Recombinase Dmc1 Research Articles

Hop2-Mnd1 functions as a DNA sequence fidelity switch in Dmc1-mediated DNA recombination

Homologous recombination during meiosis is critical for chromosome segregation and also gives rise to genetic diversity. Genetic exchange between homologous chromosomes during meiosis is mediated by the recombinase Dmc1, which is capable of recombining DNA sequences with mismatches. The Hop2-Mnd1 complex mediates Dmc1 activity. Here, we reveal a regulatory role for Hop2-Mnd1 in restricting substrate selection. Specifically, Hop2-Mnd1 upregulates Dmc1 activity with DNA substrates that are either fully homologous or contain DNA mismatches, and it also acts against DNA strand exchange between substrates solely harboring microhomology. By isolating and examining salient Hop2-Mnd1 separation-of-function variants, we show that suppressing illegitimate DNA recombination requires the Dmc1 filament interaction attributable to Hop2-Mnd1 but not its DNA binding activity. Our study provides mechanistic insights into how Hop2-Mnd1 helps maintain meiotic recombination fidelity.

BRCA2 stabilises RAD51 and DMC1 nucleoprotein filaments through a conserved interaction mode

BRCA2 is essential for DNA repair by homologous recombination in mitosis and meiosis. It interacts with recombinases RAD51 and DMC1 to facilitate the formation of nucleoprotein filaments on resected DNA ends that catalyse recombination-mediated repair. BRCA2’s BRC repeats bind and disrupt RAD51 and DMC1 filaments, whereas its PhePP motifs bind recombinases and stabilise their nucleoprotein filaments. However, the mechanism of filament stabilisation has hitherto remained unknown. Here, we report the crystal structure of a BRCA2-DMC1 complex, revealing how core interaction sites of PhePP motifs bind to recombinases. The interaction mode is conserved for RAD51 and DMC1, which selectively bind to BRCA2’s two distinct PhePP motifs via subtly divergent binding pockets. PhePP motif sequences surrounding their core interaction sites protect nucleoprotein filaments from BRC-mediated disruption. Hence, we report the structural basis of how BRCA2’s PhePP motifs stabilise RAD51 and DMC1 nucleoprotein filaments for their essential roles in mitotic and meiotic recombination.

DNA cytosine methyltransferases differentially regulate genome-wide hypermutation and interhomolog recombination in Trichoderma reesei meiosis.

Trichoderma reesei is an economically important enzyme producer with several unique meiotic features. spo11, the initiator of meiotic double-strand breaks (DSBs) in most sexual eukaryotes, is dispensable for T. reesei meiosis. T. reesei lacks the meiosis-specific recombinase Dmc1. Rad51 and Sae2, the activator of the Mre11 endonuclease complex, promote DSB repair and chromosome synapsis in wild-type and spo11Δ meiosis. DNA methyltransferases (DNMTs) perform multiple tasks in meiosis. Three DNMT genes (rid1, dim2and dimX) differentially regulate genome-wide cytosine methylation and C:G-to-T:A hypermutations in different chromosomal regions. We have identified two types of DSBs: type I DSBs require spo11 or rid1 for initiation, whereas type II DSBs do not rely on spo11 and rid1 for initiation. rid1 (but not dim2) is essential for Rad51-mediated DSB repair and normal meiosis. rid1 and rad51 exhibit a locus heterogeneity (LH) relationship, in which LH-associated proteins often regulate interconnectivity in protein interaction networks. This LH relationship can be suppressed by deleting dim2 in a haploid rid1Δ (but not rad51Δ) parental strain, indicating that dim2 and rid1 share a redundant function that acts earlier than rad51 during early meiosis. In conclusion, our studies provide the first evidence of the involvement of DNMTs during meiotic initiation and recombination.

Mutational analysis of Mei5, a subunit of Mei5-Sae3 complex, in Dmc1-mediated recombination during yeast meiosis.

