BACKGROUND: Vertebrate meiotic recombination events are concentrated in regions (hotspots) that display open chromatin marks, such as trimethylation of lysines 4 and 36 of histone 3 (H3K4me3 and H3K36me3). Mouse and human PRDM9 proteins catalyze H3K4me3 and H3K36me3 and determine hotspot positions, whereas other vertebrates lacking PRDM9 recombine in regions with chromatin already opened for another function, such as gene promoters. While these other vertebrate species lacking PRDM9 remain fertile, inactivation of the mouse Prdm9 gene, which shifts the hotspots to the functional regions (including promoters), typically causes gross fertility reduction; and the reasons for these species differences are not clear. RESULTS: We introduced Prdm9 deletions into the Rattus norvegicus genome and generated the first rat genome-wide maps of recombination-initiating double-strand break hotspots. Rat strains carrying the same wild-type Prdm9 allele shared 88% hotspots but strains with different Prdm9 alleles only 3%. After Prdm9 deletion, rat hotspots relocated to functional regions, about 40% to positions corresponding to Prdm9-independent mouse hotspots, including promoters. Despite the hotspot relocation and decreased fertility, Prdm9-deficient rats of the SHR/OlaIpcv strain produced healthy offspring. The percentage of normal pachytene spermatocytes in SHR-Prdm9 mutants was almost double than in the PWD male mouse oligospermic sterile mutants. We previously found a correlation between the crossover rate and sperm presence in mouse Prdm9 mutants. The crossover rate of SHR is more similar to sperm-carrying mutant mice, but it did not fully explain the fertility of the SHR mutants. Besides mild meiotic arrests at rat tubular stages IV (mid-pachytene) and XIV (metaphase), we also detected postmeiotic apoptosis of round spermatids. We found delayed meiosis and age-dependent fertility in both sexes of the SHR mutants. CONCLUSIONS: We hypothesize that the relative increased fertility of rat versus mouse Prdm9 mutants could be ascribed to extended duration of meiotic prophase I. While rat PRDM9 shapes meiotic recombination landscapes, it is unnecessary for recombination. We suggest that PRDM9 has additional roles in spermatogenesis and speciation-spermatid development and reproductive age-that may help to explain male-specific hybrid sterility.

• Keywords
• Fertility, Meiotic recombination, PRDM9, Rattus norvegicus,
• MeSH
• Chromatin MeSH
• DNA Breaks, Double-Stranded MeSH
• Fertility genetics   MeSH
• Histone-Lysine N-Methyltransferase genetics   MeSH
• Rats MeSH
• Meiosis * genetics   MeSH
• Mice MeSH
• Rats, Inbred SHR MeSH
• Spermatogenesis genetics   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Extramural MeSH
• Names of Substances
• Chromatin MeSH
• Histone-Lysine N-Methyltransferase MeSH
• prdm9 protein, mouse MeSH Browser

A hallmark of meiosis is the rearrangement of parental alleles to ensure genetic diversity in the gametes. These chromosome rearrangements are mediated by the repair of programmed DNA double-strand breaks (DSBs) as genetic crossovers between parental homologs. In mice, humans, and many other mammals, meiotic DSBs occur primarily at hotspots, determined by sequence-specific binding of the PRDM9 protein. Without PRDM9, meiotic DSBs occur near gene promoters and other functional sites. Studies in a limited number of mouse strains showed that functional PRDM9 is required to complete meiosis, but despite its apparent importance, Prdm9 has been repeatedly lost across many animal lineages. Both the reason for mouse sterility in the absence of PRDM9 and the mechanism by which Prdm9 can be lost remain unclear. Here, we explore whether mice can tolerate the loss of Prdm9 By generating Prdm9 functional knockouts in an array of genetic backgrounds, we observe a wide range of fertility phenotypes and ultimately demonstrate that PRDM9 is not required for completion of male meiosis. Although DSBs still form at a common subset of functional sites in all mice lacking PRDM9, meiotic outcomes differ substantially. We speculate that DSBs at functional sites are difficult to repair as a crossover and that by increasing the efficiency of crossover formation at these sites, genetic modifiers of recombination rates can allow for meiotic progression. This model implies that species with a sufficiently high recombination rate may lose Prdm9 yet remain fertile.

