title: CtIP protein dimerization is critical for its recruitment to chromosomal DNA double-stranded breaks.
abstract: CtIP (CtBP-interacting protein) associates with BRCA1 and the Mre11-Rad50-Nbs1 (MRN) complex and plays an essential role in homologous recombination (HR)-mediated DNA double-stranded break (DSB) repair. It has been described that CtIP forms dimers in mammalian cells, but the biological significance is not clear. In this study, we identified a conserved motif in the N terminus of CtIP, which is required for dimer formation. We further showed that CtIP mutants impaired in forming dimers are strongly defective in HR, end resection, and activation of the ataxia telangiectasia and Rad3-related pathway, without notable change of CtIP interactions with BRCA1 or Nbs1. In addition to HR, CtIP dimerization is also required for microhomology-mediated end joining. Live cell imaging of enhanced GFP-tagged CtIP demonstrates that the CtIP dimerization mutant fails to be localized to DSBs, whereas placing a heterologous dimerization motif to the dimerization mutant restores CtIP recruitment to DSBs. These studies suggest that CtIP dimer formation is essential for its recruitment to DSBs on chromatin upon DNA damage. Furthermore, DNA damage-induced phosphorylation of CtIP is significantly reduced in the CtIP dimerization mutants. Therefore, in addition to the C-terminal conserved domains critical for CtIP function, the dimerization motif on the N terminus of CtIP is also conserved and essential for its function in DNA damage responses. The severe repair defects of CtIP dimerization mutants are likely due to the failure in localization to chromosomal DSBs upon DNA damage.
authors.affiliation: Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, California 92037, USA.
authors.forename: Hailong
authors.initials: H
authors.lastname: Wang
authors.forename: Zhengping
authors.initials: Z
authors.lastname: Shao
authors.forename: Linda Z
authors.initials: LZ
authors.lastname: Shi
authors.forename: Patty Yi-Hwa
authors.initials: PY
authors.lastname: Hwang
authors.forename: Lan N
authors.initials: LN
authors.lastname: Truong
authors.forename: Michael W
authors.initials: MW
authors.lastname: Berns
authors.forename: David J
authors.initials: DJ
authors.lastname: Chen
authors.forename: Xiaohua
authors.initials: X
authors.lastname: Wu
mesh_terms: D020816:Amino Acid Motifs; D019313:BRCA1 Protein / Q000235:genetics / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002877:Chromosomes, Human / Q000235:genetics / Q000378:metabolism*; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair / Q000502:physiology*; D004706:Endodeoxyribonucleases; D059765:Homologous Recombination / Q000502:physiology; D006801:Humans; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000502:physiology; D055503:Protein Multimerization / Q000502:physiology*; D017434:Protein Structure, Tertiary
chemical_list: D019313:BRCA1 Protein; C492913:BRCA1 protein, human; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; C112156:NBN protein, human; D009687:Nuclear Proteins; D004706:Endodeoxyribonucleases; C410741:RBBP8 protein, human