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Swi5-Sfr1 复合物通过两种不同的机制调节 Dmc1 和 Rad51 驱动的 DNA 链交换，该过程涉及两个不同的三链中间体。

The Swi5-Sfr1 complex regulates Dmc1- and Rad51-driven DNA strand exchange proceeding through two distinct three-stranded intermediates by different mechanisms.

机构信息

Institute of Innovative Research, Tokyo Institute of Technology, Yokohama, Kanagawa 226-8503, Japan.

Department of Life Science and Technology, School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Kanagawa 226-8503, Japan.

出版信息

Nucleic Acids Res. 2024 Nov 11;52(20):12517-12533. doi: 10.1093/nar/gkae841.

DOI:10.1093/nar/gkae841
PMID:39340300
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11551746/
Abstract

In eukaryotes, Dmc1 and Rad51 are key proteins of homologous recombination. The Swi5-Sfr1 complex in fission yeast, a conserved auxiliary factor, stimulates DNA strand exchange driven by both Dmc1 and Rad51. Interestingly, biochemical analysis suggested that Swi5-Sfr1 regulates strand exchange activities of these recombinases differently, but the mechanisms were unclear. We previously developed a real-time system to analyze Rad51-driven DNA strand exchange and identified two topologically distinct three-stranded intermediates (complex 1 (C1) and complex 2 (C2)). Swi5-Sfr1 facilitates the C1-C2 transition and releases single-stranded DNA (ssDNA) from C2, acting as a strand exchange activator. In this study, we investigated fission yeast Dmc1-driven DNA strand exchange and the role of Swi5-Sfr1 in Dmc1 activity in real-time. Kinetic analysis revealed a three-step model for the Dmc1-driven reaction, similar to that of Rad51. Although Swi5-Sfr1 stimulated the Dmc1-driven reaction, it had a weaker impact than Rad51. Furthermore, Swi5-Sfr1 enhanced the association of Dmc1 with ssDNA by promoting filament nucleus formation, acting as a mediator, unlike its role with Rad51. This stimulation mechanism also differs from that of Ca2+ or ATP analog, AMP-PNP. Our findings suggest that Swi5-Sfr1 stimulates strand exchange activities of Dmc1 and Rad51 via different reaction steps.

摘要

在真核生物中，Dmc1 和 Rad51 是同源重组的关键蛋白。裂殖酵母中的 Swi5-Sfr1 复合物是一种保守的辅助因子，可刺激 Dmc1 和 Rad51 驱动的 DNA 链交换。有趣的是，生化分析表明 Swi5-Sfr1 以不同的方式调节这些重组酶的链交换活性，但机制尚不清楚。我们之前开发了一种实时分析 Rad51 驱动的 DNA 链交换的系统，并鉴定了两种拓扑上不同的三链中间体（复合物 1（C1）和复合物 2（C2））。Swi5-Sfr1 促进 C1-C2 转换并从 C2 释放单链 DNA（ssDNA），充当链交换激活剂。在这项研究中，我们在实时研究了裂殖酵母 Dmc1 驱动的 DNA 链交换以及 Swi5-Sfr1 在 Dmc1 活性中的作用。动力学分析揭示了 Dmc1 驱动反应的三步骤模型，类似于 Rad51 的反应。尽管 Swi5-Sfr1 刺激了 Dmc1 驱动的反应，但它的影响比 Rad51 弱。此外，Swi5-Sfr1 通过促进丝核体形成来增强 Dmc1 与 ssDNA 的结合，作为一种介质，与 Rad51 的作用不同。这种刺激机制也不同于 Ca2+ 或 ATP 类似物 AMP-PNP。我们的研究结果表明，Swi5-Sfr1 通过不同的反应步骤刺激 Dmc1 和 Rad51 的链交换活性。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/e5658efc8bee/gkae841fig8.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/aedee7fde49f/gkae841figgra1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/79bf3a1f2271/gkae841fig1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/045f90a11488/gkae841fig2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/88988333794c/gkae841fig3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/6b61e7ed84ca/gkae841fig4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/50a1e01d6b18/gkae841fig5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/5f0e752a26f8/gkae841fig6.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/ca3bc6b05b1c/gkae841fig7.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/e5658efc8bee/gkae841fig8.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/aedee7fde49f/gkae841figgra1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/79bf3a1f2271/gkae841fig1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/045f90a11488/gkae841fig2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/88988333794c/gkae841fig3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/6b61e7ed84ca/gkae841fig4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/50a1e01d6b18/gkae841fig5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/5f0e752a26f8/gkae841fig6.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/ca3bc6b05b1c/gkae841fig7.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1727/11551746/e5658efc8bee/gkae841fig8.jpg

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本文引用的文献

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2
The role of bivalent ions in the regulation of D-loop extension mediated by DMC1 during meiotic recombination.二价离子在减数分裂重组过程中由DMC1介导的D环延伸调控中的作用。
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Biochemical properties of fission yeast homologous recombination enzymes.
裂殖酵母同源重组酶的生化特性。
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The Rad51 paralog complex Rad55-Rad57 acts as a molecular chaperone during homologous recombination.Rad51 同源物复合物 Rad55-Rad57 在同源重组过程中充当分子伴侣。
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Real-time tracking reveals catalytic roles for the two DNA binding sites of Rad51.实时追踪揭示了 Rad51 两个 DNA 结合位点的催化作用。
Nat Commun. 2020 Jun 11;11(1):2950. doi: 10.1038/s41467-020-16750-3.
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Rad51 facilitates filament assembly of meiosis-specific Dmc1 recombinase.Rad51 促进减数分裂特异性 Dmc1 重组酶的丝形成。
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A mutant form of Dmc1 that bypasses the requirement for accessory protein Mei5-Sae3 reveals independent activities of Mei5-Sae3 and Rad51 in Dmc1 filament stability.一种突变形式的 Dmc1 可以绕过辅助蛋白 Mei5-Sae3 的需求，揭示了 Mei5-Sae3 和 Rad51 在 Dmc1 丝稳定性中的独立活性。
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Real-time Observation of the DNA Strand Exchange Reaction Mediated by Rad51.由Rad51介导的DNA链交换反应的实时观察。
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