• MeSH
• X Chromosome MeSH
• Fertility genetics   physiology   MeSH
• Histone-Lysine N-Methyltransferase genetics   physiology   MeSH
• Meiosis * MeSH
• Mice, Inbred C57BL MeSH
• Mice, Knockout MeSH
• Mice MeSH
• Spermatogenesis physiology   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, N.I.H., Intramural MeSH
• Names of Substances
• Histone-Lysine N-Methyltransferase MeSH
• prdm9 protein, mouse MeSH Browser

Hybrid sterility is one of the reproductive isolation mechanisms leading to speciation. Prdm9, the only known vertebrate hybrid-sterility gene, causes failure of meiotic chromosome synapsis and infertility in male hybrids that are the offspring of two mouse subspecies. Within species, Prdm9 determines the sites of programmed DNA double-strand breaks (DSBs) and meiotic recombination hotspots. To investigate the relation between Prdm9-controlled meiotic arrest and asynapsis, we inserted random stretches of consubspecific homology on several autosomal pairs in sterile hybrids, and analyzed their ability to form synaptonemal complexes and to rescue male fertility. Twenty-seven or more megabases of consubspecific (belonging to the same subspecies) homology fully restored synapsis in a given autosomal pair, and we predicted that two or more DSBs within symmetric hotspots per chromosome are necessary for successful meiosis. We hypothesize that impaired recombination between evolutionarily diverged chromosomes could function as one of the mechanisms of hybrid sterility occurring in various sexually reproducing species.

• Keywords
• Prdm9, chromosomes, evolutionary biology, gene expression, genomics, homology-dependent meiotic chromosome pairing, mouse, speciation, synaptonemal complex,
• MeSH
• Biological Evolution * MeSH
• Chimera genetics   MeSH
• Chromosomes genetics   MeSH
• DNA Breaks, Double-Stranded MeSH
• Histone-Lysine N-Methyltransferase genetics   MeSH
• Hybridization, Genetic MeSH
• Infertility genetics   MeSH
• Meiosis genetics   MeSH
• Infertility, Male genetics   MeSH
• Mice MeSH
• Chromosome Pairing genetics   MeSH
• Recombination, Genetic MeSH
• Reproductive Isolation MeSH
• Synaptonemal Complex genetics   MeSH
• Genetic Speciation MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Histone-Lysine N-Methyltransferase MeSH
• prdm9 protein, mouse MeSH Browser

In mammals, meiotic recombination occurs at 1- to 2-kb genomic regions termed hotspots, whose positions and activities are determined by PRDM9, a DNA-binding histone methyltransferase. We show that the KRAB domain of PRDM9 forms complexes with additional proteins to allow hotspots to proceed into the next phase of recombination. By a combination of yeast-two hybrid assay, in vitro binding, and coimmunoprecipitation from mouse spermatocytes, we identified four proteins that directly interact with PRDM9's KRAB domain, namely CXXC1, EWSR1, EHMT2, and CDYL. These proteins are coexpressed in spermatocytes at the early stages of meiotic prophase I, the limited period when PRDM9 is expressed. We also detected association of PRDM9-bound complexes with the meiotic cohesin REC8 and the synaptonemal complex proteins SYCP3 and SYCP1. Our results suggest a model in which PRDM9-bound hotspot DNA is brought to the chromosomal axis by the action of these proteins, ensuring the proper chromatin and spatial environment for subsequent recombination events.

• MeSH
• Chromatin metabolism   MeSH
• Chromosomes genetics   physiology   MeSH
• DNA metabolism   MeSH
• DNA Breaks, Double-Stranded MeSH
• Genome MeSH
• Histone-Lysine N-Methyltransferase genetics   metabolism   physiology   MeSH
• Homologous Recombination MeSH
• Meiosis physiology   MeSH
• Mice MeSH
• Protein Domains MeSH
• Recombination, Genetic physiology   MeSH
• Spermatocytes metabolism   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Chromatin MeSH
• DNA MeSH
• Histone-Lysine N-Methyltransferase MeSH
• PRDM9 protein, human MeSH Browser