Centro Andaluz de Biología Molecular y Medicina Regenerativa-CABIMER, Consejo Superior de Investigaciones Científicas; Universidad de Sevilla; Universidad Pablo de Olavide; Seville, Spain.
Institute of Molecular Biology (IMB), Mainz, Germany.
Cell Rep. 2021 Jul 27;36(4):109440. doi: 10.1016/j.celrep.2021.109440.
The minichromosome maintenance (MCM) helicase physically interacts with the recombination proteins Rad51 and Rad52 from yeast to human cells. We show, in Saccharomyces cerevisiae, that these interactions occur within a nuclease-insoluble scaffold enriched in replication/repair factors. Rad51 accumulates in a MCM- and DNA-binding-independent manner and interacts with MCM helicases located outside of the replication origins and forks. MCM, Rad51, and Rad52 accumulate in this scaffold in G1 and are released during the S phase. In the presence of replication-blocking lesions, Cdc7 prevents their release from the scaffold, thus maintaining the interactions. We identify a rad51 mutant that is impaired in its ability to bind to MCM but not to the scaffold. This mutant is proficient in recombination but partially defective in single-stranded DNA (ssDNA) gap filling and replication fork progression through damaged DNA. Therefore, cells accumulate MCM/Rad51/Rad52 complexes at specific nuclear scaffolds in G1 to assist stressed forks through non-recombinogenic functions.
微小染色体维持（MCM）解旋酶与来自酵母到人类细胞的重组蛋白 Rad51 和 Rad52 物理相互作用。我们在酿酒酵母中表明，这些相互作用发生在富含复制/修复因子的核酸内切酶不可溶支架内。Rad51 以 MCM 和 DNA 结合非依赖性的方式积累，并与位于复制起点和叉之外的 MCM 解旋酶相互作用。在 G1 期，MCM、Rad51 和 Rad52 在这个支架中积累，并在 S 期释放。在存在复制阻断损伤的情况下，Cdc7 阻止它们从支架中释放，从而维持相互作用。我们鉴定出一种 rad51 突变体，该突变体在与 MCM 结合但不与支架结合的能力上存在缺陷。该突变体在重组中是有效的，但在单链 DNA（ssDNA）缺口填充和复制叉通过受损 DNA的推进方面部分缺陷。因此，细胞在 G1 期在特定的核支架上积累 MCM/Rad51/Rad52 复合物，以通过非重组功能帮助受压的叉。
