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During meiosis, homologous recombination repairs programmed DNA double-stranded breaks. Meiotic recombination physically links the homologous chromosomes (“homologs”), creating the tension between them that is required for their segregation. The central recombinase in this process is Dmc1. Dmc1’s activity is regulated by its accessory factors including the heterodimeric protein Mei5-Sae3 and Rad51. We use a gain-of-function dmc1 mutant, dmc1-E157D, that bypasses Mei5-Sae3 to gain insight into the role of this accessory factor and its relationship to mitotic recombinase Rad51, which also functions as a Dmc1 accessory protein during meiosis. We find that Mei5-Sae3 has a role in filament formation and stability, but not in the bias of recombination partner choice that favors homolog over sister chromatids. Analysis of meiotic recombination intermediates suggests that Mei5-Sae3 and Rad51 function independently in promoting filament stability. In spite of its ability to load onto single-stranded DNA and carry out recombination in the absence of Mei5-Sae3, recombination promoted by the Dmc1 mutant is abnormal in that it forms foci in the absence of DNA breaks, displays unusually high levels of multi-chromatid and intersister joint molecule intermediates, as well as high levels of ectopic recombination products. We use super-resolution microscopy to show that the mutant protein forms longer foci than those formed by wild-type Dmc1. Our data support a model in which longer filaments are more prone to engage in aberrant recombination events, suggesting that filament lengths are normally limited by a regulatory mechanism that functions to prevent recombination-mediated genome rearrangements., Author summary During meiosis, two rounds of division follow a single round of DNA replication to create the gametes for biparental reproduction. The first round of division requires that the homologous chromosomes become physically linked to one another to create the tension that is necessary for their segregation. This linkage is achieved through DNA recombination between the two homologous chromosomes, followed by resolution of the recombination intermediate into a crossover. Central to this process is the meiosis-specific recombinase Dmc1, and its accessory factors, which provide important regulatory functions to ensure that recombination is accurate, efficient, and occurs predominantly between homologous chromosomes, and not sister chromatids. To gain insight into the regulation of Dmc1 by its accessory factors, we mutated Dmc1 such that it was no longer dependent on its accessory factor Mei5-Sae3. Our analysis reveals that Dmc1 accessory factors Mei5-Sae3 and Rad51 have independent roles in stabilizing Dmc1 filaments. Furthermore, we find that although Rad51 is required for promoting recombination between homologous chromosomes, Mei5-Sae3 is not. Lastly, we show that our Dmc1 mutant forms abnormally long filaments, and high levels of aberrant recombination intermediates and products. These findings suggest that filaments are actively maintained at short lengths to prevent deleterious genome rearrangements.

Human chromosome 15q25 is involved in several disease-associated structural rearrangements, including microdeletions and chromosomal markers with inverted duplications. Using comparative fluorescence in situ hybridization, strand-sequencing, single-molecule, real-time sequencing and Bionano optical mapping analyses, we investigated the organization of the 15q25 region in human and nonhuman primates. We found that two independent inversions occurred in this region after the fission event that gave rise to phylogenetic chromosomes XIV and XV in humans and great apes. One of these inversions is still polymorphic in the human population today and may confer differential susceptibility to 15q25 microdeletions and inverted duplications. The inversion breakpoints map within segmental duplications containing core duplicons of the GOLGA gene family and correspond to the site of an ancestral centromere, which became inactivated about 25 million years ago. The inactivation of this centromere likely released segmental duplications from recombination repression typical of centromeric regions. We hypothesize that this increased the frequency of ectopic recombination creating a hotspot of hominid inversions where dispersed GOLGA core elements now predispose this region to recurrent genomic rearrangements associated with disease., Author summary Human chromosome 15 derived from a fission event that occurred in the ancestor of great apes. Following inactivation of the ancestral centromere at 15q25 a dispersal of segmental duplications took place, providing templates for ectopic recombination and predisposing the region to genomic instability. Different disease-associated microdeletions and chromosomal markers have been described with breakpoints mapping within these segmental duplications. To gain insight into the instability at 15q25, we sought to analyze this region in human and nonhuman primates using multiple genomics techniques and demonstrated the presence of two independent inversion events that occurred during great apes evolution. One of these inversions is still polymorphic in humans and may cause, in conjunction with a GOLGA core duplicon—a ~14 kbp chromosome 15 repeat, susceptibility to non-allelic homologous recombination leading to pathogenic recurrent rearrangements. Our results support the existence of a strong relationship between inversions and core duplicons and reinforce the hypothesis that GOLGA repeats play a fundamental role both in disease and evolution.

Meiosis distinguishes cells in the germ line from their somatic counterparts. Part of meiosis involves a bizarre series of events in which cells deliberately damage their DNA and then repair it. Of the many double-strand breaks that are generated, a minority are resolved as reciprocal exchanges between homologous chromosomes [1]. In a wide variety of species, recombination diversifies genomes and is required for the formation of viable gametes. Although the functional role of crossing over should impose strong selective constraints, the rate of recombination varies among individuals. Recombination rate shows resemblance among relatives [2], differs among inbred strains raised in a common environment [3], and responds to artificial selection [4], demonstrating a genetic component to individual rate differences that remains mostly unexplained. A handful of genes responsible for variation in recombination rate have been identified, including modifiers of the total number of crossovers [5–8] and loci that determine the genomic placement of recombination events [9]. As a fundamental part of gametogenesis, meiosis also plays a role in speciation, the means by which one species becomes two. A common observation is that otherwise viable hybrids between genetically differentiated lineages suffer from reduced fertility up to the point of complete sterility—an important reproductive barrier that keeps species distinct [10]. One stage during which problems appear is meiosis, when hybrid gametogenesis sometimes arrests. According to an influential model, hybrid sterility arises from disrupted interactions between different genes that evolved in separate populations [11,12]. The search for incompatibility genes that cause hybrid sterility has so far yielded a short list [13–15]. The fact that only a small number of genes are known to control variation in recombination rate or to contribute to hybrid sterility makes the discovery that one gene mediates both traits especially noteworthy. Prdm9 encodes a histone methyltransferase that modifies chromatin and is essential for meiosis in mice [16]. Decades of persistent effort from the group of Jiri Forejt demonstrated that incompatibility between Prdm9 and other loci causes sterility in F1 hybrid males formed by crossing two subspecies of house mice [17]. Subsequently, genetic mapping in crosses, population genetic analysis, bioinformatic prediction of DNA binding sites, and association studies jointly identified Prdm9 as a primary determinant of recombination hotspot location (short stretches of sequence within which most crossovers occur) in mice and humans [9,18–20]. The dual roles of Prdm9 reveal a tantalizing link between recombination and speciation at the genetic level and motivate the search for other genes that affect both processes. Now, progress from Jiri Forejt’s lab provides fresh evidence for a genetic connection between recombination and hybrid sterility. Balcova et al. (2016) [21] profile the genome-wide recombination rate by visualizing the immunolocalization pattern of the MLH1 mismatch repair protein that resolves double-strand breaks into crossovers (Fig 1). This powerful approach enables the total number of recombination events to be counted in individual spermatocytes. The authors first use the MLH1 technique to confirm that the genome of an inbred strain representing the house mouse subspecies Mus musculus musculus experiences an average of 4.7 more crossovers than the genome of an inbred strain descended primarily from M. m. domesticus (a 19% increase). Next, Balcova et al. (2016) measure recombination rates in a panel of inbred strains, each of which harbors single chromosomes from the M. m. musculus strain on the genomic background of the M. m. domesticus strain. By comparing rates in chromosome substitution strains with the appropriate parental strain, the authors identify three chromosomes that affect the genome-wide crossover number. The locus with the largest phenotypic effect is located on the X chromosome. Rate differences among substitution strains carrying pieces of the chromosome further localize the position of this modifier of the global recombination rate to a 4.7 Mb interval (nicknamed Meir1 by the authors). This locus lies within a broader part of the X chromosome previously shown to modulate recombination in crosses involving other subspecies of mice [22,23]. Fig 1 Image illustrating immunofluorescent cytology approach to measuring the genome-wide recombination rate (from Balcova et al., 2016). Several lines of evidence suggest that this region of the X chromosome genetically links recombination and speciation. First, the same interval is associated with multiple correlates of hybrid sterility. Small testes, low sperm count, and morphological abnormalities in sperm observed in F1 males from crosses between M. m. musculus mothers and M. m. domesticus fathers all map to this genomic location [24,25]. Second, hybrid male sterility due to one of the loci in this X-linked region (Hstx2) involves an incompatibility with Prdm9 [25]. Finally, the effects of this piece of the X chromosome on recombination and sterility show similar properties, including male specificity. The findings of Balcova et al. (2016), along with previous discoveries about Prdm9, raise the intriguing possibility that recombination and speciation are mechanistically coupled. Remarkably, both the fine-scale placement of crossovers (mediated by Prdm9) and the global recombination rate (controlled by Meir1) seem to be associated with hybrid sterility. The authors propose that the incompatibility between Prdm9 and Hstx2 could inhibit the repair of double-strand breaks, which could in turn lead to the disrupted synapsis of chromosomes and meiotic arrest observed in sterile F1 males [26,27]. Regardless of the mechanism, the larger implication is that evolutionary divergence in recombination genes can generate a frequently observed form of reproductive isolation. Once the causative gene(s) and mutation(s) corresponding to Meir1 and Hstx2 are identified in this system, important remaining questions can be answered. Do the joint effects on recombination and hybrid sterility reflect the pleiotropic activities of a single gene or multiple linked genes? What are the molecular, cellular, and developmental bases of these phenotypes? What is the evolutionary history of the causative mutations? Did natural selection drive phenotypic divergence, as seems to be the case for the rapidly evolving Prdm9 [28]? The results from Balcova et al. (2016) should also motivate similar studies in other organisms. The observations that recombination rate evolves and that hybrid sterility is a key reproductive barrier are phylogenetically widespread, leaving no obvious reason that the emerging connection between recombination and speciation should be restricted to mice.

