Dataset Information

The MCM8-MCM9 complex promotes RAD51 recruitment at DNA damage sites to facilitate homologous recombination.

ABSTRACT: The minichromosome maintenance protein homologs MCM8 and MCM9 have previously been implicated in DNA replication elongation and prereplication complex (pre-RC) formation, respectively. We found that MCM8 and MCM9 physically associate with each other and that MCM8 is required for the stability of MCM9 protein in mammalian cells. Depletion of MCM8 or MCM9 in human cancer cells or the loss of function MCM9 mutation in mouse embryo fibroblasts sensitizes cells to the DNA interstrand cross-linking (ICL) agent cisplatin. Consistent with a role in the repair of ICLs by homologous recombination (HR), knockdown of MCM8 or MCM9 significantly reduces HR repair efficiency. Chromatin immunoprecipitation analysis using human DR-GFP cells or Xenopus egg extract demonstrated that MCM8 and MCM9 proteins are rapidly recruited to DNA damage sites and promote RAD51 recruitment. Thus, these two metazoan-specific MCM homologs are new components of HR and may represent novel targets for treating cancer in combination with DNA cross-linking agents.

SUBMITTER: Park J

PROVIDER: S-EPMC3624244 | biostudies-literature | 2013 Apr

REPOSITORIES: biostudies-literature

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The MCM8-MCM9 complex promotes RAD51 recruitment at DNA damage sites to facilitate homologous recombination.

Park Jonghoon J Long David T DT Lee Kyung Yong KY Abbas Tarek T Shibata Etsuko E Negishi Masamitsu M Luo Yunhai Y Schimenti John C JC Gambus Agnieszka A Walter Johannes C JC Dutta Anindya A

Molecular and cellular biology 20130211 8

The minichromosome maintenance protein homologs MCM8 and MCM9 have previously been implicated in DNA replication elongation and prereplication complex (pre-RC) formation, respectively. We found that MCM8 and MCM9 physically associate with each other and that MCM8 is required for the stability of MCM9 protein in mammalian cells. Depletion of MCM8 or MCM9 in human cancer cells or the loss of function MCM9 mutation in mouse embryo fibroblasts sensitizes cells to the DNA interstrand cross-linking (I ...[more]

PMID: 23401855

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Project description:Classical non-homologous end joining (C-NHEJ) and homologous recombination (HR) compete to repair mammalian chromosomal double strand breaks (DSBs). However, C-NHEJ has no impact on HR induced by DNA nicking enzymes. In this case, the replication fork is thought to convert the DNA nick into a one-ended DSB, which lacks a readily available partner for C-NHEJ. Whether C-NHEJ competes with HR at a non-enzymatic mammalian replication fork barrier (RFB) remains unknown. We previously showed that conservative "short tract" gene conversion (STGC) induced by a chromosomal Tus/Ter RFB is a product of bidirectional replication fork stalling. This finding raises the possibility that Tus/Ter-induced STGC proceeds via a two-ended DSB intermediate. If so, Tus/Ter-induced STGC might be subject to competition by C-NHEJ. However, in contrast to the DSB response, where genetic ablation of C-NHEJ stimulates HR, we report here that Tus/Ter-induced HR is unaffected by deletion of either of two C-NHEJ genes, Xrcc4 or Ku70. These results show that Tus/Ter-induced HR does not entail the formation of a two-ended DSB to which C-NHEJ has competitive access. We found no evidence that the alternative end-joining factor, DNA polymerase ?, competes with Tus/Ter-induced HR. We used chromatin-immunoprecipitation to compare Rad51 recruitment to a Tus/Ter RFB and to a neighboring site-specific DSB. Rad51 accumulation at Tus/Ter was more intense and more sustained than at a DSB. In contrast to the DSB response, Rad51 accumulation at Tus/Ter was restricted to within a few hundred base pairs of the RFB. Taken together, these findings suggest that the major DNA structures that bind Rad51 at a Tus/Ter RFB are not conventional DSBs. We propose that Rad51 acts as an "early responder" at stalled forks, binding single stranded daughter strand gaps on the arrested lagging strand, and that Rad51-mediated fork remodeling generates HR intermediates that are incapable of Ku binding and therefore invisible to the C-NHEJ machinery.

Dataset Information

The MCM8-MCM9 complex promotes RAD51 recruitment at DNA damage sites to facilitate homologous recombination.

Publications

The MCM8-MCM9 complex promotes RAD51 recruitment at DNA damage sites to facilitate homologous recombination.

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