the kinetochore in the mutant. These findings suggest that in rice PRD2 might not be required for spindle assembly and organization, as it does in Arabidopsis. Taken together our results indicate that plant MEI4/PRD2 homologs do play a conserved role in the formation of meiotic DSBs in DNA, but that their involvement in bipolar spindle assembly is rather species-specific.

Meiotic chromosome dynamics and double strand break formation in reptiles.

During meiotic prophase I, tightly regulated processes take place, from pairing and synapsis of homologous chromosomes to recombination, which are essential for the generation of genetically variable haploid gametes. These processes have canonical meiotic features conserved across different phylogenetic groups. However, the dynamics of meiotic prophase I in non-mammalian vertebrates are poorly known. Here, we compare four species from Sauropsida to understand the regulation of meiotic prophase I in reptiles: the Australian central bearded dragon (Pogona vitticeps), two geckos (Paroedura picta and Coleonyx variegatus) and the painted turtle (Chrysemys picta). We first performed a histological characterization of the spermatogenesis process in both the bearded dragon and the painted turtle. We then analyzed prophase I dynamics, including chromosome pairing, synapsis and the formation of double strand breaks (DSBs). We show that meiosis progression is highly conserved in reptiles with telomeres clustering forming the bouquet, which we propose promotes homologous pairing and synapsis, along with facilitating the early pairing of micro-chromosomes during prophase I (i.e., early zygotene). Moreover, we detected low levels of meiotic DSB formation in all taxa. Our results provide new insights into reptile meiosis.

ZmPRD1 is essential for double-strand break formation, but is not required for bipolar spindle assembly during maize meiosis.

Homologs of PUTATIVE RECOMBINATION INITIATION DEFECT 1 (PRD1) are known to be essential for meiotic double-strand break (DSB) formation in mouse (Mus musculus), Arabidopsis, and rice (Oryza sativa). Recent research has shown that rice PRD1 also plays an unanticipated role in meiotic bipolar spindle assembly, revealing that PRD1 has multiple functions in plant meiosis. In this study, we characterize the meiotic function of PRD1 in maize (Zea mays; ZmPRD1). Our results show that Zmprd1 mutant plants display normal vegetative growth but have complete male and female sterility. Meiotic DSB formation is fully abolished in mutant meiocytes, leading to failure in homologous pairing, synapsis, and recombination. ZmPRD1 exhibits a different pattern of chromosome localization compared to its rice homologs. The ZmPRD1 protein interacts with several DSB-forming proteins, but does not directly interact with the kinetochore proteins REC8 and SGO1. Possibly as a result of this, there are no significant abnormalities of bipolar spindle assembly in Zmprd1 meiocytes. Overall, our results demonstrate that ZmPRD1 is essential for DSB formation and homologous recombination in maize meiosis. However, the recently-identified function of PRD1 in bipolar spindle assembly during rice meiosis is not conserved in maize.

Pathogenic variants of meiotic double strand break (DSB) formation genes PRDM9 and ANKRD31 in premature ovarian insufficiency

PurposeThe etiology of premature ovarian insufficiency (POI) is heterogeneous, and genetic factors account for 20–25% of the patients. The primordial follicle pool is determined by the meiosis process, which is initiated by programmed DNA double strand breaks (DSB) and homologous recombination. The objective of the study is to explore the role of DSB formation genes in POI pathogenesis.MethodsVariants in DSB formation genes were analyzed from a database of exome sequencing in 1,030 patients with POI. The pathogenic effects of the potentially causative variants were verified by further functional studies.ResultsThree pathogenic heterozygous variants in PRDM9 and two in ANKRD31 were identified in seven patients. Functional studies showed the variants in PRDM9 impaired its methyltransferase activity, and the ANKRD31 variations disturbed its interaction with another DSB formation factor REC114 by haploinsufficiency effect, indicating the pathogenic effects of the two genes on ovarian function were dosage dependent.ConclusionOur study identified pathogenic variants of PRDM9 and ANKRD31 in POI patients, shedding new light on the contribution of meiotic DSB formation genes in ovarian development, further expanding the genetic architecture of POI.

The synaptonemal complex central region modulates crossover pathways and feedback control of meiotic double-strand break formation.