Recombination is an engine of genetic diversity and therefore constitutes a key process in evolutionary biology and genetics. While the outcome of crossover recombination can readily be detected as shuffled alleles by following the inheritance of markers in pedigreed families, the more precise location of both crossover and non-crossover recombination events has been difficult to pinpoint. As a consequence, we lack a detailed portrait of the recombination landscape for most organisms and knowledge on how this landscape impacts on sequence evolution at a local scale. To localize recombination events with high resolution in an avian system, we performed whole-genome re-sequencing at high coverage of a complete three-generation collared flycatcher pedigree. We identified 325 crossovers at a median resolution of 1.4 kb, with 86% of the events localized to, Author Summary Homologous chromosomes exchange genetic material during cell division at meiosis by the process of crossover recombination. Although such crossover events are visually identifiable by cytogenetic techniques, it has remained a challenge to pinpoint the location of crossovers at the DNA sequence level. An emerging novel possibility to approach this challenge is to exploit the high resolution offered by re-sequencing of multiple individuals in species in which a genome assembly is available. Specifically, by sequencing members of a family, the inheritance of chromosomal segments can be followed and the location of crossover as well as non-crossover recombination determined. We performed such an endeavour in the collared flycatcher, a songbird species that has been in focus for extensive ecological and evolutionary research. We found that crossover events were concentrated to certain ‘hot-spot’ regions, and that the density of such events was highest in and close to genes. A higher rate of crossover recombination was found in males than in females and in males, but not in females, the rate of crossover increased towards ends of chromosomes. The location of non-crossovers was significantly associated with that of crossovers. We could further document an unbalanced transmission of genetic variants at non-crossover events via to the process of GC-biased gene conversion.

In many mammals, including humans and mice, the zinc finger histone methyltransferase PRDM9 performs the first step in meiotic recombination by specifying the locations of hotspots, the sites of genetic recombination. PRDM9 binds to DNA at hotspots through its zinc finger domain and activates recombination by trimethylating histone H3K4 on adjacent nucleosomes through its PR/SET domain. Recently, the isolated PR/SET domain of PRDM9 was shown capable of also trimethylating H3K36 in vitro, raising the question of whether this reaction occurs in vivo during meiosis, and if so, what its function might be. Here, we show that full-length PRDM9 does trimethylate H3K36 in vivo in mouse spermatocytes. Levels of H3K4me3 and H3K36me3 are highly correlated at hotspots, but mutually exclusive elsewhere. In vitro, we find that although PRDM9 trimethylates H3K36 much more slowly than it does H3K4, PRDM9 is capable of placing both marks on the same histone molecules. In accord with these results, we also show that PRDM9 can trimethylate both K4 and K36 on the same nucleosomes in vivo, but the ratio of K4me3/K36me3 is much higher for the pair of nucleosomes adjacent to the PRDM9 binding site compared to the next pair further away. Importantly, H3K4me3/H3K36me3-double-positive nucleosomes occur only in regions of recombination: hotspots and the pseudoautosomal (PAR) region of the sex chromosomes. These double-positive nucleosomes are dramatically reduced when PRDM9 is absent, showing that this signature is PRDM9-dependent at hotspots; the residual double-positive nucleosomes most likely come from the PRDM9-independent PAR. These results, together with the fact that PRDM9 is the only known mammalian histone methyltransferase with both H3K4 and H3K36 trimethylation activity, suggest that trimethylation of H3K36 plays an important role in the recombination process. Given the known requirement of H3K36me3 for double strand break repair by homologous recombination in somatic cells, we suggest that it may play the same role in meiosis., Author Summary Genetic recombination is the meiotic process by which novel combinations of alleles are passed on to the next generation. This process accelerates evolution by creating genetic diversity, and is also essential for successful meiosis. In mammals, the enzyme PRDM9 initiates recombination and determines the subset of sites within the genome—called recombination hotspots—that recombine. PRDM9 does this by binding DNA at hotspots and placing epigenetic marks. Previously, PRDM9 was only known to place the H3K4me3 mark at hotspots in living cells. Here, we show that PRDM9 places the H3K36me3 mark at hotspots, and that H3K4me3 and H3K36me3 coincide only at regions of recombination in germ cells. We prove that this coincidence is driven almost entirely by PRDM9; there is dramatically less coincidence between H3K4me3 and H3K36me3 when PRDM9 is absent. These results reveal a new enzymatic function for PRDM9, and a new epigenetic signature at hotspots that may restrict recombination to these sites. Since aberrant recombination can cause aneuploidy resulting in fetal loss, and abnormal genome rearrangements that underlie many congenital syndromes and some cancers, a thorough understanding of this fundamental process has potentially far-reaching implications.

Few experimental studies have examined the role that sexual recombination plays in bacterial evolution, including the effects of horizontal gene transfer on genome structure. To address this limitation, we analyzed genomes from an experiment in which Escherichia coli K-12 Hfr (high frequency recombination) donors were periodically introduced into 12 evolving populations of E. coli B and allowed to conjugate repeatedly over the course of 1000 generations. Previous analyses of the evolved strains from this experiment showed that recombination did not accelerate adaptation, despite increasing genetic variation relative to asexual controls. However, the resolution in that previous work was limited to only a few genetic markers. We sought to clarify and understand these puzzling results by sequencing complete genomes from each population. The effects of recombination were highly variable: one lineage was mostly derived from the donors, while another acquired almost no donor DNA. In most lineages, some regions showed repeated introgression and others almost none. Regions with high introgression tended to be near the donors’ origin of transfer sites. To determine whether introgressed alleles imposed a genetic load, we extended the experiment for 200 generations without recombination and sequenced whole-population samples. Beneficial alleles in the recipient populations were occasionally driven extinct by maladaptive donor-derived alleles. On balance, our analyses indicate that the plasmid-mediated recombination was sufficiently frequent to drive donor alleles to fixation without providing much, if any, selective advantage., Author summary Bacteria often transfer genes encoding antibiotic resistance as well as other important traits, but the extent of intergenomic recombination—in effect, sex—is highly variable across bacterial species. Why? A better understanding of how and why bacteria exchange genes would help people combat the spread of infectious disease as well as shed light on the evolutionary origins of sex. Here, we sequenced genomes from an evolution experiment with Escherichia coli in which recombination was extensive but, unexpectedly, did not speed up the rate of adaptation. In this experiment, the effective rate of chromosomal recombination was much higher than previously inferred for natural E. coli populations. In fact, the rate was so high that introduced genes sometimes drove established beneficial alleles to extinction in the experimental populations. In effect, genes that were physically linked to the genes causing recombination had a strong transmission advantage, whether or not they provided any selective advantage to the recipient cells.

