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Meiotic DNA breaks activate a streamlined response that largely avoids protein level changes -DIA

ABSTRACT: To understand the proteome and phosphoproteome level responses to meiotic DNA breaks, we have compared protein phosphorylation and protein levels in cells spo11-YF (cannot form meiotic DNA breaks) cells to SPO11 (can form meiotic breaks) cells.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Saccharomyces Cerevisiae (baker's Yeast)

TISSUE(S): Diploid Cell

SUBMITTER: Funda Kar

LAB HEAD: Christine Vogel

PROVIDER: PXD031781 | Pride | 2022-08-15

REPOSITORIES: pride

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			Action	DRS
	190620_h10355_rep1_phospho_dia.raw	Raw
	190620_h10423_rep1_phospho_dia.raw	Raw
	190621_h10423_rep2_phospho_dia.raw	Raw
	190622_h10355_rep2_phospho_dia.raw	Raw
	190623_h10355_rep3_phospho_dia.raw	Raw

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Project description:DNA double-strand breaks (DSBs) initiate meiotic recombination. Past DSB-mapping studies have used rad50S or sae2? mutants, which are defective in break processing, to accumulate DSBs, and report large (= 50 kb) “DSB-hot” regions that are separated by “DSB-cold” domains of similar size. Substantial recombination occurs in some DSB-cold regions, suggesting that DSB patterns are not normal in rad50S or sae2? mutants. We therefore developed novel methods that detect DSBs using ssDNA enrichment and microarray hybridization, and that use background-based normalization to allow cross-comparison between array datasets, to map genome-wide the DSBs that accumulate in processing-capable, repair-defective dmc1î and dmc1î rad51î mutants. DSBs were observed at known hotspots, but also in most previously-identified “DSB-cold” regions, including near centromeres and telomeres. While about 40% of the genome is DSB-cold in rad50S mutants, analysis of meiotic ssDNA from dmc1? shows that most of these regions have significant DSB activity. Thus, DSBs are distributed much more uniformly than was previously believed. Southern-blot assays of DSBs in selected regions in dmc1?, rad50S and wild-type cells confirm these findings. Comparisons of DSB signals in dmc1, dmc1 rad51, and dmc1 spo11 mutant strains identify Dmc1 as the primary strand transfer activity genome-wide, and Spo11-induced lesions as initiating all meiotic recombination. Keywords: DSB mapping, ChIP-chip, single strand DNA , BND cellulose We use two different strategies to map the genome-wide distribution of meiotic DSBs in the yeast Saccharomyces cerevisiae. The first is a chromatin immunoprecipitation (ChIP) based approach that targets the Spo11p protein, which remains covalently attached to DSB ends in the rad50S mutant background. The second approach involves BND cellulose enrichment of the single strand DNA (ssDNA) recombination intermediate formed by end-resection at DSB sites following Spo11p removal. We use dmc1 and dmc1 rad51 mutants that accumulates meiotic single strand DNA intermediates

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Meiotic DNA breaks activate a streamlined response that largely avoids protein level changes -DIA

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