Differential expression and subcellular localization of murine BRCA1 and BRCA1-delta 11 isoforms in murine and human cell lines.
BRCA1 mutations are involved in breast and ovarian cancer predisposition in humans. The biological functions of the murine BRCA1 gene have been extensively studied but little is known about murine BRCA1 proteins. To better characterize these proteins, we have cloned the full-length murine BRCA1 cDNA and a splice variant deleted of exon 11, BRCA1-delta 11, by RT-PCR method. Three polyclonal antibodies raised against various parts of murine BRCA1 were used in our study: D16, M20 and 5MO, which were generated in our laboratory. This allowed us to analyze the expression and subcellular localization of both isoforms in murine and human cell lines by immunoblotting, immunoprecipitation, cell fractionation and immunofluorescence. Endogenous BRCA1 was detected in murine cell lines but not splice variant BRCA1-delta 11, whereas both ectopically expressed murine isoforms were detected in transfected human Bosc 23 cells. Subcellular fractionation and immunofluorescence results showed that the BRCA1 protein was mainly located in the nucleus, whereas BRCA1-delta 11 was preferentially cytoplasmic. The conservation of exon 11 splicing and the differential subcellular localization of BRCA1 and BRCA1-delta 11 in human and mouse suggest that these proteins could play distinct roles and that they could differentially act in the pathological mechanisms leading to the development of breast and ovarian cancer. The characterization of the murine BRCA1 proteins and antibodies will be useful to further study BRCA1 functions in murine models.
Bachelier
R
R
Laboratoire Génétique et Cancer, UMR 5641 CNRS, Faculté de Médecine, Lyon, France.
Dalla Venezia
N
N
Mazoyer
S
S
Frappart
L
L
Lenoir
G M
GM
Vincent
A
A
D016475:3T3 Cells; D017398:Alternative Splicing; D000818:Animals; D019313:BRCA1 Protein / Q000032:analysis / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D005091:Exons; D015723:Gene Library; D019398:Genes, BRCA1; D014644:Genetic Variation; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D020033:Protein Isoforms / Q000032:analysis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017384:Sequence Deletion; D013724:Teratoma
D019313:BRCA1 Protein; D017931:DNA Primers; D020033:Protein Isoforms