Meiotic recombination normally takes place between allelic sequences on homologs. This process can also occur between non-allelic homologous sequences. Such ectopic interaction events can lead to chromosome rearrangements and are normally avoided. However, much remains unknown about how these ectopic interaction events are sensed and eliminated. In this study, using a screen in rice, we characterized a homolog of HUS1 and explored its function in meiotic recombination. In Oshus1 mutants, in conjunction with nearly normal homologous pairing and synapsis, vigorous, aberrant ectopic interactions occurred between nonhomologous chromosomes, leading to multivalent formation and subsequent chromosome fragmentation. These ectopic interactions relied on programed meiotic double strand breaks and were formed in a manner independent of the OsMER3-mediated interference-sensitive crossover pathway. Although early homologous recombination events occurred normally, the number of interference-sensitive crossovers was reduced in the absence of OsHUS1. Together, our results indicate that OsHUS1 might be involved in regulating ectopic interactions during meiosis, probably by forming the canonical RAD9-RAD1-HUS1 (9-1-1) complex., Author Summary Meiosis is a special type of cell division that generates gametes for sexual reproduction. During meiosis, recombination not only occurs between allelic sequences on homologs, but also between non-allelic homologous sequences at dispersed loci. Such ectopic recombination is the main cause of chromosomal alterations and accounts for numerous genomic disorders in humans. To ensure genomic integrity, those ectopic recombinations must be quickly resolved. Despite the importance of ectopic recombination suppression, the mechanism underlying this process still remains largely unknown. Here, using rice as a model system, we identified the rice HUS1 homolog, a member of the RAD9-RAD1-HUS1 (9-1-1) complex, and elucidated its roles in meiotic recombination. In Oshus1, vigorous ectopic interactions occur between nonhomologous chromosomes, and the number of crossovers is reduced. We suspect that OsHUS1 participates in regulating ectopic interactions during meiosis, probably by forming the canonical RAD9-RAD1-HUS1 (9-1-1) complex.

Mammalian common fragile sites are loci of frequent chromosome breakage and putative recombination hotspots. Here, we utilized Replication Slow Zones (RSZs), a budding yeast homolog of the mammalian common fragile sites, to examine recombination activities at these loci. We found that rates of URA3 inactivation of a hisG-URA3-hisG reporter at RSZ and non-RSZ loci were comparable under all conditions tested, including those that specifically promote chromosome breakage at RSZs (hydroxyurea [HU], mec1Δ sml1Δ, and high temperature), and those that suppress it (sml1Δ and rrm3Δ). These observations indicate that RSZs are not recombination hotspots and that chromosome fragility and recombination activity can be uncoupled. Results confirmed recombinogenic effects of HU, mec1Δ sml1Δ, and rrm3Δ and identified temperature as a regulator of mitotic recombination. We also found that these conditions altered the nature of recombination outcomes, leading to a significant increase in the frequency of URA3 inactivation via loss of heterozygosity (LOH), the type of genetic alteration involved in cancer development. Further analyses revealed that the increase was likely due to down regulation of intrachromatid and intersister (IC/IS) bias in mitotic recombination, and that RSZs exhibited greater sensitivity to HU dependent loss of IC/IS bias than non RSZ loci. These observations suggest that recombinogenic conditions contribute to genome rearrangements not only by increasing the overall recombination activity, but also by altering the nature of recombination outcomes by their effects on recombination partner choice. Similarly, fragile sites may contribute to cancer more frequently than non-fragile loci due their enhanced sensitivity to certain conditions that down-regulate the IC/IS bias rather than intrinsically higher rates of recombination., Author Summary Chromosome rearrangements are frequently associated with human cancers. Such rearrangement can result from a DNA break followed by an erroneous repair. Mammalian common fragile sites are one of the most extensively studied naturally occurring breakage prone regions of the genome. It has been proposed that fragile sites are recombination hotspots and that increased recombination activity at these loci contribute to cancer. We examined this hypothesis using a model organism, budding yeast Saccharomyces cerevisiae, where a homolog of the mammalian common fragile sites has been identified. Unexpectedly, our results showed that the rate of recombination at the fragile sites was not any higher than non fragile sites, even under the conditions that promoted chromosome breakage at the fragile sites. However, we found that the frequency of loss of heterozygosity (LOH) and translocation, the type of recombination outcomes known to contribute to cancer, to be significantly elevated at fragile sites under certain conditions. These findings suggest that the fragile sites might indeed contribute to cancer more frequently than non-fragile loci, but the reason for this is likely to be due the nature of the recombination outcome(s) rather than higher rates of recombination.

Faithful chromosome segregation during meiosis requires that homologous chromosomes associate and recombine. Chiasmata, the cytological manifestation of recombination, provide the physical link that holds the homologs together as a pair, facilitating their orientation on the spindle at meiosis I. Formation of most crossover (CO) events requires the assistance of a group of proteins collectively known as ZMM. HFM1/Mer3 is in this group of proteins and is required for normal progression of homologous recombination and proper synapsis between homologous chromosomes in a number of model organisms. Our work is the first study in mammals showing the in vivo function of mouse HFM1. Cytological observations suggest that initial steps of recombination are largely normal in a majority of Hfm1−/− spermatocytes. Intermediate and late stages of recombination appear aberrant, as chromosomal localization of MSH4 is altered and formation of MLH1foci is drastically reduced. In agreement, chiasma formation is reduced, and cells arrest with subsequent apoptosis at diakinesis. Our results indicate that deletion of Hfm1 leads to the elimination of a major fraction but not all COs. Formation of chromosome axial elements and homologous pairing is apparently normal, and Hfm1−/− spermatocytes progress to the end of prophase I without apparent developmental delay or apoptosis. However, synapsis is altered with components of the central region of the synaptonemal complex frequently failing to extend the full length of the chromosome axes. We propose that initial steps of recombination are sufficient to support homology recognition, pairing, and initial chromosome synapsis and that HFM1 is required to form normal numbers of COs and to complete synapsis., Author Summary Meiosis is a specialized type of cell division during which haploid gametes are generated to offset doubling of the chromosome number that occurs at fertilization. Prior to segregation in the first meiotic division, meiotic chromosomes pair and recombine. Chiasmata, products of recombination, provide the temporary physical link between the chromosomes of each pair. This association is required to prepare the chromosomes for segregation to the opposite poles of the spindle at the first meiotic division, so that ultimately each gamete receives only one copy of each chromosome. For this reason, errors in recombination are commonly linked to human infertility, spontaneous abortions, and severe aneuploid-based birth defects. We observed that the absence of HFM1/Mer3 results in deficient recombination and in incomplete meiosis-specific modification of chromosome structure. Our findings thus indicate that the mouse HFM1 protein is required for normal progression of important aspects of meiosis that act to insure that the correct numbers of chromosomes are transferred to the next generation.

