The forkhead-associated domain of NBS1 is essential for nuclear foci formation after irradiation but not essential for hRAD50[middle dot]hMRE11[middle dot]NBS1 complex DNA repair activity.
NBS1 (p95), the protein responsible for Nijmegen breakage syndrome, shows a weak homology to the yeast Xrs2 protein at the N terminus region, known as the forkhead-associated (FHA) domain and the BRCA1 C terminus domain. The protein interacts with hMRE11 to form a complex with a nuclease activity for initiation of both nonhomologous end joining and homologous recombination. Here, we show in vivo direct evidence that NBS1 recruits the hMRE11 nuclease complex into the cell nucleus and leads to the formation of foci by utilizing different functions from several domains. The amino acid sequence at 665-693 on the C terminus of NBS1, where a novel identical sequence with yeast Xrs2 protein was found, is essential for hMRE11 binding. The hMRE11-binding region is necessary for both nuclear localization of the complex and for cellular radiation resistance. On the other hand, the FHA domain regulates nuclear foci formation of the multiprotein complex in response to DNA damage but is not essential for nuclear transportation of the complex and radiation resistance. Because the FHA/BRCA1 C terminus domain is widely conserved in eukaryotic nuclear proteins related to the cell cycle, gene regulation, and DNA repair, the foci formation could be associated with many phenotypes of Nijmegen breakage syndrome other than radiation sensitivity.
Tauchi
H
H
Department of Radiation Biology, Research Institute for Radiation Biology and Medicine, Hiroshima University, Kasumi 1-2-3, Minami-ku, Hiroshima 734-8553, Japan.
Kobayashi
J
J
Morishima
K
K
Matsuura
S
S
Nakamura
A
A
Shiraishi
T
T
Ito
E
E
Masnada
D
D
Delia
D
D
Komatsu
K
K
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism* / Q000528:radiation effects; D002645:Chickens; D004249:DNA Damage / Q000528:radiation effects; D004260:DNA Repair; D004268:DNA-Binding Proteins; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; D005455:Fluorescent Antibody Technique; D005656:Fungal Proteins / Q000378:metabolism*; D005720:Gamma Rays; D006801:Humans; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D014162:Transfection; D020798:Two-Hybrid System Techniques
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D046911:Macromolecular Substances; D046912:Multiprotein Complexes; D009687:Nuclear Proteins; C086930:RAD50 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; C094191:MRE11 protein, S cerevisiae