Interhomolog recombination in meiosis is mediated by the Dmc1 recombinase. The Mei5-Sae3 complex of Saccharomyces cerevisiae promotes Dmc1 assembly and functions with Dmc1 for homology-mediated repair of meiotic DNA double-strand breaks. How Mei5-Sae3 facilitates Dmc1 assembly remains poorly understood. In this study, we created and characterized several mei5 mutants featuring the amino acid substitutions of basic residues. We found that Arg97 of Mei5, conserved in its ortholog, SFR1 (complex with SWI5), RAD51 mediator, in humans and other organisms, is critical for complex formation with Sae3 for Dmc1 assembly. Moreover, the substitution of either Arg117 or Lys133 with Ala in Mei5 resulted in the production of a C-terminal truncated Mei5 protein during yeast meiosis. Notably, the shorter Mei5-R117A protein was observed in meiotic cells but not in mitotic cells when expressed, suggesting a unique regulation of Dmc1-mediated recombination by posttranslational processing of Mei5-Sae3.

The Dmc1 recombinase physically interacts with and promotes the meiotic crossover functions of the Mlh1-Mlh3 endonuclease.

The accurate segregation of homologous chromosomes during the Meiosis I reductional division in most sexually reproducing eukaryotes requires crossing over between homologs. In baker's yeast approximately 80% of meiotic crossovers result from Mlh1-Mlh3 and Exo1 acting to resolve double-Holliday junction intermediates in a biased manner. Little is known about how Mlh1-Mlh3 is recruited to recombination intermediates to perform its role in crossover resolution. We performed a gene dosage screen in baker's yeast to identify novel genetic interactors with Mlh1-Mlh3. Specifically, we looked for genes whose lowered dosage reduced meiotic crossing over using sensitized mlh3 alleles that disrupt the stability of the Mlh1-Mlh3 complex and confer defects in mismatch repair but do not disrupt meiotic crossing over. To our surprise we identified genetic interactions between MLH3 and DMC1, the recombinase responsible for recombination between homologous chromosomes during meiosis. We then showed that Mlh3 physically interacts with Dmc1 in vitro and in vivo. Partial complementation of Mlh3 crossover functions was observed when MLH3 was expressed under the control of the CLB1 promoter (NDT80 regulon), suggesting that Mlh3 function can be provided late in meiotic prophase at some functional cost. A model for how Dmc1 could facilitate Mlh1-Mlh3's role in crossover resolution is presented.

Recombinase mediator Mei5-Sae3 stimulates Dmc1 recombinase assembly on RPA-coated ssDNA by stabilizing nucleation clusters

Recombinase mediator Mei5-Sae3 stimulates Dmc1 recombinase assembly on RPA-coated ssDNA by stabilizing nucleation clusters

BRCA2-HSF2BP oligomeric ring disassembly by BRME1 promotes homologous recombination.

In meiotic homologous recombination (HR), BRCA2 facilitates loading of the recombinases RAD51 and DMC1 at the sites of double-strand breaks (DSBs). The HSF2BP-BRME1 complex interacts with BRCA2. Its absence causes a severe reduction in recombinase loading at meiotic DSB. We previously showed that, in somatic cancer cells ectopically producing HSF2BP, DNA damage can trigger HSF2BP-dependent degradation of BRCA2, which prevents HR. Here, we report that, upon binding to BRCA2, HSF2BP forms octameric rings that are able to interlock into a large ring-shaped 24-nucleotide oligomer. Addition of BRME1 leads to dissociation of both of these ring structures and cancels the disruptive effect of HSF2BP on cancer cell resistance to DNA damage. It also prevents BRCA2 degradation during interstrand DNA crosslink repair in Xenopus egg extracts. We propose that, during meiosis, the control of HSF2BP-BRCA2 oligomerization by BRME1 ensures timely assembly of the ring complex that concentrates BRCA2 and controls its turnover, thus promoting HR.

SCFRMF mediates degradation of the meiosis-specific recombinase DMC1

Meiotic recombination requires the specific RecA homolog DMC1 recombinase to stabilize strand exchange intermediates in most eukaryotes. Normal DMC1 levels are crucial for its function, yet the regulatory mechanisms of DMC1 stability are unknown in any organism. Here, we show that the degradation of Arabidopsis DMC1 by the 26S proteasome depends on F-box proteins RMF1/2-mediated ubiquitination. Furthermore, RMF1/2 interact with the Skp1 ortholog ASK1 to form the ubiquitin ligase complex SCFRMF1/2. Genetic analyses demonstrate that RMF1/2, ASK1 and DMC1 act in the same pathway downstream of SPO11-1 dependent meiotic DNA double strand break formation and that the proper removal of DMC1 is crucial for meiotic crossover formation. Moreover, six DMC1 lysine residues were identified as important for its ubiquitination but not its interaction with RMF1/2. Our results reveal mechanistic insights into how the stability of a key meiotic recombinase that is broadly conserved in eukaryotes is regulated.