The synaptonemal complex (SC) is a proteinaceous structure that mediates homolog engagement and genetic recombination during meiosis. In budding yeast, Zip-Mer-Msh (ZMM) proteins promote crossover (CO) formation and initiate SC formation. During SC elongation, the SUMOylated SC component Ecm11 and the Ecm11-interacting protein Gmc2 facilitate the polymerization of Zip1, an SC central region component. Through physical recombination, cytological, and genetic analyses, we found that ecm11 and gmc2 mutants exhibit chromosome-specific defects in meiotic recombination. CO frequencies on a short chromosome (chromosome III) were reduced, whereas CO and non-crossover frequencies on a long chromosome (chromosome VII) were elevated. Further, in ecm11 and gmc2 mutants, more double-strand breaks (DSBs) were formed on a long chromosome during late prophase I, implying that the Ecm11–Gmc2 (EG) complex is involved in the homeostatic regulation of DSB formation. The EG complex may participate in joint molecule (JM) processing and/or double-Holliday junction resolution for ZMM-dependent CO-designated recombination. Absence of the EG complex ameliorated the JM-processing defect in zmm mutants, suggesting a role for the EG complex in suppressing ZMM-independent recombination. Our results suggest that the SC central region functions as a compartment for sequestering recombination-associated proteins to regulate meiosis specificity during recombination.

The molecular control of meiotic double-strand break (DSB) formation and its significance in human infertility.

Meiosis is an essential step in gametogenesis which is the key process in sexually reproducing organisms as meiotic aberrations may result in infertility. In meiosis, programmed DNA double-strand break (DSB) formation is one of the fundamental processes that are essential for maintaining homolog interactions and correcting segregation of chromosomes. Although the number and distribution of meiotic DSBs are tightly regulated, still abnormalities in DSB formation are known to cause meiotic arrest and infertility. This review is a detailed account of molecular bases of meiotic DSB formation, its evolutionary conservation, and variations in different species. We further reviewed the mutations of DSB formation genes in association with human infertility and also proposed the future directions and strategies about the study of meiotic DSB formation.

ZmMTOPVIB Enables DNA Double-Strand Break Formation and Bipolar Spindle Assembly during Maize Meiosis.

The meiotic TopoVI B subunit (MTopVIB) plays an essential role in double-strand break formation in mouse (Mus musculus), Arabidopsis (Arabidopsis thaliana), and rice (Oryza sativa), and recent work reveals that rice MTopVIB also plays an unexpected role in meiotic bipolar spindle assembly, highlighting multiple functions of MTopVIB during rice meiosis. In this work, we characterized the meiotic TopVIB in maize (Zea mays; ZmMTOPVIB). The ZmmtopVIB mutant plants exhibited normal vegetative growth but male and female sterility. Meiotic double-strand break formation was abolished in mutant meiocytes. Despite normal assembly of axial elements, mutants showed severely affected synapsis and disrupted homologous pairing. Importantly, we showed that bipolar spindle assembly was also affected in ZmmtopVIB, resulting in triad and polyad formation. Overall, our results demonstrate that ZmMTOPVIB plays critical roles in double-strand break formation and homologous recombination. In addition, our results suggest that the function of MTOPVIB in bipolar spindle assembly is likely conserved across different monocots.

Genetic Interactions of Histone Modification Machinery Set1 and PAF1C with the Recombination Complex Rec114-Mer2-Mei4 in the Formation of Meiotic DNA Double-Strand Breaks.

Homologous recombination is essential for chromosome segregation during meiosis I. Meiotic recombination is initiated by the introduction of double-strand breaks (DSBs) at specific genomic locations called hotspots, which are catalyzed by Spo11 and its partners. DSB hotspots during meiosis are marked with Set1-mediated histone H3K4 methylation. The Spo11 partner complex, Rec114-Mer2-Mei4, essential for the DSB formation, localizes to the chromosome axes. For efficient DSB formation, a hotspot with histone H3K4 methylation on the chromatin loops is tethered to the chromosome axis through the H3K4 methylation reader protein, Spp1, on the axes, which interacts with Mer2. In this study, we found genetic interaction of mutants in a histone modification protein complex called PAF1C with the REC114 and MER2 in the DSB formation in budding yeast Saccharomyces cerevisiae. Namely, the paf1c mutations rtf1 and cdc73 showed synthetic defects in meiotic DSB formation only when combined with a wild-type-like tagged allele of either the REC114 or MER2. The synthetic defect of the tagged REC114 allele in the DSB formation was seen also with the set1, but not with spp1 deletion. These results suggest a novel role of histone modification machinery in DSB formation during meiosis, which is independent of Spp1-mediated loop-axis tethering.

