The Mre11-Rad50-Nbs1 complex acts both upstream and downstream of ataxia telangiectasia mutated and Rad3-related protein (ATR) to regulate the S-phase checkpoint following UV treatment.
The Mre11-Rad50-Nbs1 (MRN) complex is required for mediating the S-phase checkpoint following UV treatment, but the underlying mechanism is not clear. Here we demonstrate that at least two mechanisms are involved in regulating the S-phase checkpoint in an MRN-dependent manner following UV treatment. First, when replication forks are stalled, MRN ... is required upstream of ataxia telangiectasia mutated and Rad3-related protein (ATR) to facilitate ATR activation in a substrate and dosage-dependent manner. In particular, MRN is required for ATR-directed phosphorylation of RPA2, a critical event in mediating the S-phase checkpoint following UV treatment. Second, MRN is a downstream substrate of ATR. Nbs1 is phosphorylated by ATR at Ser-343 when replication forks are stalled, and this phosphorylation event is also important for down-regulating DNA replication following UV treatment. Moreover, we demonstrate that MRN and ATR/ATR-interacting protein (TRIP) interact with each other, and the forkhead-associated/breast cancer C-terminal domains (FHA/BRCT) of Nbs1 play a significant role in mediating this interaction. Mutations in the FHA/BRCT domains do not prevent ATR activation but specifically impair ATR-mediated Nbs1 phosphorylation at Ser-343, which results in a defect in the S-phase checkpoint. These data suggest that MRN plays critical roles both upstream and downstream of ATR to regulate the S-phase checkpoint when replication forks are stalled.
Mesh Terms:
Acid Anhydride Hydrolases, Ataxia Telangiectasia Mutated Proteins, Cell Cycle Proteins, Cell Line, Cell Line, Tumor, DNA Damage, DNA Repair Enzymes, DNA-Binding Proteins, Gene Expression Regulation, Neoplastic, Glutathione Transferase, Humans, MRE11 Homologue Protein, Nuclear Proteins, Phosphorylation, Protein Serine-Threonine Kinases, Protein Structure, Tertiary, S Phase, Serine, Ultraviolet Rays
Acid Anhydride Hydrolases, Ataxia Telangiectasia Mutated Proteins, Cell Cycle Proteins, Cell Line, Cell Line, Tumor, DNA Damage, DNA Repair Enzymes, DNA-Binding Proteins, Gene Expression Regulation, Neoplastic, Glutathione Transferase, Humans, MRE11 Homologue Protein, Nuclear Proteins, Phosphorylation, Protein Serine-Threonine Kinases, Protein Structure, Tertiary, S Phase, Serine, Ultraviolet Rays
J Biol Chem
Date: Aug. 03, 2007
PubMed ID: 17526493
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