Meiotic recombination differs between males and females; however, when and how these differences are established is unknown. Here we identify extensive sex differences at the initiation of recombination by mapping hotspots of meiotic DNA double-strand breaks in male and female mice. Contrary to past findings in humans, few hotspots are used uniquely in either sex. Instead, grossly different recombination landscapes result from up to fifteen-fold differences in hotspot usage between males and females. Indeed, most recombination occurs at sex-biased hotspots. Sex-biased hotspots seem to be partly determined by chromosome structure, and DNA methylation, which is absent in females at the onset of meiosis, has a substantial role. Sex differences are also evident later in meiosis as the rate at which meiotic breaks are repaired as crossovers differs between males and females in distal regions. The suppression of distal crossovers may help to minimize age-related aneuploidy that arises owing to cohesion loss during dictyate arrest in females.

Homologous recombination is required for proper segregation of homologous chromosomes during meiosis. It occurs predominantly at recombination hotspots that are defined by the DNA binding specificity of the PRDM9 protein. PRDM9 contains three conserved domains typically involved in regulation of transcription; yet, the role of PRDM9 in gene expression control is not clear. Here, we analyze the germline transcriptome of Prdm9−/− male mice in comparison to Prdm9+/+ males and find no apparent differences in the mRNA and miRNA profiles. We further explore the role of PRDM9 in meiosis by analyzing the effect of the KRAB, SSXRD, and post-SET zinc finger deletions in a cell culture expression system and the KRAB domain deletion in mice. We found that although the post-SET zinc finger and the KRAB domains are not essential for the methyltransferase activity of PRDM9 in cell culture, the KRAB domain mutant mice show only residual PRDM9 methyltransferase activity and undergo meiotic arrest. In aggregate, our data indicate that domains typically involved in regulation of gene expression do not serve that role in PRDM9, but are likely involved in setting the proper chromatin environment for initiation and completion of homologous recombination.

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Widespread indications for use of molecular diagnostics in various aspects of clinical medicine have driven proliferation of testing. The rapid adoption and continuous technological evolution of molecular diagnostics have often strained the development and maintenance of a functional underlying framework of coding, coverage, and reimbursement policies, thereby presenting challenges to various stakeholders, including molecular professionals, payers, and patients. A multidisciplinary working group convened by the Association for Molecular Pathology Economic Affairs Committee was tasked to describe the complex landscape of molecular pathology economics and highlight opportunities for member engagement. In this article, on the basis of review and synthesis of government Supported exclusively by the Association for Molecular Pathology.

13Homologous recombination in meiosis is initiated by programmed DNA double strand 14 breaks (DSBs) and DSB repair as a crossover is essential to prevent chromosomal 15 abnormalities in gametes. Sex differences in recombination have been previously 16 observed by analyses of recombination end-products. To understand when and how 17 sex differences are established, we built genome-wide maps of meiotic DSBs in both 18 male and female mice. We found that most recombination initiates at sex-biased DSB 19 hotspots. Local context, the choice of DSB targeting pathway and sex-specific patterns 20 of DNA methylation give rise to these differences. Sex differences are not limited to the 21 initiation stage, as the rate at which DSBs are repaired as crossovers appears to differ 22 between the sexes in distal regions. This uneven repair patterning may be linked to the 23 higher aneuploidy rate in females. Together, these data demonstrate that sex 24 differences occur early in meiotic recombination. 25 Main text 26Genetic recombination brings homologous chromosomes together during meiotic 27 prophase and facilitates their orderly segregation at the first meiotic division. 28Recombination is initiated by programmed DNA double strand breaks (DSBs) that are 29 subsequently repaired as either crossovers (COs) or as non-crossovers (NCOs). The 30 pattern and frequency of recombination can differ between males and females of the 31 same species: the female CO rate is higher in humans and mice 1 while in most studied 32 mammals COs are highly concentrated at sub-telomeric regions in males, but not in 33 females 2-6 . This pattern is not universal however and in some species, such as pigs, 34 3 subtelomeric crossovers are elevated in females 7 . To date, sex differences in 35 recombination have been studied by comparing the genetic end products of 36 recombination, primarily COs, between the sexes. However, CO-based estimates of 37 recombination rates are limited by the number of sampled meioses and the maximum 38 resolution of such analyses is determined by the spacing of sequence polymorphisms. 39Most importantly, sex-specific variation that manifests at the initiation of meiotic 40 recombination has not been studied and it remains an open question whether sex 41 differences originate at the time of DSB formation or whether they arise later, during 42 DSB repair. We therefore generated quantitative, high resolution and genome-wide 43 maps of meiotic DSBs in both male and female mice to examine when and where sex 44 biases in recombination are established, and to elucidate the mechanism(s) that give 45 rise to these biases. 46To map meiotic DSBs in female meiosis we exploited a method we previously 47 developed 8 to map DSB hotspots in mouse 9-11 and human 12 males. This variant of 48 ChIP-Seq (single stranded DNA sequencing, SSDS) detects single stranded DNA 49 (ssDNA) bound to the DMC1 protein, an early intermediate in the DSB repair 50 process 12,13 . In female mice, meiotic DSBs form in the fetal ovary and the number of 51 cells ...