Premium
A novel model to characterize structure and function of BRCA1
Author(s) -
Lin Dong,
Izadpanah Reza,
Braun Stephen E.,
Alt Eckhard
Publication year - 2018
Publication title -
cell biology international
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.932
H-Index - 77
eISSN - 1095-8355
pISSN - 1065-6995
DOI - 10.1002/cbin.10846
Subject(s) - homologous recombination , biology , dna repair , recombineering , dna damage , gene , mutant , genetics , dna , microbiology and biotechnology
BRCA1 plays a central role in DNA repair. Although N‐terminal RING and C‐terminal BRCT domains are studied well, the functions of the central region of BRCA1 are poorly characterized. Here, we report a structural and functional analysis of BRCA1 alleles and functional human BRCA1 in chicken B‐lymphocyte cell line DT40. The combination of “homologous recombineering” and “RT‐cassette” enables modifications of chicken BRCA1 gene in Escherichia coli . Mutant BRCA1 knock‐in DT40 cell lines were generated using BRCA1 mutation constructs by homologous recombination with a targeting efficiency of up to 100%. Our study demonstrated that deletion of motifs 2–9 BRCA1 Δ/Δ181‐1415 ( Caenorhabditis elegans BRCA1 mimic) or deletion of motif 1 BRCA1 Δ/Δ126‐136 decreased cell viability following cisplatin treatment. Furthermore, deletion of motifs 5 and 6 BRCA1 Δ/Δ525‐881 within DNA‐binding region, even the conserved 7‐amino acid deletion BRCA1 Δ/Δ872‐878 within motif 6, caused a decreased cell viability upon cisplatin treatment. Surprisingly, human BRCA1 is functional in DT40 cells as indicated by DNA damage‐induced Rad 51 foci formation in human BRCA1 knock‐in DT40 cells. These results demonstrate that those conserved motifs within the central region are essential for DNA repair functions of BRCA1. These findings provide a valuable tool for the development of new therapeutic modalities of breast cancer linked to BRCA1.