Homologous recombination repair is regulated by domains at the N- and C-terminus of NBS1 and is dissociated with ATM functions.
The proteins responsible for radiation sensitive disorders, NBS1, kinase ataxia-telangiectasia-(A-T)-mutated (ATM) and MRE11, interact through the C-terminus of NBS1 in response to the generation of DNA double-strand breaks (DSBs) and are all implicated in checkpoint regulation and DSB repair, such as homologous recombination (HR). We measured the ability of several NBS1 mutant clones and A-T cells to regulate HR repair using the DR-GFP or SCneo systems. ATM deficiency did not reduce the HR repair frequency of an induced DSB, and it was confirmed by findings that HR frequencies are only slightly affected by deletion of ATM-binding site at the extreme C-terminus of NBS1. In contrast, The HR-regulating ability is dramatically reduced by deletion of the MRE11-binding domain at the C-terminus of NBS1 and markedly inhibited by mutations in the FHA/BRCT domains at the N-terminus. This impaired capability in HR is consistent with a failure to observe MRE11 foci formation. Furthermore, normal HR using sister chromatid was completely inhibited by the absence of FHA/BRCT domains. These results suggested that the N- and C-terminal domains of NBS1 are the major regulatory domains for HR pathways, very likely through the recruitment and retention of the MRE11 nuclease to DSB sites in an ATM-independent fashion.
Sakamoto
S
S
Department of Radiation Repair Dynamics, Radiation Biology Center, Kyoto University, Kyoto, Japan.
Iijima
K
K
Mochizuki
D
D
Nakamura
K
K
Teshigawara
K
K
Kobayashi
J
J
Matsuura
S
S
Tauchi
H
H
Komatsu
K
K
D000225:Adenine; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000235:genetics*; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics*; D005347:Fibroblasts / Q000502:physiology; D005787:Gene Frequency; D006801:Humans; D009687:Nuclear Proteins / Q000235:genetics*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics*; D011995:Recombination, Genetic; D013941:Thymine; D025521:Tumor Suppressor Proteins / Q000235:genetics*
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; 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; D000225:Adenine; D013941:Thymine