Department of Biology, Institute of Biochemistry, Eidgenössische Technische Hochschule (ETH), 8093, Zürich, Switzerland.
Institute for Research in Biomedicine, Università della Svizzera italiana (USI), Faculty of Biomedical Sciences, 6500, Bellinzona, Switzerland.
Nat Commun. 2022 May 2;13(1):2374. doi: 10.1038/s41467-022-29841-0.
The conserved Mre11-Rad50 complex is crucial for the detection, signaling, end tethering and processing of DNA double-strand breaks. While it is known that Mre11-Rad50 foci formation at DNA lesions accompanies repair, the underlying molecular assembly mechanisms and functional implications remained unclear. Combining pathway reconstitution in electron microscopy, biochemical assays and genetic studies, we show that S. cerevisiae Mre11-Rad50 with or without Xrs2 forms higher-order assemblies in solution and on DNA. Rad50 mediates such oligomerization, and mutations in a conserved Rad50 beta-sheet enhance or disrupt oligomerization. We demonstrate that Mre11-Rad50-Xrs2 oligomerization facilitates foci formation, DNA damage signaling, repair, and telomere maintenance in vivo. Mre11-Rad50 oligomerization does not affect its exonuclease activity but drives endonucleolytic cleavage at multiple sites on the 5'-DNA strand near double-strand breaks. Interestingly, mutations in the human RAD50 beta-sheet are linked to hereditary cancer predisposition and our findings might provide insights into their potential role in chemoresistance.
保守的 Mre11-Rad50 复合物对于 DNA 双链断裂的检测、信号转导、末端连接和加工至关重要。虽然已知 Mre11-Rad50 焦点在 DNA 损伤处的形成伴随着修复，但潜在的分子组装机制和功能意义仍不清楚。通过电子显微镜、生化测定和遗传研究相结合，我们表明酿酒酵母 Mre11-Rad50 无论是否带有 Xrs2，在溶液中和 DNA 上都形成更高阶的组装。Rad50 介导这种寡聚化，并且保守的 Rad50 β-折叠中的突变增强或破坏寡聚化。我们证明 Mre11-Rad50-Xrs2 寡聚化促进体内焦点形成、DNA 损伤信号转导、修复和端粒维持。Mre11-Rad50 寡聚化不影响其外切核酸酶活性，但在双链断裂附近的 5'-DNA 链上的多个位点驱动内切核酸酶切割。有趣的是，人类 RAD50 β-折叠中的突变与遗传性癌症易感性有关，我们的发现可能为它们在化学抗性中的潜在作用提供了一些见解。
