https://scholars.lib.ntu.edu.tw/handle/123456789/635770
標題: | Hop2-Mnd1 and Swi5-Sfr1 stimulate Dmc1 filament assembly using distinct mechanisms | 作者: | Lee, Wei Iwasaki, Hiroshi Tsubouchi, Hideo HUNG-WEN LI |
公開日期: | 8-九月-2023 | 卷: | 51 | 期: | 16 | 起(迄)頁: | 8550 | 來源出版物: | Nucleic acids research | 摘要: | In meiosis, Dmc1 recombinase and the general recombinase Rad51 are responsible for pairing homologous chromosomes and exchanging strands. Fission yeast (Schizosaccharomyces pombe) Swi5-Sfr1 and Hop2-Mnd1 stimulate Dmc1-driven recombination, but the stimulation mechanism is unclear. Using single-molecule fluorescence resonance energy transfer (smFRET) and tethered particle motion (TPM) experiments, we showed that Hop2-Mnd1 and Swi5-Sfr1 individually enhance Dmc1 filament assembly on single-stranded DNA (ssDNA) and adding both proteins together allows further stimulation. FRET analysis showed that Hop2-Mnd1 enhances the binding rate of Dmc1 while Swi5-Sfr1 specifically reduces the dissociation rate during the nucleation, about 2-fold. In the presence of Hop2-Mnd1, the nucleation time of Dmc1 filaments shortens, and doubling the ss/double-stranded DNA (ss/dsDNA) junctions of DNA substrates reduces the nucleation times in half. Order of addition experiments confirmed that Hop2-Mnd1 binds on DNA to recruit and stimulate Dmc1 nucleation at the ss/dsDNA junction. Our studies directly support the molecular basis of how Hop2-Mnd1 and Swi5-Sfr1 act on different steps during the Dmc1 filament assembly. DNA binding of these accessory proteins and nucleation preferences of recombinases thus dictate how their regulation can take place. |
URI: | https://scholars.lib.ntu.edu.tw/handle/123456789/635770 | ISSN: | 0305-1048 1362-4962 |
DOI: | 10.1093/nar/gkad561 |
顯示於: | 化學系 |
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