Meiotic recombination safeguards proper segregation of homologous chromosomes into gametes, affects genetic variation within species, and contributes to meiotic chromosome recognition, pairing and synapsis. The Prdm9 gene has a dual role, it controls meiotic recombination by determining the genomic position of crossover hotspots and, in infertile hybrids of house mouse subspecies Mus m. musculus (Mmm) and Mus m. domesticus (Mmd), it further functions as the major hybrid sterility gene. In the latter role Prdm9 interacts with the hybrid sterility X 2 (Hstx2) genomic locus on Chromosome X (Chr X) by a still unknown mechanism. Here we investigated the meiotic recombination rate at the genome-wide level and its possible relation to hybrid sterility. Using immunofluorescence microscopy we quantified the foci of MLH1 DNA mismatch repair protein, the cytological counterparts of reciprocal crossovers, in a panel of inter-subspecific chromosome substitution strains. Two autosomes, Chr 7 and Chr 11, significantly modified the meiotic recombination rate, yet the strongest modifier, designated meiotic recombination 1, Meir1, emerged in the 4.7 Mb Hstx2 genomic locus on Chr X. The male-limited transgressive effect of Meir1 on recombination rate parallels the male-limited transgressive role of Hstx2 in hybrid male sterility. Thus, both genetic factors, the Prdm9 gene and the Hstx2/Meir1 genomic locus, indicate a link between meiotic recombination and hybrid sterility. A strong female-specific modifier of meiotic recombination rate with the effect opposite to Meir1 was localized on Chr X, distally to Meir1. Mapping Meir1 to a narrow candidate interval on Chr X is an important first step towards positional cloning of the respective gene(s) responsible for variation in the global recombination rate between closely related mouse subspecies., Author Summary During differentiation of germ cells into gametes, a maternal and a paternal copy of each chromosome have to find each other, pair, and synapse in order to ensure proper chromosome segregation into the gametes. Because of the unique ability to identify homologous DNA sequences between homologous chromosomes, meiotic recombination is an essential step in proper chromosome pairing and synapsis in the majority of species. However, when the paternal and maternal sets of chromosomes come from different (sub)species, the recognition of homologs can be disturbed and result in sterility of male hybrids. In this study we investigated the genetic control of variation in the global recombination rate between two closely related mouse subspecies with regard to the known infertility of their F1 hybrids. We show that the variation in the global recombination rate between both subspecies is under the control of three genomic loci. The strongest one appeared within the hybrid sterility X2 genomic locus on Chromosome X. Our findings will allow positional cloning of the gene and will shed new light on the role of meiotic recombination in reproductive isolation between closely related species.

Meiotic recombination ensures proper chromosome segregation in many sexually reproducing organisms. Despite this crucial function, rates of recombination are highly variable within and between taxa, and the genetic basis of this variation remains poorly understood. Here, we exploit natural variation in the inbred, sequenced lines of the Drosophila melanogaster Genetic Reference Panel (DGRP) to map genetic variants affecting recombination rate. We used a two-step crossing scheme and visible markers to measure rates of recombination in a 33 cM interval on the X chromosome and in a 20.4 cM interval on chromosome 3R for 205 DGRP lines. Though we cannot exclude that some biases exist due to viability effects associated with the visible markers used in this study, we find ~2-fold variation in recombination rate among lines. Interestingly, we further find that recombination rates are uncorrelated between the two chromosomal intervals. We performed a genome-wide association study to identify genetic variants associated with recombination rate in each of the two intervals surveyed. We refined our list of candidate variants and genes associated with recombination rate variation and selected twenty genes for functional assessment. We present strong evidence that five genes are likely to contribute to natural variation in recombination rate in D. melanogaster; these genes lie outside the canonical meiotic recombination pathway. We also find a weak effect of Wolbachia infection on recombination rate and we confirm the interchromosomal effect. Our results highlight the magnitude of population variation in recombination rate present in D. melanogaster and implicate new genetic factors mediating natural variation in this quantitative trait., Author Summary During meiosis, homologous chromosomes exchange genetic material through recombination. In most sexually reproducing species, recombination is necessary for chromosomes to properly segregate. Recombination defects can generate gametes with an incorrect number of chromosomes, which is devastating for organismal fitness. Despite the central role of recombination for chromosome segregation, recombination is highly variable process both within and between species. Though it is clear that this variation is due at least in part to genetics, the specific genes contributing to variation in recombination within and between species remain largely unknown. This is particularly true in the model organism, Drosophila melanogaster. Here, we use the D. melanogaster Genetic Reference Panel to determine the scale of population-level variation in recombination rate and to identify genes significantly associated with this variation. We estimated rates of recombination on two different chromosomes in 205 strains of D. melanogaster. We also used genome-wide association mapping to identify genetic factors associated with recombination rate variation. We find that recombination rate on the two chromosomes are independent traits. We further find that population-level variation in recombination is mediated by many loci of small effect, and that the genes contributing to variation in recombination rate are outside of the well-characterized meiotic recombination pathway.

Recombination is a fundamental biological process with profound evolutionary implications. Theory predicts that recombination increases the effectiveness of selection in natural populations. Yet, direct tests of this prediction have been restricted to qualitative trends due to the lack of detailed characterization of recombination rate variation across genomes and within species. The use of imprecise recombination rates can also skew population genetic analyses designed to assess the presence and mode of selection across genomes. Here we report the first integrated high-resolution description of genomic and population variation in recombination, which also distinguishes between the two outcomes of meiotic recombination: crossing over (CO) and gene conversion (GC). We characterized the products of 5,860 female meioses in Drosophila melanogaster by genotyping a total of 139 million informative SNPs and mapped 106,964 recombination events at a resolution down to 2 kilobases. This approach allowed us to generate whole-genome CO and GC maps as well as a detailed description of variation in recombination among individuals of this species. We describe many levels of variation in recombination rates. At a large-scale (100 kb), CO rates exhibit extreme and highly punctuated variation along chromosomes, with hot and coldspots. We also show extensive intra-specific variation in CO landscapes that is associated with hotspots at low frequency in our sample. GC rates are more uniformly distributed across the genome than CO rates and detectable in regions with reduced or absent CO. At a local scale, recombination events are associated with numerous sequence motifs and tend to occur within transcript regions, thus suggesting that chromatin accessibility favors double-strand breaks. All these non-independent layers of variation in recombination across genomes and among individuals need to be taken into account in order to obtain relevant estimates of recombination rates, and should be included in a new generation of population genetic models of the interaction between selection and linkage., Author Summary Most sexual eukaryotes require recombination between homologous chromosomes for the proper formation of haploid gametes from diploid germ cells. Evolutionarily, recombination increases the effectiveness of selection in natural populations, thus explaining the pervasiveness of recombination and sex. Recombination is also a central parameter in population genetic studies designed to detect the presence of selection acting across genomes. Current evolutionary analyses are hindered, however, by the use of imprecise recombination rates that can influence the results and skew their interpretation. This limitation is associated with the lack of detailed characterization of natural variation in recombination across genomes and within species. Our study in Drosophila melanogaster represents the first integrated, whole-genome description of recombination that alleviates these deficiencies in any organism. Our results and conclusions will help to characterize the molecular basis of the observed variation in recombination across genomes and have an immediate impact on population genetic analyses of selection, laying the foundation for a new generation of population models that will better capture natural variation in recombination and its consequences.

Accurate chromosome segregation during meiosis requires that homologous chromosomes pair and become physically connected so that they can orient properly on the meiosis I spindle. These connections are formed by homologous recombination closely integrated with the development of meiosis-specific, higher-order chromosome structures. The yeast Pch2 protein has emerged as an important factor with roles in both recombination and chromosome structure formation, but recent analysis suggested that TRIP13, the mouse Pch2 ortholog, is not required for the same processes. Using distinct Trip13 alleles with moderate and severe impairment of TRIP13 function, we report here that TRIP13 is required for proper synaptonemal complex formation, such that autosomal bivalents in Trip13-deficient meiocytes frequently displayed pericentric synaptic forks and other defects. In males, TRIP13 is required for efficient synapsis of the sex chromosomes and for sex body formation. Furthermore, the numbers of crossovers and chiasmata are reduced in the absence of TRIP13, and their distribution along the chromosomes is altered, suggesting a role for TRIP13 in aspects of crossover formation and/or control. Recombination defects are evident very early in meiotic prophase, soon after DSB formation. These findings provide evidence for evolutionarily conserved functions for TRIP13/Pch2 in both recombination and formation of higher order chromosome structures, and they support the hypothesis that TRIP13/Pch2 participates in coordinating these key aspects of meiotic chromosome behavior., Author Summary Meiosis is the specialized cell division that gives rise to reproductive cells such as sperm and eggs. During meiosis in most organisms, genetic information is exchanged between homologous maternal and paternal chromosomes through the process of homologous recombination. This recombination forms connections between homologous chromosomes that allow them to segregate accurately when the meiotic cell divides. Recombination defects can result in reproductive cells with abnormal chromosome numbers, which are a major cause of developmental disorders and spontaneous abortions in humans. Meiotic recombination is tightly controlled such that each pair of chromosomes undergoes at least one crossover recombination event despite a low average number of crossovers per chromosome. Recombination is coordinated with the development of specialized, meiosis-specific chromosome structures that stabilize pairing interactions between homologous maternal and paternal chromosomes. We show here that the mouse TRIP13 protein is required for normal execution of many aspects of meiotic recombination and chromosome structure development that it was not previously known to influence. Intriguingly, many of these new roles appear to parallel known functions of a homologous protein from budding yeast, called Pch2. These findings thus indicate that TRIP13/Pch2 functions are more widely conserved throughout evolution than thought before.