Causative Mutations and Mechanism of Androgenetic Hydatidiform Moles

Androgenetic complete hydatidiform moles are human pregnancies with no embryos and affect 1 in every 1,400 pregnancies. They have mostly androgenetic monospermic genomes with all the chromosomes originating from a haploid sperm and no maternal chromosomes. Androgenetic complete hydatidiform moles were described in 1977, but how they occur has remained an open question. We identified bi-allelic deleterious mutations in MEI1, TOP6BL/C11orf80, and REC114, with roles in meiotic double-strand breaks formation in women with recurrent androgenetic complete hydatidiform moles. We investigated the occurrence of androgenesis in Mei1-deficient female mice and discovered that 8% of their oocytes lose all their chromosomes by extruding them with the spindles into the first polar body. We demonstrate that Mei1-/- oocytes are capable of fertilization and 5% produce androgenetic zygotes. Thus, we uncover a meiotic abnormality in mammals and a mechanism for the genesis of androgenetic zygotes that is the extrusion of all maternal chromosomes and their spindles into the first polar body.

Pericentromere-Specific Cohesin Complex Prevents Meiotic Pericentric DNA Double-Strand Breaks and Lethal Crossovers

In most eukaryotes, meiotic crossovers are essential for error-free chromosome segregation but are specifically repressed near centromeres to prevent missegregation. Recognized for >85 years, the molecular mechanism of this repression has remained unknown. Meiotic chromosomes contain two distinct cohesin complexes: pericentric complex (for segregation) and chromosomal arm complex (for crossing over). We show that the pericentric-specific complex also actively represses pericentric meiotic double-strand break (DSB) formation and, consequently, crossovers. We uncover the mechanism by which fission yeast heterochromatin protein Swi6 (mammalian HP1-homolog) prevents recruitment of activators of meiotic DSB formation. Localizing missing activators to wild-type pericentromeres bypasses repression and generates abundant crossovers but reduces gamete viability. The molecular mechanism elucidated here likely extends to other species, including humans, where pericentric crossovers can result in disorders, such as Down syndrome. These mechanistic insights provide new clues to understand the roles played by multiple cohesin complexes, especially in human infertility and birth defects.

Interdependent and separable functions of Caenorhabditis elegans MRN-C complex members couple formation and repair of meiotic DSBs

Faithful inheritance of genetic information through sexual reproduction relies on the formation of crossovers between homologous chromosomes during meiosis, which, in turn, relies on the formation and repair of numerous double-strand breaks (DSBs). As DSBs pose a potential threat to the genome, mechanisms that ensure timely and error-free DSB repair are crucial for successful meiosis. Here, we identify NBS-1, the Caenorhabditis elegans ortholog of the NBS1 (mutated in Nijmegen Breakage Syndrome) subunit of the conserved MRE11-RAD50-NBS1/Xrs2 (MRN) complex, as a key mediator of DSB repair via homologous recombination (HR) during meiosis. Loss of nbs-1 leads to severely reduced loading of recombinase RAD-51, ssDNA binding protein RPA, and pro-crossover factor COSA-1 during meiotic prophase progression; aggregated and fragmented chromosomes at the end of meiotic prophase; and 100% progeny lethality. These phenotypes reflect a role for NBS-1 in processing of meiotic DSBs for HR that is shared with its interacting partners MRE-11-RAD-50 and COM-1 (ortholog of Com1/Sae2/CtIP). Unexpectedly, in contrast to MRE-11 and RAD-50, NBS-1 is not required for meiotic DSB formation. Meiotic defects of the nbs-1 mutant are partially suppressed by abrogation of the nonhomologous end-joining (NHEJ) pathway, indicating a role for NBS-1 in antagonizing NHEJ during meiosis. Our data further reveal that NBS-1 and COM-1 play distinct roles in promoting HR and antagonizing NHEJ. We propose a model in which different components of the MRN-C complex work together to couple meiotic DSB formation with efficient and timely engagement of HR, thereby ensuring crossover formation and restoration of genome integrity before the meiotic divisions.

The PHD finger protein Spp1 has distinct functions in the Set1 and the meiotic DSB formation complexes.