FIGL1 coordinates with dosage-sensitive BRCA2 in modulating meiotic recombination in maize.

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.

Hop2-Mnd1 and Swi5-Sfr1 stimulate Dmc1 filament assembly using distinct mechanisms.

In meiosis, Dmc1 recombinase and the general recombinase Rad51 are responsible for pairing homologous chromosomes and exchanging strands. Fission yeast (Schizosaccharomyces pombe) Swi5-Sfr1 and Hop2-Mnd1 stimulate Dmc1-driven recombination, but the stimulation mechanism is unclear. Using single-molecule fluorescence resonance energy transfer (smFRET) and tethered particle motion (TPM) experiments, we showed that Hop2-Mnd1 and Swi5-Sfr1 individually enhance Dmc1 filament assembly on single-stranded DNA (ssDNA) and adding both proteins together allows further stimulation. FRET analysis showed that Hop2-Mnd1 enhances the binding rate of Dmc1 while Swi5-Sfr1 specifically reduces the dissociation rate during the nucleation, about 2-fold. In the presence of Hop2-Mnd1, the nucleation time of Dmc1 filaments shortens, and doubling the ss/double-stranded DNA (ss/dsDNA) junctions of DNA substrates reduces the nucleation times in half. Order of addition experiments confirmed that Hop2-Mnd1 binds on DNA to recruit and stimulate Dmc1 nucleation at the ss/dsDNA junction. Our studies directly support the molecular basis of how Hop2-Mnd1 and Swi5-Sfr1 act on different steps during the Dmc1 filament assembly. DNA binding of these accessory proteins and nucleation preferences of recombinases thus dictate how their regulation can take place.

SPIDR is required for homologous recombination during mammalian meiosis.

Meiotic recombinases RAD51 and DMC1 mediate strand exchange in the repair of DNA double-strand breaks (DSBs) by homologous recombination. This is a landmark event of meiosis that ensures genetic diversity in sexually reproducing organisms. However, the regulatory mechanism of DMC1/RAD51-ssDNA nucleoprotein filaments during homologous recombination in mammals has remained largely elusive. Here, we show that SPIDR (scaffold protein involved in DNA repair) regulates the assembly or stability of RAD51/DMC1 on ssDNA. Knockout of Spidr in male mice causes complete meiotic arrest, accompanied by defects in synapsis and crossover formation, which leads to male infertility. In females, loss of Spidr leads to subfertility; some Spidr-/- oocytes are able to complete meiosis. Notably, fertility is rescued partially by ablation of the DNA damage checkpoint kinase CHK2 in Spidr-/- females but not in males. Thus, our study identifies SPIDR as an essential meiotic recombination factor in homologous recombination in mammals.

A cytological revisit on parthenogenetic Artemia and the deficiency of a meiosis-specific recombinase DMC1 in the possible transition from bisexuality to parthenogenesis.

Although parthenogenesis is widespread in nature and known to have close relationships with bisexuality, the transitional mechanism is poorly understood. Artemia is an ideal model to address this issue because bisexuality and "contagious" obligate parthenogenesis independently exist in its congeneric members. In the present study, we first performed chromosome spreading and immunofluorescence to compare meiotic processes of Artemia adopting two distinct reproductive ways. The results showed that, unlike conventional meiosis in bisexual Artemia, meiosis II in parthenogenic Artemia is entirely absent and anaphase I is followed by a single mitosis-like equational division. Interspecific comparative transcriptomics showed that two central molecules in homologous recombination (HR), Dmc1 and Rad51, exhibited significantly higher expression in bisexual versus parthenogenetic Artemia. qRT-PCR indicated that the expression of both genes peaked at the early oogenesis and gradually decreased afterward. Knocking-down by RNAi of Dmc1 in unfertilized females of bisexual Artemia resulted in a severe deficiency of homologous chromosome pairing and produced univalents at the middle oogenesis stage, which was similar to that of parthenogenic Artemia, while in contrast, silencing Rad51 led to no significant chromosome morphological change. Our results indicated that Dmc1 is vital for HR in bisexual Artemia, and the deficiency of Dmc1 may be correlated with or even possibly one of core factors in the transition from bisexuality to parthenogenesis.