In meiosis, programmed DNA breaks repaired by homologous recombination (HR) can be processed into inter-homolog crossovers that promote the accurate segregation of chromosomes. In general, more programmed DNA double-strand breaks (DSBs) are formed than the number of inter-homolog crossovers, and the excess DSBs must be repaired to maintain genomic stability. Sister-chromatid (inter-sister) recombination is postulated to be important for the completion of meiotic DSB repair. However, this hypothesis is difficult to test because of limited experimental means to disrupt inter-sister and not inter-homolog HR in meiosis. We find that the conserved Structural Maintenance of Chromosomes (SMC) 5 and 6 proteins in Caenorhabditis elegans are required for the successful completion of meiotic homologous recombination repair, yet they appeared to be dispensable for accurate chromosome segregation in meiosis. Mutations in the smc-5 and smc-6 genes induced chromosome fragments and dismorphology. Chromosome fragments associated with HR defects have only been reported in mutants, which have disrupted inter-homolog crossover. Surprisingly, the smc-5 and smc-6 mutations did not disrupt the formation of chiasmata, the cytologically visible linkages between homologous chromosomes formed from meiotic inter-homolog crossovers. The mutant fragmentation defect appeared to be preferentially enhanced by the disruptions of inter-homolog recombination but not by the disruptions of inter-sister recombination. Based on these findings, we propose that the C. elegans SMC-5/6 proteins are required in meiosis for the processing of homolog-independent, presumably sister-chromatid-mediated, recombination repair. Together, these results demonstrate that the successful completion of homolog-independent recombination is crucial for germ cell genomic stability., Author Summary Sperm and oocytes are essential for the faithful transmission of genetic information during sexual reproduction. As germ cells mature into sperm and oocytes, DNA double-strand breaks (DSBs) are deliberately created on each chromosome and a subset of DSBs is repaired to form meiotic crossovers between homologous chromosomes. Because germ cells must undergo this programmed process of deliberate DNA damage and repair, identifying repair factors active in germ cells and determining the requirement of their functions in meiotic DSB repair are important first steps in understanding infertility and developmental disorders caused by defective sperm and oocytes. In this manuscript, we find that the evolutionarily conserved SMC-5 and SMC-6 proteins fulfill a critical role in preserving genomic stability in germ cells in C. elegans. Our findings further describe the genetic mechanisms by which the C. elegans SMC-5/6 proteins function in meiotic DSB repair. These data reveal that inter-sister homologous recombination, a repair mechanism thought to function as a back-up repair method in meiosis, serves a more significant role in normal meiosis than was previously appreciated.

Studies of human trisomies indicate a remarkable relationship between abnormal meiotic recombination and subsequent nondisjunction at maternal meiosis I or II. Specifically, failure to recombine or recombination events located either too near to or too far from the centromere have been linked to the origin of human trisomies. It should be possible to identify these abnormal crossover configurations by using immunofluorescence methodology to directly examine the meiotic recombination process in the human female. Accordingly, we initiated studies of crossover-associated proteins (e.g., MLH1) in human fetal oocytes to analyze their number and distribution on nondisjunction-prone human chromosomes and, more generally, to characterize genome-wide levels of recombination in the human female. Our analyses indicate that the number of MLH1 foci is lower than predicted from genetic linkage analysis, but its localization pattern conforms to that expected for a crossover-associated protein. In studies of individual chromosomes, our observations provide evidence for the presence of “vulnerable” crossover configurations in the fetal oocyte, consistent with the idea that these are subsequently translated into nondisjunctional events in the adult oocyte., Author Summary One of the defining events in the formation of eggs and sperm is meiotic recombination, a process in which DNA is exchanged between “partner” chromosomes. If this process is perturbed, chromosomes often go astray during meiotic division, resulting in eggs or sperm with too many or too few chromosomes. In humans, the resulting embryos are almost always abnormal and are a major source of miscarriages or congenital birth defects (e.g., Down syndrome). Over the past decade, techniques have become available that make possible the examination of meiotic recombination “as it happens” in the human egg, and this has allowed us to define the properties of this process. In the present study, we used this methodology to characterize meiotic recombination in human fetal oocytes and to ask whether we could identify abnormal recombination patterns. Such patterns were, indeed, observed for each chromosome we studied, although different abnormalities were observed among the individual chromosomes. Thus, factors operating before birth influence the likelihood of chromosome misdivision in the adult oocyte, although those that make chromosome 21 misdivide (leading to Down syndrome) may not be the same as those that make other chromosomes misdivide.

The behavior of chromosomes during meiosis has been likened to a middle school dance, where partners find one another, form couples that move about and trade information, and then separate to opposite sides of the dance hall. With chromosomes, as with the dancers, forming exclusive couples often is difficult—individuals can be attracted to more than one partner or find themselves trapped behind or between other couples—and, failing to form a couple effectively, end up on the wrong side of the dance hall. For chromosomes, this failure of pairing and segregation leads to an unbalanced chromosome complement (aneuploidy), with its attendant problems of sterility and genetic disease. Two papers in this issue of PLoS Genetics [1],[2] demonstrate that telomere-promoted movements influence nearly every step in chromosome pairing and meiotic recombination, opening a new avenue to address questions that have intrigued biologists and vexed clinicians for over a hundred years. Chromosome movement is implicit in the classically recognized stages of meiotic prophase, but descriptions of directly visualized movements have been rare (see [3]). Early in prophase, chromosomes transition from having their centromeres clustered near the spindle pole (the Rabl orientation) to having their telomeres clustered at the nuclear periphery adjacent to the spindle pole (the bouquet orientation; see Figure 1). The bouquet stage ends with dispersal of telomeres across the inner nuclear envelope as chromosomes finalize their intimate pairing by forming synaptonemal complexes (SCs) that link chromosome pairs closely along their lengths. The formation of these intimate, exclusive partnerships would seem to finish the task at hand and to end the need for active, whole-chromosome movements, but this turns out not to be the case. Figure 1 Chromosomes Pass through Distinct Organizational Phases as They Negotiate Meiotic Prophase. Movements that persist throughout meiotic prophase were first described by Hiraoka's group in the fission yeast Schizosaccharomyces pombe where, following bouquet formation, telomeres remain at the spindle pole while it leads the nucleus along microtubules, back and forth through the cell, until just before the first meiotic division [4]. Meiotic prophase is noncanonical in S. pombe in that synaptonemal complexes are not formed and recombination is not regulated to avoid forming crossovers near one another (i.e., there is no positive crossover interference). This has led some to question the generality of persistent movements. Recently, however, similarly persistent rapid prophase movements (RPMs) have been described in the budding yeast Saccharomyces cerevisiae [5]–[7]. Although these movements are of individual chromosomes rather than of the whole genomic complement, and although they appear to be promoted by actin rather than by microtubules, each system involves SUN domain–containing proteins that are known to mediate transnuclear envelope linkages, in the present case tethering telomeres to the cytoskeleton [8]–[10]. Such linkages also are present in mammalian meiotic nuclei [11],[12], indicating a widely conserved mechanism and suggesting conserved function(s). Before its role in these movements was recognized, the budding yeast Ndj1 protein was known to promote bouquet formation, the normal kinetics of SC formation, and the usual pattern of meiotic recombination; to maintain low levels of ectopic recombination (genetic exchanges between homologous DNA sequences in nonallelic locations); and, ultimately, to reduce the frequency of aneuploidy [13]–[17]. Ndj1 also plays a role in anchoring telomeres to the inner nuclear envelope [15], apparently by stabilizing the association of telomeres with the transmembrane SUN protein, Mps3 [10]. Reports that the meiosis-specific budding yeast protein Csm4 is similar to Ndj1 in being required to prevent aneuploidy [18] led the authors of the two current papers to ask, in remarkable molecular detail, whether the meiotic requirements for Csm4 are similar to those for Mps3 and Ndj1. The simple answer is “Yes,” but the angel is in the details. The authors find that Csm4, unlike Mps3 and Ndj1, is not required to anchor telomeres to the nuclear envelope but is required for telomeres to engage in the RPMs (see [7],[8]). Nevertheless, the impact on the progress of recombination, in all its currently understood molecular intricacies, is similar—delays in the appearances of recombination intermediates begin very early in prophase and persist or lengthen as prophase progresses. The implication of these observations is that the RPMs are the critical factor rather than telomere tethering to the nuclear envelope per se. So then, what is the role of the RPMs? Here, the authors diverge somewhat in their answers. The paper from the Shinohara lab proposes that RPMs promote the biochemistry of recombination more or less directly, perhaps by affecting chromosome structure [1]. The paper from the Alani and Kleckner labs proposes that RPMs function during an early phase when the cell determines which early recombination intermediates will become crossovers. They suggest that delays in this phase, perhaps due to a requirement for Ndj1 and Csm4 to resolve chromosome interlocks (at zygotene, see Figure 1), generates the subsequent defects [2]. Tests of these hypotheses will require considerable ingenuity in experimental design. A simple and striking conclusion from these papers is that mechanical energy, pumped into the nucleus via the telomeres, contributes critically to the work of genetic recombination. Identification of the MNC complex (Mps3, Ndj1, Csm4) in Sa. cerevisiae and of related structures and pathways in other organisms is only the beginning to understanding how these transnuclear envelope tethers are constructed and regulated. Understanding how these connections and the movements they foster contribute to the faithful segregation of chromosomes in meiosis will be challenging and rewarding, like the middle school dance.