Histone H3K4 methylation is a feature of meiotic recombination hotspots shared by many organisms including plants and mammals. Meiotic recombination is initiated by programmed double-strand break (DSB) formation that in budding yeast takes place in gene promoters and is promoted by histone H3K4 di/trimethylation. This histone modification is recognized by Spp1, a PHD finger containing protein that belongs to the conserved histone H3K4 methyltransferase Set1 complex. During meiosis, Spp1 binds H3K4me3 and interacts with a DSB protein, Mer2, to promote DSB formation close to gene promoters. How Set1 complex- and Mer2- related functions of Spp1 are connected is not clear. Here, combining genome-wide localization analyses, biochemical approaches and the use of separation of function mutants, we show that Spp1 is present within two distinct complexes in meiotic cells, the Set1 and the Mer2 complexes. Disrupting the Spp1-Set1 interaction mildly decreases H3K4me3 levels and does not affect meiotic recombination initiation. Conversely, the Spp1-Mer2 interaction is required for normal meiotic recombination initiation, but dispensable for Set1 complex-mediated histone H3K4 methylation. Finally, we provide evidence that Spp1 preserves normal H3K4me3 levels independently of the Set1 complex. We propose a model where Spp1 works in three ways to promote recombination initiation: first by depositing histone H3K4 methylation (Set1 complex), next by “reading” and protecting histone H3K4 methylation, and finally by making the link with the chromosome axis (Mer2-Spp1 complex). This work deciphers the precise roles of Spp1 in meiotic recombination and opens perspectives to study its functions in other organisms where H3K4me3 is also present at recombination hotspots.

An in vivo genetic screen in Drosophila identifies the orthologue of human cancer/testis gene SPO11 among a network of targets to inhibit lethal(3)malignant brain tumour growth.

Using transgenic RNAi technology, we have screened over 4000 genes to identify targets to inhibit malignant growth caused by the loss of function of lethal(3)malignant brain tumour in Drosophila in vivo. We have identified 131 targets, which belong to a wide range of gene ontologies. Most of these target genes are not significantly overexpressed in mbt tumours hence showing that, rather counterintuitively, tumour-linked overexpression is not a good predictor of functional requirement. Moreover, we have found that most of the genes upregulated in mbt tumours remain overexpressed in tumour-suppressed double-mutant conditions, hence revealing that most of the tumour transcriptome signature is not necessarily correlated with malignant growth. One of the identified target genes is meiotic W68 (mei-W68), the Drosophila orthologue of the human cancer/testis gene Sporulation-specific protein 11 (SPO11), the enzyme that catalyses the formation of meiotic double-strand breaks. We show that Drosophila mei-W68/SPO11 drives oncogenesis by causing DNA damage in a somatic tissue, hence providing the first instance in which a SPO11 orthologue is unequivocally shown to have a pro-tumoural role. Altogether, the results from this screen point to the possibility of investigating the function of human cancer relevant genes in a tractable experimental model organism like Drosophila.

The PRDM9 KRAB domain is required for meiosis and involved in protein interactions

PR domain-containing protein 9 (PRDM9) is a major regulator of the localization of meiotic recombination hotspots in the human and mouse genomes. This role involves its DNA-binding domain, which is composed of a tandem array of zinc fingers, and PRDM9-dependent trimethylation of histone H3 at lysine 4. PRDM9 is a member of the PRDM family of transcription regulators, but unlike other family members, it contains a Krüppel-associated box (KRAB)-related domain that is predicted to be a potential protein interaction domain. Here, we show that truncation of the KRAB domain of mouse PRDM9 leads to loss of PRDM9 function and altered meiotic prophase and gametogenesis. In addition, we identified proteins that interact with the KRAB domain of PRDM9 in yeast two-hybrid assay screens, particularly CXXC1, a member of the COMPASS complex. We also show that CXXC1 interacts with IHO1, an essential component of the meiotic double-strand break (DSB) machinery. As CXXC1 is orthologous to Saccharomyces cerevisiae Spp1 that links DSB sites to the DSB machinery on the chromosome axis, we propose that these molecular interactions involved in the regulation of meiotic DSB formation are conserved in mouse meiosis.

The Landscape of Mouse Meiotic Double-Strand Break Formation, Processing, and Repair

Heritability and genome stability are shaped by meiotic recombination, which is initiated via hundreds of DNA double-strand breaks (DSBs). The distribution of DSBs throughout the genome is notrandom, but mechanisms molding this landscaperemain poorly understood. Here, we exploit genome-wide maps of mouse DSBs at unprecedented nucleotide resolution to uncover previously invisible spatial features of recombination. At fine scale, we reveal a stereotyped hotspot structure-DSBs occur within narrow zones between methylated nucleosomes-and identify relationships between SPO11, chromatin, and the histone methyltransferase PRDM9. At large scale, DSB formation issuppressed on non-homologous portions of the sex chromosomes via the DSB-responsive kinase ATM, which also shapes the autosomal DSB landscape at multiple size scales. We also provide a genome-wide analysis of exonucleolytic DSB resection lengths and elucidate spatial relationships between DSBs and recombination products. Our results paint a comprehensive picture of features governing successive steps in mammalian meiotic recombination.