Functional analysis of FHA and BRCT domains of NBS1 in chromatin association and DNA damage responses.
Rad50/Mre11/NBS1 (R/M/N) is a multi-functional protein complex involved in DNA repair, cell cycle checkpoint activation, DNA replication and replication block-induced responses. Ionizing radiation (IR) induces the phosphorylation of NBS1 and nuclear foci formation of the complex. Although it has been suggested that the R/M/N complex is associated with DNA damage sites, we present here biochemical evidence for chromatin association of the complex. We show that the chromatin association of R/M/N is independent of IR and ataxia telangiectasia mutated (ATM). We also demonstrate that optimal chromatin association of the Rad50/Mre11/NBS1 proteins requires both the conserved forkhead-associated (FHA) and breast cancer C-terminus (BRCT) domains of NBS1. Moreover, both these domains of NBS1 are required for its phosphorylation on Ser343 but not on Ser278. Importantly, both the FHA and BRCT domains are essential for IR-induced foci (IRIF) formation of R/M/N and S phase checkpoint activation, but only the BRCT domain is needed for cell survival after IR. These data demonstrate that the FHA and BRCT domains of NBS1 are crucial for the functions of the R/M/N complex.
Zhao
Song
S
Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center, San Antonio, TX 78245, USA.
Renthal
William
W
Lee
Eva Y-H P
EY
D017766:Acid Anhydride Hydrolases; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000502:physiology*; D002461:Cell Line, Transformed; D002470:Cell Survival / Q000528:radiation effects; D002843:Chromatin / Q000378:metabolism*; D004249:DNA Damage; D004260:DNA Repair; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D000076228:MRE11 Homologue Protein; D046911:Macromolecular Substances; D009687:Nuclear Proteins / Q000737:chemistry* / Q000502:physiology*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D017434:Protein Structure, Tertiary; D011839:Radiation, Ionizing; D016196:S Phase; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins
D018797:Cell Cycle Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; C121766:MRE11 protein, human; D046911:Macromolecular Substances; C112156:NBN protein, human; D009687:Nuclear Proteins; D025521:Tumor Suppressor Proteins; C576324:ATM 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