In most eukaryotes, the prophase of the first meiotic division is characterized by a high level of homologous recombination between homologous chromosomes. Recombination events are not distributed evenly within the genome, but vary both locally and at large scale. Locally, most recombination events are clustered in short intervals (a few kilobases) called hotspots, separated by large intervening regions with no or very little recombination. Despite the importance of regulating both the frequency and the distribution of recombination events, the genetic factors controlling the activity of the recombination hotspots in mammals are still poorly understood. We previously characterized a recombination hotspot located close to the Psmb9 gene in the mouse major histocompatibility complex by sperm typing, demonstrating that it is a site of recombination initiation. With the goal of uncovering some of the genetic factors controlling the activity of this initiation site, we analyzed this hotspot in both male and female germ lines and compared the level of recombination in different hybrid mice. We show that a haplotype-specific element acts at distance and in trans to activate about 2,000-fold the recombination activity at Psmb9. Another haplotype-specific element acts in cis to repress initiation of recombination, and we propose this control to be due to polymorphisms located within the initiation zone. In addition, we describe subtle variations in the frequency and distribution of recombination events related to strain and sex differences. These findings show that most regulations observed act at the level of initiation and provide the first analysis of the control of the activity of a meiotic recombination hotspot in the mouse genome that reveals the interactions of elements located both in and outside the hotspot., Author Summary In most sexually reproducing species, during meiosis a high level of recombination between homologous chromosomes is induced. These events are not evenly distributed in the genome but clustered in small regions called hotspots. The genetic factors controlling their activity in mammals are still poorly understood. We have performed experiments to identify factors that influence the recombination activity of a hotspot in the mouse genome. By detecting the recombination products by a PCR-based method, we show that the variation of hotspot activity (up to 2,000-fold) is mainly due to differences of initiation frequencies, rather than differences at later steps of recombination. In addition, we identify several levels of controls. First, the initiation of recombination is activated by a haplotype-specific element, localized outside the hotspot and acting in trans (when heterozygous, this element allows for recombination initiation on both homologous chromosomes). This suggests a unique type of regulation requiring the presence of a diffusible factor and/or of communications between homologous chromosomes before recombination. A second element represses the recombination initiation in cis, which might indicate the influence of local polymorphisms affecting initiation events. Our results provide the first functional analysis of the control of recombination initiation sites for meiotic recombination in mammals.

Recombination increases dramatically during meiosis to promote genetic exchange and generate recombinant progeny. Interestingly, meiotic recombination is unevenly distributed throughout genomes, and, as a consequence, genetic and physical map distances do not have a simple linear relationship. Recombination hotspots and coldspots have been described in many organisms and often reflect global features of chromosome structure. In particular, recombination frequencies are often distorted within or outside sex-determining regions of the genome. Here, we report that recombination is elevated adjacent to the mating-type locus (MAT) in the pathogenic basidiomycete Cryptococcus neoformans. Among fungi, C. neoformans has an unusually large MAT locus, and recombination is suppressed between the two >100-kilobase mating-type specific alleles. When genetic markers were introduced at defined physical distances from MAT, we found the meiotic recombination frequency to be ~20% between MAT and a flanking marker at 5, 10, 50, or 100 kilobases from the right border. As a result, the physical/genetic map ratio in the regions adjacent to MAT is distorted ~10- to 50-fold compared to the genome-wide average. Moreover, recombination frequently occurred on both sides of MAT and negative interference between crossovers was observed. MAT heterozygosity was not required for enhanced recombination, implying that this process is not due to a physical distortion from the two non-paired alleles and could also occur during same-sex mating. Sequence analysis revealed a correlation between high G + C content and these hotspot regions. We hypothesize that the presence of recombinational activators may have driven several key events during the assembly and reshaping of the MAT locus and may have played similar roles in the origins of both metabolic and biosynthetic gene clusters. Our findings suggest that during meiosis the MAT locus may be exchanged onto different genetic backgrounds and therefore have broad evolutionary implications with respect to mating-type switching in both model and pathogenic yeasts., Synopsis It is hypothesized that sexual reproduction enables genetic recombination between individuals to generate diversity, thus increasing population fitness. However, a well-known meiotic feature is that recombination is not randomly distributed across the genome: “coldspots” and “hotspots” exist, implying some regions undergo exchange more frequently. In this paper, the authors report the discovery of recombination hotspots linked to the sex-determining region mating-type locus (MAT) in the human pathogenic fungus Cryptococcus neoformans. Through genetic analysis, hotspots were found to reside on both sides of MAT and are associated with DNA regions marked by high G + C base pair composition. Moreover, recombination on one side of MAT is associated with a recombination on the other. As a result, the MAT locus can be replaced onto the homologous chromosome as a unit—an effective switch of MAT. Based on these findings, they propose that the MAT-linked recombination hotspots impacted key steps during MAT evolution. This study has broader implications on how gene clusters (including those involved in metabolism or secondary metabolite production) are assembled and maintained and explains how recombination is distorted in sex-determining regions in eukaryotes.

Although homologous recombination is an important pathway for the repair of double-stranded DNA breaks in mitotically dividing eukaryotic cells, these events can also have negative consequences, such as loss of heterozygosity (LOH) of deleterious mutations. We mapped about 140 spontaneous reciprocal crossovers on the right arm of the yeast chromosome IV using single-nucleotide-polymorphism (SNP) microarrays. Our mapping and subsequent experiments demonstrate that inverted repeats of Ty retrotransposable elements are mitotic recombination hotspots. We found that the mitotic recombination maps on the two homologs were substantially different and were unrelated to meiotic recombination maps. Additionally, about 70% of the DNA lesions that result in LOH are likely generated during G1 of the cell cycle and repaired during S or G2. We also show that different genetic elements are associated with reciprocal crossover conversion tracts depending on the cell cycle timing of the initiating DSB., Author Summary Double-strand breaks (DSBs) are DNA lesions that can be fatal to a cell if left unrepaired. They can be caused by exogenous sources, such as gamma radiation, or endogenous stresses, such as high levels of transcription. Yeast cells primarily repair DSBs that are initiated outside of meiosis by mitotic recombination, which can result in physical exchanges between chromosomes, known as crossovers. We created a mitotic recombination map of one chromosome arm, representing 10% of the genome. This recombination map allows us to determine which regions of the chromosome arm are more susceptible to DNA damage than other regions. We were able to determine that most DSBs that result in detectable genomic changes were initiated prior to DNA replication and that some secondary DNA structures can be recombination hotspots. Recombination can also occur during meiosis, as a method of ensuring proper chromosome segregation. However, previously reported meiotic recombination maps have no correlation with our mitotic recombination map.