Three-color dSTORM Imaging and Analysis of Recombination Foci in Mouse Spread Meiotic Nuclei.

During the first meiotic prophase in mouse, repair of SPO11-induced DNA double-strand breaks (DSBs), facilitating homologous chromosome synapsis, is essential to successfully complete the first meiotic cell division. Recombinases RAD51 and DMC1 play an important role in homology search, but their mechanistic contribution to this process is not fully understood. Super-resolution, single-molecule imaging of RAD51 and DMC1 provides detailed information on recombinase accumulation on DSBs during meiotic prophase. Here, we present a detailed protocol of recombination foci analysis of three-color direct stochastic optical reconstruction microscopy (dSTORM) imaging of SYCP3, RAD51, and DMC1, fluorescently labeled by antibody staining in mouse spermatocytes. This protocol consists of sample preparation, data acquisition, pre-processing, and data analysis. The sample preparation procedure includes an updated version of the nuclear spreading of mouse testicular cells, followed by immunocytochemistry and the preparation steps for dSTORM imaging. Data acquisition consists of three-color dSTORM imaging, which is extensively described. The pre-processing that converts fluorescent signals to localization data also includes channel alignment and image reconstruction, after which regions of interest (ROIs) are identified based on RAD51 and/or DMC1 localization patterns. The data analysis steps then require processing of the fluorescent signal localization within these ROIs into discrete nanofoci, which can be further analyzed. This multistep approach enables the systematic investigation of spatial distributions of proteins associated with individual DSB sites and can be easily adapted for analyses of other foci-forming proteins. All computational scripts and software are freely accessible, making them available to a broad audience. Key features Preparation of spread nuclei, resulting in a flattened preparation with easy antibody-accessible chromatin-associated proteins on dSTORM-compatible coverslips. dSTORM analysis of immunofluorescent repair foci in meiotic prophase nuclei. Detailed descriptions of data acquisition, (pre-)processing, and nanofoci feature analysis applicable to all proteins that assemble in immunodetection as discrete foci. Graphical overview.

OsRAD51 Plays a Vital Role in Promoting Homologous Recombination in Rice Meiosis.

Meiotic recombination plays a pivotal role in achieving accurate chromosomal segregation and increasing genetic diversity. In the homologous recombination pathway, the detailed mechanisms of how OsRAD51 and OsDMC1 work in rice meiosis remain to be explored. Here, we obtained different types of mutants for Osrad51a1, Osrad51a2, Osdmc1a, and Osdmc1b through CRISPR/Cas9. Both Osrad51a1 and Osrad51a2 exhibited normal vegetative growth and fertility. Osrad51 (Osrad51a1 Osrad51a2) mutant plants show normal vegetative growth but exhibit complete sterility, indicating that OsRAD51A1 and OsRAD51A2 are functionally redundant in rice fertility. In contrast to the wild type, Osrad51 chromosomes are not paired perfectly at pachytene and synaptonemal complex (SC) formation is deficient. Moreover, univalents and multivalent associations were observed at metaphase I, chromosome fragments presented at anaphase I, and crossover formation is basically suppressed in Osrad51 pollen mother cells (PMCs). OsRAD51 foci emerge at leptotene and disappear from late pachytene and chromosome localization of OsRAD51 depends on the formation of double-strand breaks (DSBs). Most OsRAD51 foci can co-localize with OsDMC1 signals. OsRAD51 is essential for the loading of OsDMC1 onto chromosomes, and vice versa. In addition, both OsRAD51 and OsDMC1 can interact with OsFIGL1 and OsBRCA2, two important components in rice meiosis. Moreover, the Osrad51 Osdmc1 (Osrad51a1 Osrad51a2 Osdmc1a Osdmc1b) quadruple mutant PMCs exhibited similar defective phenotypes as Osrad51 in homologous pairing, synapsis, and DSB repair. Taken together, our results suggest that the recombinases DMC1 and RAD51 may functionally depend on each other and play important roles in meiotic recombination during meiosis in rice.

