title: Mutations in the BRCT binding site of BRCA1 result in hyper-recombination.
abstract: We introduced a K1702M mutation in the BRCA1 BRCT domain known to prevent the binding of proteins harboring pS-X-X-F motifs such as Abraxas-RAP80, BRIP1, and CtIP. Surprisingly, rather than impairing homologous recombination repair (HRR), expression of K1702M resulted in hyper-recombination coinciding with an accumulation of cells in S-G2 and no effect on nonhomologous end-joining. These cells also showed increased RAD51 and RPA nuclear staining. More pronounced effects were seen with a naturally occurring BRCT mutant (M1775R) that also produced elevated levels of ssDNA, in part co-localizing with RPA, in line with excessive DNA resection. M1775R induced unusual, thread-like promyelocytic leukemia (PML) nuclear bodies and clustered RPA foci rather than the typical juxtaposed RPA-PML foci seen with wild-type BRCA1. Interestingly, K1702M hyper-recombination diminished with a second mutation in the BRCA1 RING domain (I26A) known to reduce BRCA1 ubiquitin-ligase activity. Thesein vitro findings correlated with elevated nuclear RAD51 and RPA staining of breast cancer tissue from a patient with the M1775R mutation. Altogether, the disruption of BRCA1 (BRCT)-pS-X-X-F protein binding results in ubiquitination-dependent hyper-recombination via excessive DNA resection and the appearance of atypical PML-NBs. Thus, certain BRCA1 mutations that cause hyper-recombination instead of reduced DSB repair might lead to breast cancer.
authors.affiliation: Department of Radiation Oncology, Virginia Commonwealth University, Richmond, 23298, USA.
authors.forename: Seth M
authors.initials: SM
authors.lastname: Dever
authors.forename: Sarah E
authors.initials: SE
authors.lastname: Golding
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Rosenberg
authors.forename: Bret R
authors.initials: BR
authors.lastname: Adams
authors.forename: Michael O
authors.initials: MO
authors.lastname: Idowu
authors.forename: John M
authors.initials: JM
authors.lastname: Quillin
authors.forename: Nicholas
authors.initials: N
authors.lastname: Valerie
authors.forename: Bo
authors.initials: B
authors.lastname: Xu
authors.forename: Lawrence F
authors.initials: LF
authors.lastname: Povirk
authors.forename: Kristoffer
authors.initials: K
authors.lastname: Valerie
mesh_terms: D020816:Amino Acid Motifs; D019313:BRCA1 Protein / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites; D001943:Breast Neoplasms / Q000235:genetics; D002352:Carrier Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004260:DNA Repair; D004706:Endodeoxyribonucleases; D005260:Female; D005822:Genetic Vectors; D057809:HEK293 Cells; D006801:Humans; D009154:Mutation; D009687:Nuclear Proteins / Q000378:metabolism; D000071156:Promyelocytic Leukemia Protein; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051135:Rad51 Recombinase / Q000378:metabolism; D011995:Recombination, Genetic; D014157:Transcription Factors / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000378:metabolism; D054875:Ubiquitination
chemical_list: D019313:BRCA1 Protein; D002352:Carrier Proteins; D009687:Nuclear Proteins; D000071156:Promyelocytic Leukemia Protein; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C070576:PML protein, human; D004247:DNA; D051135:Rad51 Recombinase; D004706:Endodeoxyribonucleases; C410741:RBBP8 protein, human