title: "DNA Binding Region" of BRCA1 Affects Genetic Stability through modulating the Intra-S-Phase Checkpoint.
abstract: The breast cancer associated gene 1 (BRCA1) contains 3 domains: an N-terminal RING domain with ubiquitin E3 ligase activity, C-terminal BRCT protein interaction domain and a central region. RING and BRCT domains are well characterized, yet the function of the central region remains unclear. In this study, we identified an essential DNA binding region (DBR: 421-701 amino acids) within the central region of human BRCA1, and found that BRCA1 brings DNA together and preferably binds to splayed-arm DNA in a sequence-independent manner. To investigate the biological role of the DBR, we generated mouse ES cells, which lack the DBR (ΔDBR) by using the TALEN method. The ΔDBR cells exhibited decreased survival as compared to the wild type (WT) cells treated with a PARP inhibitor, however they have an intact ability to conduct DNA repair mediated by homologous recombination (HR). The ΔDBR cells continued to incorporate more EdU in the presence of hydroxyurea (HU), which causes replication stress and exhibited reduced viability than the WT cells. Moreover, phosphorylation of CHK1, which regulates the intra-S phase checkpoint, was moderately decreased in ΔDBR cells. These data suggest that DNA binding by BRCA1 affects the stability of DNA replication folks, resulting in weakened intra-S-phase checkpoint control in the ΔDBR cells. The ΔDBR cells also exhibited an increased number of abnormal chromosome structures as compared with WT cells, indicating that the ΔDBR cells have increased genetic instability. Thus, we demonstrated that the DBR of BRCA1 modulates genetic stability through the intra-S-phase checkpoint activated by replication stress.
authors.affiliation: 1. Genetics of Development and Disease Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, USA.
authors.forename: Takaaki
authors.initials: T
authors.lastname: Masuda
authors.affiliation: 2. Faculty of Health Sciences, University of Macau, Macau SAR, China.
authors.forename: Xiaoling
authors.initials: X
authors.lastname: Xu
authors.affiliation: 3. Laboratory of Bioengineering and Physical Science, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, USA.
authors.forename: Emilios K
authors.initials: EK
authors.lastname: Dimitriadis
authors.affiliation: 1. Genetics of Development and Disease Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, USA.
authors.forename: Tyler
authors.initials: T
authors.lastname: Lahusen
authors.affiliation: 1. Genetics of Development and Disease Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, USA.; 2. Faculty of Health Sciences, University of Macau, Macau SAR, China.
authors.forename: Chu-Xia
authors.initials: CX
authors.lastname: Deng
mesh_terms: D000818:Animals; D019313:BRCA1 Protein / Q000737:chemistry*; D001665:Binding Sites; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D002869:Chromosome Aberrations; D004260:DNA Repair; D004261:DNA Replication / Q000235:genetics / Q000502:physiology; D004268:DNA-Binding Proteins / Q000737:chemistry*; D006801:Humans; D051379:Mice; D018625:Microscopy, Atomic Force; D016297:Mutagenesis, Site-Directed; D000072417:Protein Domains; D059807:S Phase Cell Cycle Checkpoints; D020539:Sequence Analysis, Protein
chemical_list: D019313:BRCA1 Protein; C492913:BRCA1 protein, human; D004268:DNA-Binding Proteins
keywords: BRCA1; DNA binding; Genomic instability; Intra-S-phase checkpoint