Selective and/or neutral processes may govern variation in DNA content and, ultimately, genome size. The observation in several organisms of a negative correlation between recombination rate and intron size could be compatible with a neutral model in which recombination is mutagenic for length changes. We used whole-genome data on small insertions and deletions within transposable elements from chicken and zebra finch to demonstrate clear links between recombination rate and a number of attributes of reduced DNA content. Recombination rate was negatively correlated with the length of introns, transposable elements, and intergenic spacer and with the rate of short insertions. Importantly, it was positively correlated with gene density, the rate of short deletions, the deletion bias, and the net change in sequence length. All these observations point at a pattern of more condensed genome structure in regions of high recombination. Based on the observed rates of small insertions and deletions and assuming that these rates are representative for the whole genome, we estimate that the genome of the most recent common ancestor of birds and lizards has lost nearly 20% of its DNA content up until the present. Expansion of transposable elements can counteract the effect of deletions in an equilibrium mutation model; however, since the activity of transposable elements has been low in the avian lineage, the deletion bias is likely to have had a significant effect on genome size evolution in dinosaurs and birds, contributing to the maintenance of a small genome. We also demonstrate that most of the observed correlations between recombination rate and genome contraction parameters are seen in the human genome, including for segregating indel polymorphisms. Our data are compatible with a neutral model in which recombination drives vertebrate genome size evolution and gives no direct support for a role of natural selection in this process., Author Summary One major implication from genetic work done several decades ago is that the genome contains a lot of sequences that do not constitute genes or other functional elements. The total amount of DNA—the genome size—is thus not necessarily an indicator of DNA complexity or organismal complexity, an observation often referred to as the C-value paradox (C-value being a measure of DNA content). What then is it that determines genome size? One model posits that the evolution of genome size is not a consequence of natural selection but is instead governed by the incidence and character of naturally occurring mutations that affect the length of DNA, a process that is not affected by selection. Here we present the results of an analysis of how recombination affects the size of avian and human genomes. We find strong evidence that the rate of recombination is a driving force of genome size evolution. In regions of the genome where recombination occurs frequently, the loss of DNA caused by small deletions is particularly pronounced. Our simulations show that the effect of such recombination-driven genome contraction can be profound over evolutionary time scales. These observations lead to a model in which recombination is mutagenic for length changes and that the incidence of deletions increases with increasing recombination rate. Although we cannot formally exclude that natural selection contributes to the observed relationship between recombination and genome contraction, we find no evidence to support such a scenario.

In this study we have analysed AtASY3, a coiled-coil domain protein that is required for normal meiosis in Arabidopsis. Analysis of an Atasy3-1 mutant reveals that loss of the protein compromises chromosome axis formation and results in reduced numbers of meiotic crossovers (COs). Although the frequency of DNA double-strand breaks (DSBs) appears moderately reduced in Atasy3-1, the main recombination defect is a reduction in the formation of COs. Immunolocalization studies in wild-type meiocytes indicate that the HORMA protein AtASY1, which is related to Hop1 in budding yeast, forms hyper-abundant domains along the chromosomes that are spatially associated with DSBs and early recombination pathway proteins. Loss of AtASY3 disrupts the axial organization of AtASY1. Furthermore we show that the AtASY3 and AtASY1 homologs BoASY3 and BoASY1, from the closely related species Brassica oleracea, are co-immunoprecipitated from meiocyte extracts and that AtASY3 interacts with AtASY1 via residues in its predicted coiled-coil domain. Together our results suggest that AtASY3 is a functional homolog of Red1. Since studies in budding yeast indicate that Red1 and Hop1 play a key role in establishing a bias to favor inter-homolog recombination (IHR), we propose that AtASY3 and AtASY1 may have a similar role in Arabidopsis. Loss of AtASY3 also disrupts synaptonemal complex (SC) formation. In Atasy3-1 the transverse filament protein AtZYP1 forms small patches rather than a continuous SC. The few AtMLH1 foci that remain in Atasy3-1 are found in association with the AtZYP1 patches. This is sufficient to prevent the ectopic recombination observed in the absence of AtZYP1, thus emphasizing that in addition to its structural role the protein is important for CO formation., Author Summary Homologous recombination (HR) during prophase I of meiosis leads to the formation of physical connections, known as chiasmata, between homologous chromosomes (homologs). Chiasmata are essential for accurate homolog segregation at the first meiotic division. HR is initiated by the formation of DNA double-strand breaks (DSBs). As DNA replication prior to meiosis results in the duplication of each homolog to form two identical sister chromatids, a DSB in one sister chromatid could potentially be repaired using the other as the repair template rather than one of the two non-sister chromatids of the homolog. If this route were predominant, the formation of chiasmata would be disfavored and chromosome segregation would be compromised. However, during meiosis there is a strong bias towards inter-homolog recombination (IHR). In this study we have identified AtASY3, a component of the proteinaceous axes that organize the chromosomes during meiosis in Arabidopsis. We find that AtASY3 interacts with AtASY1, a previously identified axis protein that is essential for crossover formation. We show that loss of AtASY3 disrupts the axis-organization of AtASY1. This results in a substantial reduction in chiasmata, and there is extensive chromosome mis-segregation. We propose that loss of AtASY3 affects the efficiency of the inter-homolog bias.

Homologous recombination is an important mechanism for the repair of DNA damage in mitotically dividing cells. Mitotic crossovers between homologues with heterozygous alleles can produce two homozygous daughter cells (loss of heterozygosity), whereas crossovers between repeated genes on non-homologous chromosomes can result in translocations. Using a genetic system that allows selection of daughter cells that contain the reciprocal products of mitotic crossing over, we mapped crossovers and gene conversion events at a resolution of about 4 kb in a 120-kb region of chromosome V of Saccharomyces cerevisiae. The gene conversion tracts associated with mitotic crossovers are much longer (averaging about 12 kb) than the conversion tracts associated with meiotic recombination and are non-randomly distributed along the chromosome. In addition, about 40% of the conversion events have patterns of marker segregation that are most simply explained as reflecting the repair of a chromosome that was broken in G1 of the cell cycle., Author Summary Most higher organisms have two copies of several different types of chromosomes. For example, the human female has 23 pairs of chromosomes. Although the chromosome pairs have very similar sequences, they are not identical. Members of a chromosome pair can swap segments from one chromosome to the other; these exchanges are called “recombination.” Most previous studies of recombination have been done in cells undergoing meiosis, the process that leads to the formation of eggs and sperm (gametes). Recombination, however, can also occur in cells that are dividing mitotically. In our study, we examine the properties of mitotic recombination in yeast. We show that mitotic recombination differs from meiotic recombination in two important ways. First, the sizes of the chromosome segments that are non-reciprocally transferred during mitotic recombination are much larger than those transferred during meiotic exchange. Second, in meiosis, most recombination events involve the repair of a single chromosome break, whereas in mitosis, about half of the recombination events appear to involve the repair of two chromosome breaks.

