Dataset Information

Arabidopsis meiotic-specific recombinase DMC1 IP-MS

ABSTRACT: To search for regulators of DMC1 stability, we generated DMC1-FLAG transgenic Arabidopsis plants and conducted immunoprecipitation-mass spectrometry (IP-MS) assay. 6,477 peptides corresponding to 2309 proteins were identified and 1009 proteins remained after removing background interactions using plants transformed with the corresponding empty vector (FLAG). GO term analysis was performed with the obtained data and revealed that ubiquitin proteasome pathway components are significantly enriched among the candidate proteins.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Arabidopsis Thaliana (mouse-ear Cress)

TISSUE(S): Plant Cell

SUBMITTER: Wanyue Xu

LAB HEAD: Yingxiang Wang

PROVIDER: PXD038126 | Pride | 2023-10-24

REPOSITORIES: Pride

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Publications

SCF<sup>RMF</sup> mediates degradation of the meiosis-specific recombinase DMC1.

Xu Wanyue W Yu Yue Y Jing Juli J Wu Zhen Z Zhang Xumin X You Chenjiang C Ma Hong H Copenhaver Gregory P GP He Yan Y Wang Yingxiang Y Wang Yingxiang Y

Nature communications 20230819 1

Meiotic recombination requires the specific RecA homolog DMC1 recombinase to stabilize strand exchange intermediates in most eukaryotes. Normal DMC1 levels are crucial for its function, yet the regulatory mechanisms of DMC1 stability are unknown in any organism. Here, we show that the degradation of Arabidopsis DMC1 by the 26S proteasome depends on F-box proteins RMF1/2-mediated ubiquitination. Furthermore, RMF1/2 interact with the Skp1 ortholog ASK1 to form the ubiquitin ligase complex SCF<sup> ...[more]

PMID: 37598222

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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

Dataset Information

Arabidopsis meiotic-specific recombinase DMC1 IP-MS

Publications

SCF<sup>RMF</sup> mediates degradation of the meiosis-specific recombinase DMC1.

Similar Datasets

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