Abstract 2455: BRCA1 and BARD1 expression and localization in breast cell lines

Abstract

Abstract One of the principal functions of BRCA1 is to keep the integrity of DNA by homologous recombination which repairs DNA double strand breaks, thus its localization is mainly nuclear in a normal cell. BRCA1 nuclear function is enhanced by its interaction with BARD1, which promotes the import and nuclear retention of BRCA1 by masking its nuclear export signal. Therefore, the interaction of these two proteins is relevant in BRCA1 nuclear function. On the other hand, in tumor cell lines, splicing variants of BARD1 lacking the BRCA1 interaction domain have been described. The expression of these splicing variants of BARD1 could contribute to the loss of nuclear localization of BRCA1. Recently, our group found that a high percentage of triple negative breast cancer tumors showed cytoplasmic or no expression of BRCA1 (unpublished data). Our interest is to study the molecular mechanisms that explain the cytoplasmic localization of BRCA1 in breast cancer cells. We studied mRNA expression of BRCA1, BARD1 and BARD1β through RT-PCR and expression of both proteins by western blot, immunocytochemistry and immunofluorescence in two breast cancer cell lines (HCC1937: ER-, mutated in BRCA1, and T47D: ER+) and one non-tumor breast cell line (MCF10A). BRCA1 and BARD1 expression was coherent between immunocytochemistry and immunofluorescence assay. The HCC1937 cell line presented a weak BRCA1 protein expression mainly in the cytoplasm. The other cell lines had a high nuclear and weak cytoplasmic expression. BARD1 protein is expressed in the nucleus and cytoplasm in all cell lines. BARD1 full mRNA had an equal expression in the three cell lines, and its isoform BARD1β, which do not interact with BRCA1, was expressed at higher levels in tumor cell lines than in the non-tumor cell lines. However, T47D cell line showed nuclear localization of BRCA1 despite high levels of BARD1β mRNA. This suggests that BARD1β either does not express at protein level, or that full-length BARD1 levels are enough to keep BRCA1 in the nucleus. The study of molecular causes of BRCA1 localization in cell lines will allow us to better understand subcellular localization of BRCA1 in tumors and its interaction with BARD1. FONDECYT 1120200. Citation Format: Cristobal Herrera, Elisa Pérez, Elías Blanco, Patricia Gajardo, Pilar Carvallo. BRCA1 and BARD1 expression and localization in breast cell lines. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2455. doi:10.1158/1538-7445.AM2014-2455