Maniccia Anna W, Lewis Catherine, Begum Nurjahan, Xu Jingyao, Cui Jianqi, Chipitsyna Galina, Aysola Kartik, Reddy Vaishali, Bhat Ganapathy, Fujimura Yasuo, Henderson Beric, Reddy E Shyam P, Rao Veena N
Cancer Biology Program, Department of OB/GYN, Morehouse School of Medicine, Georgia Cancer Center for Excellence, Atlanta, Georgia, USA.
J Cell Physiol. 2009 Jun;219(3):634-41. doi: 10.1002/jcp.21708.
BRCA1 is a tumor suppressor gene that is mutated in families with breast and ovarian cancer. Several BRCA1 splice variants are found in different tissues, but their subcellular localization and functions are poorly understood at the moment. We previously described BRCA1 splice variant BRCA1a to induce apoptosis and function as a tumor suppressor of triple negative breast, ovarian and prostate cancers. In this study we have analyzed the function of BRCA1 isoforms (BRCA1a and BRCA1b) and compared them to the wild-type BRCA1 protein using several criteria like studying expression in normal and tumor cells by RNase protection assays, subcellular localization/fractionation by immunofluorescence microscopy and Western blot analysis, transcription regulation of biological relevant proteins and growth suppression in breast cancer cells. We are demonstrating for the first time that ectopically expressed GFP-tagged BRCA1, BRCA1a, and BRCA1b proteins are localized to the mitochondria, repress ELK-1 transcriptional activity and possess antiproliferative activity on breast cancer cells. These results suggest that the exon 9, 10, and 11 sequences (aa 263-1365) which contain two nuclear localization signals, p53, Rb, c-Myc, gamma-tubulin, Stat, Rad51, Rad50 binding domains, angiopoietin-1 repression domain are not absolutely required for mitochondrial localization and growth suppressor function of these proteins. Since mitochondrial dysfunction is a hallmark of cancer, we can speculate that the mitochondrial localization of BRCA1 proteins may be functionally significant in regulating both the mitochondrial DNA damage as well as apoptotic activity of BRCA1 proteins and mislocalization causes cancer. J. Cell. Physiol. 219: 634-641, 2009. (c) 2009 Wiley-Liss, Inc.
BRCA1是一种肿瘤抑制基因,在乳腺癌和卵巢癌家族中发生突变。在不同组织中发现了几种BRCA1剪接变体,但目前对它们的亚细胞定位和功能了解甚少。我们之前描述过BRCA1剪接变体BRCA1a可诱导细胞凋亡,并作为三阴性乳腺癌、卵巢癌和前列腺癌的肿瘤抑制因子发挥作用。在本研究中,我们分析了BRCA1同工型(BRCA1a和BRCA1b)的功能,并使用多种标准将它们与野生型BRCA1蛋白进行比较,如通过核糖核酸酶保护分析研究正常细胞和肿瘤细胞中的表达、通过免疫荧光显微镜和蛋白质印迹分析进行亚细胞定位/分级分离、生物相关蛋白的转录调控以及乳腺癌细胞中的生长抑制。我们首次证明,异位表达的绿色荧光蛋白标记的BRCA1、BRCA1a和BRCA1b蛋白定位于线粒体,抑制ELK-1转录活性,并对乳腺癌细胞具有抗增殖活性。这些结果表明,包含两个核定位信号、p53、Rb、c-Myc、γ-微管蛋白、Stat、Rad51、Rad50结合结构域、血管生成素-1抑制结构域的外显子9、10和11序列(氨基酸263 - 1365)对于这些蛋白的线粒体定位和生长抑制功能并非绝对必需。由于线粒体功能障碍是癌症的一个标志,我们可以推测BRCA1蛋白的线粒体定位在调节线粒体DNA损伤以及BRCA1蛋白的凋亡活性方面可能具有功能意义,而定位错误会导致癌症。《细胞生理学杂志》219: 634 - 641, 2009。(c) 2009威利 - 利斯出版公司。