Project description:During sexual reproduction half of the genetic material is deposited in gametes and a complete set of chromosomes is restored upon fertilisation. Reduction of the genetic information prior gametogenesis occurs in meiosis where crossovers (COs) between homologous chromosomes secure an exchange of their genetic information. COs are not evenly distributed along chromosomes and are suppressed in chromosomal regions encompassing compact, hypermethylated centromeric and pericentromeric DNA. Therefore, it has been postulated that DNA hypermethylation is inhibitory to COs. Here by analysing meiotic recombination in mutant plants with hypomethylated DNA we observed unexpected and counterintuitive effects of the DNA methylation losses on COs distribution, further promoting recombination in the naturally hypomethylated euchromatic chromosome arms, while inhibiting it in heterochromatic regions encompassing centromeric and pericentromeric hypermethylated DNA. Importantly, the total number of COs was not affected, implying that the loss of DNA methylation only led to a global redistribution of COs along chromosomes. To determine by which mechanisms altered levels of DNA methylation influenced recombination, namely whether this occurred directly in cis or indirectly in trans by changing expression of genes encoding recombination components, we analysed COs distribution in WT lines with randomly scattered and well mapped hypomethylated chromosomal segments. Results of these experiments, supported by expression profiling data, suggest that DNA methylation affects meiotic recombination in cis. As DNA methylation is subjected to significant variation even within a single species, our results implicate that it could influence evolution of plant genomes through the control of meiotic recombination. 2 samples: Col, epiRIL12, with 3 replicates each
Project description:The diploid isolate EM93 is the main ancestor to the widely used Saccharomyces cerevisiae haploid laboratory strain, S288C. In this study, we first hybridise DNA from eight EM93 segregants from two tetrads to Affymetrix genechip S. cerevisiae Tiling 1.0R Arrays to generate a high-resolution overview of the genetic differences between EM93 and S288C. We show that EM93 is heterozygous for many polymorphisms, including large sequence polymorphisms, such as deletions and a Saccharomyces paradoxus introgression. We also find that many of the large sequence polymorphisms are associated with Ty-elements and sub-telomeric regions. We next utilize the hybridization profiles of the 3.2 million Tiling Array probes to identify 2,965 genetic markers, which we then used to design an EM93 genotyping array. By genotyping 120 EM93 tetrads, we deduced the structures of all EM93 chromosomes and found that the average EM93 meiosis produces 144 detectable recombination events, consisting of 87 crossover and 57 non-crossover events; we also identified 203 recombination hot-spots and provide evidence for crossover interference. We find that the effect of heterozygous large sequence polymorphisms on recombination is chromosome position-dependent, with sub-telomeric large sequence polymorphisms having no discernable effect on recombination but non-sub-telomeric large sequence polymorphisms reducing recombination. Finally, we find that recombination hotspots show limited conservation, with some but not all strain-specific hotspots being due to heterozygous non-sub-telomeric large sequence polymorphisms.
Project description:During sexual reproduction half of the genetic material is deposited in gametes and a complete set of chromosomes is restored upon fertilisation. Reduction of the genetic information prior gametogenesis occurs in meiosis where crossovers (COs) between homologous chromosomes secure an exchange of their genetic information. COs are not evenly distributed along chromosomes and are suppressed in chromosomal regions encompassing compact, hypermethylated centromeric and pericentromeric DNA. Therefore, it has been postulated that DNA hypermethylation is inhibitory to COs. Here by analysing meiotic recombination in mutant plants with hypomethylated DNA we observed unexpected and counterintuitive effects of the DNA methylation losses on COs distribution, further promoting recombination in the naturally hypomethylated euchromatic chromosome arms, while inhibiting it in heterochromatic regions encompassing centromeric and pericentromeric hypermethylated DNA. Importantly, the total number of COs was not affected, implying that the loss of DNA methylation only led to a global redistribution of COs along chromosomes. To determine by which mechanisms altered levels of DNA methylation influenced recombination, namely whether this occurred directly in cis or indirectly in trans by changing expression of genes encoding recombination components, we analysed COs distribution in WT lines with randomly scattered and well mapped hypomethylated chromosomal segments. Results of these experiments, supported by expression profiling data, suggest that DNA methylation affects meiotic recombination in cis. As DNA methylation is subjected to significant variation even within a single species, our results implicate that it could influence evolution of plant genomes through the control of meiotic recombination.
Project description:Genetic recombination generates novel trait combinations, and understanding how recombination is distributed across the genome is key to modern genetics. The PRDM9 protein defines recombination hotspots; however, megabase-scale recombination patterning is independent of PRDM9. The single round of DNA replication, which precedes recombination in meiosis, may establish these patterns; therefore, we devised an approach to study meiotic replication that includes robust and sensitive mapping of replication origins. We find that meiotic DNA replication is distinct; reduced origin firing slows replication in meiosis, and a distinctive replication pattern in human males underlies the subtelomeric increase in recombination. We detected a robust correlation between replication and both contemporary and historical recombination and found that replication origin density coupled with chromosome size determines the recombination potential of individual chromosomes. Our findings and methods have implications for understanding the mechanisms underlying DNA replication, genetic recombination, and the landscape of mammalian germline variation.
Project description:B lymphopoiesis requires that immunoglobulin genes be accessible to RAG1-RAG2 recombinase. However, the RAG proteins bind widely to open chromatin, which suggests that additional mechanisms must restrict RAG-mediated DNA cleavage. Here we show that developmental downregulation of interleukin 7 (IL-7)-receptor signaling in small pre-B cells induced expression of the bromodomain-family member BRWD1, which was recruited to a specific epigenetic landscape at Igk dictated by pre–BCR-dependent Erk activation. BRWD1 enhanced RAG recruitment, increased gene accessibility and positioned nucleosomes 5? to each J? recombination signal sequence. BRWD1 thus targets recombination to Igk and places recombination within the context of signaling cascades that control B cell development. Our findings represent a paradigm in which,at any particular antigen-receptor locus, specialized mechanisms enforce lineage- and stage-specific recombination. ChIP-seq for 1 transcription factor and 2 histone modifications in flow purified mouse small pre-B cells. ATAC-seq and RNA-seq in WT and Brwd-Mut mouse flow purified small pre-B cells.
Project description:High grade serous cancer (HGSC) is frequently characterizsed by homologous recombination (HR) DNA repair deficiency, and while most such tumours are sensitive to initial treatment, acquired resistance is common. We undertook a multi-omics approach to interrogating mechanisms of resistance, using multiple autopsy samples collected from 15 women with HR-deficient HGSC. We observed frequent reversion mutations, resistance mechanisms restoring HR by other means, a high frequency of whole-genome duplication (WGD) suggestive of an evolutionary advantage, and global changes in immune composition with evidence of immune escape. Collectively these findings have implications for therapeutic intervention for HR-deficient HGSC.