BRCA1 controls homologous recombination at Tus/Ter-stalled mammalian replication forks

摘要

Replication fork stalling can promote genomic instability, predisposing to cancer and other diseases. Stalled replication forks may be processed by sister chromatid recombination (SCR), generating error-free or error-prone homologous recombination (HR) outcomes. In mammalian cells, a long-standing hypothesis proposes that the major hereditary breast/ovarian cancer predisposition gene products, BRCA1 and BRCA2, control HR/SCR at stalled replication forks. Although BRCA1 and BRCA2 affect replication fork processing, direct evidence that BRCA gene products regulate homologous recombination at stalled chromosomal replication forks is lacking, due to a dearth of tools for studying this process. Here we report that the Escherichia coli Tus/Ter complex can be engineered to induce site-specific replication fork stalling and chromosomal HR/SCR in mouse cells. Tus/Ter-induced homologous recombination entails processing of bidirectionally arrested forks. We find that the Brcal car-boxy (C)-terminal tandem BRCT repeat and regions of Brcal encoded by exon 11 -two Brcal elements implicated in tumour suppression-control Tus/Ter-induced homologous recombination. Inactivation of either Brcal or Brca2 increases the absolute frequency of 'long-tract' gene conversions at Tus/Ter-stalled forks, an outcome not observed in response to a site-specific endonuclease-mediated chromosomal double-strand break. Therefore, homologous recombination at stalled forks is regulated differently from homologous recombination at double-strand breaks arising independently of a replication fork. We propose that aberrant long-tract homologous recombination at stalled replication forks contributes to genomic instability and breast/ovarian cancer predisposition in BRCA mutant cells.%一个复制叉的停滞能启动导致癌症的病理性事件。虽然已有人提出肿瘤抑制因子BRCA1和BRCA2也许会在停滞的复制叉上调控同源重组(HR),但此前却一直没有可能在哺乳动物细胞中获得关于这一点的直接证据，原因是没有一个能够在其中验证这一点的实验系统。Ralph Scully及同事现在建立了一个系统，它允许对哺乳动物细胞中某一特定的停滞复制叉进行调控。细菌Tus/Ter复合物能够造成一个复制叉的双向停滞，这需要由BRCA1/BRCA2介导的HR才能使其恢复。然而，异常的长段HR被发现在没有BRCA蛋白时会出现在停滞的复制叉上，这为这些细胞易于发生癌变提供了一个基础。