title: Rad50 zinc hook is important for the Mre11 complex to bind chromosomal DNA double-stranded breaks and initiate various DNA damage responses.
abstract: The Mre11-Rad50-Nbs1 (MRN) complex plays critical roles in checkpoint activation and double-stranded break (DSB) repair. The Rad50 zinc hook domain mediates zinc-dependent intercomplex associations of MRN, which is important for DNA tethering. Studies in yeast suggest that the Rad50 zinc hook domain is essential for MRN functions, but its role in mammalian cells is not clear. We demonstrated that the human Rad50 hook mutants are severely defective in various DNA damage responses including ATM (Ataxia telangiectasia mutated) activation, homologous recombination, sensitivity to IR, and activation of the ATR pathway. By using live cell imaging, we observed that the Rad50 hook mutants fail to be recruited to chromosomal DSBs, suggesting a novel mechanism underlying the severe defects observed for the Rad50 hook mutants. In vitro analysis showed that Zn(2+) promotes wild type but not the hook mutant of MR to bind double-stranded DNA. In vivo, the Rad50 hook mutants are defective in being recruited to chromosomal DSBs in both H2AX-proficient and -deficient cells, suggesting that the Rad50 hook mutants are impaired in direct binding to chromosomal DSB ends. We propose that the Rad50 zinc hook domain is important for the initial binding of MRN to DSBs, leading to ATM activation to phosphorylate H2AX, which recruits more MRN to the DSB-flanking chromosomal regions. Our studies reveal a critical role for the Rad50 zinc hook domain in establishing and maintaining MRN recruitment to chromosomal DSBs and suggest an important mechanism of how the Rad50 zinc hook domain contributes to DNA repair and checkpoint activation.
authors: Department of Molecular and Experimental Medicine, The Scripps Research Institute, La, Jolla, California 92037, USA.
authors: Jing
authors: J
authors: He
authors: Linda Z
authors: LZ
authors: Shi
authors: Lan N
authors: LN
authors: Truong
authors: Chi-Sheng
authors: CS
authors: Lu
authors: Niema
authors: N
authors: Razavian
authors: Yongjiang
authors: Y
authors: Li
authors: Alejandro
authors: A
authors: Negrete
authors: Joseph
authors: J
authors: Shiloach
authors: Michael W
authors: MW
authors: Berns
authors: Xiaohua
authors: X
authors: Wu
mesh_terms: D017766:Acid Anhydride Hydrolases; D020816:Amino Acid Motifs; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000737:chemistry; D002469:Cell Separation; D002875:Chromosomes / Q000648:ultrastructure; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage; D045643:DNA Repair Enzymes / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D005434:Flow Cytometry; D020868:Gene Silencing; D016678:Genome; D023281:Genomics; D057809:HEK293 Cells; D006657:Histones / Q000737:chemistry; D006801:Humans; D000076228:MRE11 Homologue Protein; D008856:Microscopy, Fluorescence / Q000379:methods; D009154:Mutation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry; D011995:Recombination, Genetic; D025521:Tumor Suppressor Proteins / Q000737:chemistry; D015032:Zinc / Q000737:chemistry*
chemical_list: D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C468783:H2AX protein, human; D006657:Histones; C121766:MRE11 protein, human; 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; D015032:Zinc