Localization of BRCA1 protein in breast cancer tissue and cell lines with mutations

Abstract

BackgroundThe breast and ovarian cancer susceptibility gene (BRCA1) encodes a tumor suppressor. The BRCA1 protein is found primarily in cell nuclei and plays an important role in the DNA damage response and transcriptional regulation. Deficiencies in DNA repair capabilities have been associated with higher histopathological grade and worse prognosis in breast cancer.MethodsIn order to investigate the subcellular distribution of BRCA1 in tumor tissue we randomly selected 22 breast carcinomas and tested BRCA1 protein localization in frozen and contiguous formalin-fixed, paraffin embedded (FFPE) tissue, using pressure cooker antigen-retrieval and the MS110 antibody staining. To assess the impact of BRCA1 germline mutations on protein localization, we retrospectively tested 16 of the tumor specimens to determine whether they contained the common Ashkenazi Jewish founder mutations in BRCA1 (185delAG, 5382insC), and BRCA2 (6174delT). We also compared co-localization of BRCA1 and nucleolin in MCF7 cells (wild type) and a mutant BRCA1 cell line, HCC1937 (5382insC).ResultsIn FFPE tissue, with MS110 antibody staining, we frequently found reduced BRCA1 nuclear staining in breast tumor tissue compared to normal tissue, and less BRCA1 staining with higher histological grade in the tumors. However, in the frozen sections, BRCA1 antibody staining showed punctate, intra-nuclear granules in varying numbers of tumor, lactating, and normal cells. Two mutation carriers were identified and were confirmed by gene sequencing. We have also compared co-localization of BRCA1 and nucleolin in MCF7 cells (wild type) and a mutant BRCA1 cell line, HCC1937 (5382insC) and found altered sub-nuclear and nucleolar localization patterns consistent with a functional impact of the mutation on protein localization.ConclusionsThe data presented here support a role for BRCA1 in the pathogenesis of sporadic and inherited breast cancers. The use of well-characterized reagents may lead to further insights into the function of BRCA1 and possibly the further development of targeted therapeutics.

BRCA1 protein expression and subcellular localization in primary breast cancer: Automated digital microscopy analysis of tissue microarrays

“…However, BRCA1 mutation screening is helpful only for individuals who receive a clear-cut result (pathogenic mutation with a known function or negative for any mutation) and not for others who have BRCA1 mutations with uncertain significance. Our data indicated that BRCA1 expression and subcellular localization might stand as an independent prognostic factor in breast cancer which is supported by similar findings from previous studies [ 17 , 23 , 38 ]. However, this research would benefit greatly from larger population-based studies that might encourage health care workers to incorporate BRCA1 staining in breast cancer prognosis algorithm.…”

Selective mode of action of plumbagin through BRCA1 deficient breast cancer stem cells

“…We confirmed the low, cytosolic expression of BRCA1 in HCC1937 while in HCC1937/wt BRCA1, the BRCA1 protein was expressed and localized specifically to the nucleus (Additional file 3 : Figure S2). This was expected as the truncated, non-functional BRCA1 expressed by HCC1937 has been observed in nucleus as well as the cytoplasm [ 32 ]. Cell proliferation assay revealed that PB is highly selective towards the HCC1937 cells with 5 μM concentration inhibiting proliferation in 56 % cells while in HCC1937/wt BRCA1 only 25 % inhibition of proliferation was observed (Fig.…”

Establishment and characterization of a receptor-negative, hormone-nonresponsive breast cancer cell line from an Iraqi patient

“… 25 AMJ13 cells were positive for BRCA1 nuclear staining, but to a lesser degree than for BRCA2, which was positive for both nuclear and cytoplasmic staining. Tulchin et al 26 frequently found reduced BRCA1 nuclear staining in breast tumor tissue when compared with normal tissue, and less BRCA1 staining with higher histological tumor grade. Strong positive staining of the nucleus and cytoplasm suggests mutation in BRCA1 because of its role in cell cycle control and DNA repair.…”