Repair of DNA double-strand breaks in plant meiosis: role of eukaryotic RecA recombinases and their modulators.

Homologous recombination during meiosis is crucial for the DNA double-strand breaks (DSBs) repair that promotes the balanced segregation of homologous chromosomes and enhances genetic variation. In most eukaryotes, two recombinases RAD51 and DMC1 form nucleoprotein filaments on single-stranded DNA generated at DSB sites and play a central role in the meiotic DSB repair and genome stability. These nucleoprotein filaments perform homology search and DNA strand exchange to initiate repair using homologous template-directed sequences located elsewhere in the genome. Multiple factors can regulate the assembly, stability, and disassembly of RAD51 and DMC1 nucleoprotein filaments. In this review, we summarize the current understanding of the meiotic functions of RAD51 and DMC1 and the role of their positive and negative modulators. We discuss the current models and regulators of homology searches and strand exchange conserved during plant meiosis. Manipulation of these repair factors during plant meiosis also holds a great potential to accelerate plant breeding for crop improvements and productivity.

Multiple Roles of SMC5/6 Complex during Plant Sexual Reproduction.

Chromatin-based processes are essential for cellular functions. Structural maintenance of chromosomes (SMCs) are evolutionarily conserved molecular machines that organize chromosomes throughout the cell cycle, mediate chromosome compaction, promote DNA repair, or control sister chromatid attachment. The SMC5/6 complex is known for its pivotal role during the maintenance of genome stability. However, a dozen recent plant studies expanded the repertoire of SMC5/6 complex functions to the entire plant sexual reproductive phase. The SMC5/6 complex is essential in meiosis, where its activity must be precisely regulated to allow for normal meiocyte development. Initially, it is attenuated by the recombinase RAD51 to allow for efficient strand invasion by the meiosis-specific recombinase DMC1. At later stages, it is essential for the normal ratio of interfering and non-interfering crossovers, detoxifying aberrant joint molecules, preventing chromosome fragmentation, and ensuring normal chromosome/sister chromatid segregation. The latter meiotic defects lead to the production of diploid male gametes in Arabidopsis SMC5/6 complex mutants, increased seed abortion, and production of triploid offspring. The SMC5/6 complex is directly involved in controlling normal embryo and endosperm cell divisions, and pioneer studies show that the SMC5/6 complex is also important for seed development and normal plant growth in cereals.

The Arabidopsis HOP2 gene has a role in preventing illegitimate connections between nonhomologous chromosome regions.

Meiotic homologous chromosomes synapse and undergo crossing over (CO). In many eukaryotes, both synapsis and crossing over require the induction of double stranded breaks (DSBs) and subsequent repair via homologous recombination. In these organisms, two key proteins are recombinases RAD51 and DMC1. Recombinase-modulators HOP2 and MND1 assist RAD51 and DMC1 and also are required for synapsis and CO. We have investigated the hop2-1 phenotype in Arabidopsis during the segregation stages of both meiosis and mitosis. Despite a general lack of synapsis during prophase I, we observed extensive, stable interconnections between nonhomologous chromosomes in diploid hop2-1 nuclei in first and second meiotic divisions. Using γH2Ax as a marker of unrepaired DSBs, we detected γH2AX foci from leptotene through early pachytene but saw no foci from mid-pachytene onward. We conclude that the bridges seen from metaphase I onward are due to mis-repaired DSBs, not unrepaired ones. Examining haploids, we found that wild type haploids produce only univalents, but hop2-1 haploids like hop2-1 diploids have illegitimate connections stable enough to produce bridged chromosomes during segregation. Our results suggest that HOP2 has a significant active role in preventing repairs that use nonhomologous chromosomes during meiosis. We also found evidence that HOP2 plays a role in preventing illegitimate repair of radiation-induced DSBs in rapidly dividing petal cells. We conclude that HOP2 in Arabidopsis plays both a positive role in promoting synapsis and a separable role in preventing DSB repair using nonhomologous chromosomes. SIGNIFICANCE STATEMENT : The fidelity of homologous recombination (HR) during meiosis is essential to the production of viable gametes and for maintaining genome integrity in vegetative cells. HOP2 is an important protein for accurate meiotic HR in plants. We have found evidence of high levels of illegitimate repairs between nonhomologous chromosomes during meiosis and in irradiated petal cells in hop2-1 mutants, suggesting a role for HOP2 beyond its established role in synapsis and crossing over.

