The Mre11/Rad50/Nbs1 complex plays an important role in the prevention of DNA rereplication in mammalian cells.
The Mre11/Nbs1/Rad50 complex (MRN) plays multiple roles in the maintenance of genome stability, including repair of double-stranded breaks (DSBs) and activation of the S-phase checkpoint. Here we demonstrate that MRN is required for the prevention of DNA rereplication in mammalian cells. DNA replication is strictly regulated by licensing control so that the genome is replicated once and only once per cell cycle. Inactivation of Nbs1 or Mre11 leads to a substantial increase of DNA rereplication induced by overexpression of the licensing factor Cdt1. Our studies reveal that multiple mechanisms are likely involved in the MRN-mediated suppression of rereplication. First, both Mre11 and Nbs1 are required for facilitating ATR activation when Cdt1 is overexpressed, which in turn suppresses rereplication. Second, Cdt1 overexpression induces ATR-mediated phosphorylation of Nbs1 at Ser343 and this phosphorylation depends on the FHA and BRCT domains of Nbs1. Mutations at Ser343 or in the FHA and BRCT domains lead to more severe rereplication when Cdt1 is overexpressed. Third, the interaction of the Mre11 complex with RPA is important for the suppression of rereplication. This suggests that modulating RPA activity via a direct interaction of MRN is likely one of the effector mechanisms to suppress rereplication. Moreover, we demonstrate that MRN is also required for preventing the accumulation of DSBs when rereplication is induced. Therefore, our studies suggest new roles of MRN in the maintenance of genome stability through preventing rereplication and rereplication-associated DSBs when licensing control is compromised.
Lee
Alan Yueh-Luen
AY
Department of Molecular Experimental Medicine, The Scripps Research Institute, La Jolla, California 92037, USA.
Liu
Enbo
E
Wu
Xiaohua
X
D017766:Acid Anhydride Hydrolases; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D045643:DNA Repair Enzymes / Q000378:metabolism*; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D000076228:MRE11 Homologue Protein; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D051716:Replication Protein A / Q000378:metabolism
C467199:CDT1 protein, human; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C121766:MRE11 protein, human; C112156:NBN protein, human; D009687:Nuclear Proteins; D051716:Replication Protein A; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D000076228:MRE11 Homologue Protein; D017766:Acid Anhydride Hydrolases; C112158:RAD50 protein, human; D045643:DNA Repair Enzymes