Previous studies of repetitive elements (REs) have implicated a mechanistic role in generating new chimerical genes. Such examples are consistent with the classic model for exon shuffling, which relies on non-homologous recombination. However, recent data for chromosomal aberrations in model organisms suggest that ectopic homology-dependent recombination may also be important. Lack of a dataset comprising experimentally verified young duplicates has hampered an effective examination of these models as well as an investigation of sequence features that mediate the rearrangements. Here we use ∼7,000 cDNA probes (∼112,000 primary images) to screen eight species within the Drosophila melanogaster subgroup and identify 17 duplicates that were generated through ectopic recombination within the last 12 mys. Most of these are functional and have evolved divergent expression patterns and novel chimeric structures. Examination of their flanking sequences revealed an excess of repetitive sequences, with the majority belonging to the transposable element DNAREP1 family, associated with the new genes. Our dataset strongly suggests an important role for REs in the generation of chimeric genes within these species., Author Summary In numerous organisms, many new genes have been found to originate through dispersed gene duplication and exon/domain shuffling. What recombination mechanisms were involved in the duplication and the shuffling processes? Lack of the intermediate products of recombination that share adequate sequence identity between homologous sequences, or the parental sequences from which the new genes were derived, often makes answering these questions difficult. We identified a number of young genes that originated in recently diverged branches in the evolutionary tree of the eight Drosophila melanogaster subgroup species, by using fluorescence in situ hybridization with polytene chromosomes. We analyzed the genomic regions surrounding 17 new dispersed duplicate genes and observed that most of these genes are flanked by repetitive elements (REs), including a large and diverged transposable element family, DNAREP1. Several copies of these REs are kept in both new and parental gene regions, and their degeneration is correlated with the increasing ages of the identified new genes. These data suggest that REs mediate the recombination responsible for the new gene origination.

Human recombination rates vary along the chromosomes as well as between the two sexes. There is growing evidence that epigenetic factors may have an important influence on recombination rates, as well as on crossover position. Using both public database analysis and wet-bench approaches, we revisited the relationship between increased rates of meiotic recombination and genome imprinting. We constructed metric linkage disequilibrium (LD) maps for all human chromosomal regions known to contain one or more imprinted genes. We show that imprinted regions contain significantly more LD units (LDU) and have significantly more haplotype blocks of smaller sizes than flanking nonimprinted regions. There is also an excess of hot-spots of recombination at imprinted regions, and this is likely to do with the presence of imprinted genes, per se. These findings indicate that imprinted chromosomal regions are historical “hot-spots” of recombination. We also demonstrate, by direct segregation analysis at the 11p15.5 imprinted region, that there is remarkable agreement between sites of meiotic recombination and steps in LD maps. Although the increase in LDU/Megabase at imprinted regions is not associated with any significant enrichment for any particular sequence class, major sequence determinants of recombination rates seem to differ between imprinted and control regions. Interestingly, fine-mapping of recombination events within the most male meiosis–specific recombination hot-spot of Chromosome 11p15.5 indicates that many events may occur within or directly adjacent to regions that are differentially methylated in somatic cells. Taken together, these findings support the involvement of a combination of specific DNA sequences and epigenetic factors as major determinants of hot-spots of recombination at imprinted chromosomal regions., Synopsis Now that the finished reference sequence of the human genome is available, focus has shifted towards understanding fundamental aspects of its functions. Meiotic recombination between maternal and paternal chromosomes serves an important mechanistic and evolutionary role in the transmission of the genome. Although significant progress has been made towards fine-mapping meiotic recombination events along human chromosomes, the characterization of factors that influence the position and frequency of crossovers remains a challenge. These authors have used data generated by the International HapMap Project as well as experimental analysis of a collection of three-generation Centre d'Etude du Polymorphisme Humain (CEPH) families, to show that chromosomal regions containing imprinted genes (i.e., genes transcribed only from one allele in a parent-of-origin–specific manner) exhibit higher rates of meiotic recombination than nonimprinted chromosomal regions. This characteristic is common for all major human populations. The major sequence determinants of recombination rates are likely to be different at imprinted and nonimprinted regions. Moreover, epigenetic modifications associated with imprinted regions may play an important role in increasing the frequency of meiotic crossovers and determining their position. Taken together these results suggest that a complex series of factors control meiotic recombination in the human.

Meiosis relies on the SPO11 endonuclease to generate the recombinogenic DNA double strand breaks (DSBs) required for homologous chromosome synapsis and segregation. The number of meiotic DSBs needs to be sufficient to allow chromosomes to search for and find their homologs, but not excessive to the point of causing genome instability. Here we report that the mammal-specific gene Tex19.1 promotes Spo11-dependent recombination in mouse spermatocytes. We show that the chromosome asynapsis previously reported in Tex19.1-/- spermatocytes is preceded by reduced numbers of recombination foci in leptotene and zygotene. Tex19.1 is required for normal levels of early Spo11-dependent recombination foci during leptotene, but not for upstream events such as MEI4 foci formation or accumulation of H3K4me3 at recombination hotspots. Furthermore, we show that mice carrying mutations in Ubr2, which encodes an E3 ubiquitin ligase that interacts with TEX19.1, phenocopy the Tex19.1-/- recombination defects. These data suggest that Tex19.1 and Ubr2 are required for mouse spermatocytes to accumulate sufficient Spo11-dependent recombination to ensure that the homology search is consistently successful, and reveal a hitherto unknown genetic pathway promoting meiotic recombination in mammals.

Meiotic recombination is essential for the repair of programmed double strand breaks (DSBs) to generate crossovers (COs) during meiosis. The efficient processing of meiotic recombination intermediates not only needs various resolvases but also requires proper meiotic chromosome structure. The Smc5/6 complex belongs to the structural maintenance of chromosome (SMC) family and is closely related to cohesin and condensin. Although the Smc5/6 complex has been implicated in the processing of recombination intermediates during meiosis, it is not known how Smc5/6 controls meiotic DSB repair. Here, using Caenorhabditis elegans we show that the SMC-5/6 complex acts synergistically with HIM-6, an ortholog of the human Bloom syndrome helicase (BLM) during meiotic recombination. The concerted action of the SMC-5/6 complex and HIM-6 is important for processing recombination intermediates, CO regulation and bivalent maturation. Careful examination of meiotic chromosomal morphology reveals an accumulation of inter-chromosomal bridges in smc-5; him-6 double mutants, leading to compromised chromosome segregation during meiotic cell divisions. Interestingly, we found that the lethality of smc-5; him-6 can be rescued by loss of the conserved BRCA1 ortholog BRC-1. Furthermore, the combined deletion of smc-5 and him-6 leads to an irregular distribution of condensin and to chromosome decondensation defects reminiscent of condensin depletion. Lethality conferred by condensin depletion can also be rescued by BRC-1 depletion. Our results suggest that SMC-5/6 and HIM-6 can synergistically regulate recombination intermediate metabolism and suppress ectopic recombination by controlling chromosome architecture during meiosis.

The pseudoautosomal region (PAR) is a short region of homology between the mammalian X and Y chromosomes, which has undergone rapid evolution. A crossover in the PAR is essential for the proper disjunction of X and Y chromosomes in male meiosis, and PAR deletion results in male sterility. This leads the human PAR with the obligatory crossover, PAR1, to having an exceptionally high male crossover rate, which is 17-fold higher than the genome-wide average. However, the mechanism by which this obligatory crossover occurs remains unknown, as does the fine-scale positioning of crossovers across this region. Recent research in mice has suggested that crossovers in PAR may be mediated independently of the protein PRDM9, which localises virtually all crossovers in the autosomes. To investigate recombination in this region, we construct the most fine-scale genetic map containing directly observed crossovers to date using African-American pedigrees. We leverage recombination rates inferred from the breakdown of linkage disequilibrium in human populations and investigate the signatures of DNA evolution due to recombination. Further, we identify direct PRDM9 binding sites using ChIP-seq in human cells. Using these independent lines of evidence, we show that, in contrast with mouse, PRDM9 does localise peaks of recombination in the human PAR1. We find that recombination is a far more rapid and intense driver of sequence evolution in PAR1 than it is on the autosomes. We also show that PAR1 hotspot activities differ significantly among human populations. Finally, we find evidence that PAR1 hotspot positions have changed between human and chimpanzee, with no evidence of sharing among the hottest hotspots. We anticipate that the genetic maps built and validated in this work will aid research on this vital and fascinating region of the genome.