Mechanisms of distinctive mismatch tolerance between Rad51 and Dmc1 in homologous recombination.

Homologous recombination (HR) is a primary DNA double-strand breaks (DSBs) repair mechanism. The recombinases Rad51 and Dmc1 are highly conserved in the RecA family; Rad51 is mainly responsible for DNA repair in somatic cells during mitosis while Dmc1 only works during meiosis in germ cells. This spatiotemporal difference is probably due to their distinctive mismatch tolerance during HR: Rad51 does not permit HR in the presence of mismatches, whereas Dmc1 can tolerate certain mismatches. Here, the cryo-EM structures of Rad51–DNA and Dmc1–DNA complexes revealed that the major conformational differences between these two proteins are located in their Loop2 regions, which contain invading single-stranded DNA (ssDNA) binding residues and double-stranded DNA (dsDNA) complementary strand binding residues, stabilizing ssDNA and dsDNA in presynaptic and postsynaptic complexes, respectively. By combining molecular dynamic simulation and single-molecule FRET assays, we identified that V273 and D274 in the Loop2 region of human RAD51 (hRAD51), corresponding to P274 and G275 of human DMC1 (hDMC1), are the key residues regulating mismatch tolerance during strand exchange in HR. This HR accuracy control mechanism provides mechanistic insights into the specific roles of Rad51 and Dmc1 in DNA double-strand break repair and may shed light on the regulatory mechanism of genetic recombination in mitosis and meiosis.

The Inactivation of Arabidopsis UBC22 Results in Abnormal Chromosome Segregation in Female Meiosis, but Not in Male Meiosis.

Protein ubiquitination is important for the regulation of meiosis in eukaryotes, including plants. However, little is known about the involvement of E2 ubiquitin-conjugating enzymes in plant meiosis. Arabidopsis UBC22 is a unique E2 enzyme, able to catalyze the formation of ubiquitin dimers through lysine 11 (K11). Previous work has shown that ubc22 mutants are defective in megasporogenesis, with most ovules having no or abnormally functioning megaspores; furthermore, some mutant plants show distinct phenotypes in vegetative growth. In this study, we showed that chromosome segregation and callose deposition were abnormal in mutant female meiosis while male meiosis was not affected. The meiotic recombinase DMC1, required for homologous chromosome recombination, showed a dispersed distribution in mutant female meiocytes compared to the presence of strong foci in WT female meiocytes. Based on an analysis of F1 plants produced from crosses using a mutant as the female parent, about 24% of female mutant gametes had an abnormal content of DNA, resulting in frequent aneuploids among the mutant plants. These results show that UBC22 is critical for normal chromosome segregation in female meiosis but not for male meiosis, and they provide important leads for studying the role of UBC22 and K11-linked ubiquitination.

Analysis of meiosis in Pristionchus pacificus reveals plasticity in homolog pairing and synapsis in the nematode lineage.

Meiosis is conserved across eukaryotes yet varies in the details of its execution. Here we describe a new comparative model system for molecular analysis of meiosis, the nematode Pristionchus pacificus, a distant relative of the widely studied model organism Caenorhabditis elegans. P. pacificus shares many anatomical and other features that facilitate analysis of meiosis in C. elegans. However, while C. elegans has lost the meiosis-specific recombinase Dmc1 and evolved a recombination-independent mechanism to synapse its chromosomes, P. pacificus expresses both DMC-1 and RAD-51. We find that SPO-11 and DMC-1 are required for stable homolog pairing, synapsis, and crossover formation, while RAD-51 is dispensable for these key meiotic processes. RAD-51 and DMC-1 localize sequentially to chromosomes during meiotic prophase and show nonoverlapping functions. We also present a new genetic map for P. pacificus that reveals a crossover landscape very similar to that of C. elegans, despite marked divergence in the regulation of synapsis and crossing-over between these lineages.