ROLE OF THE BRCA1 GENE IN SPORADIC CANCER

遗传性乳腺癌和卵巢癌易感性的鉴定 BRCA1基因提高了人们对这种疾病的认识， 增加了对有家族病史的妇女的监测，以及 基因突变检测。然而，我们对这个角色的了解 该基因在散发性肿瘤中的表达及其在正常和肿瘤中的功能 增长和发展是稀少的。这项提议是对 过去两年进行的研究旨在调查这些 问题。本提案概述了以下实验：1)由于 BRCA1很少成为体细胞突变乳腺癌的靶点，是吗？ 因为失去表达而失去活力？使用显微解剖的正常上皮， 原位癌和浸润性癌，既有表达又有启动子 甲基化将使用基于聚合酶链式反应的方法进行评估。2)BRCA1 在整个细胞周期中，mRNA的表达受到严格的调控。改建 在表达上，无论是时间性质的层面，都可能发现它的 功能。因此，基础的和细胞周期调节的启动子元件 将会被确认。利用最小启动子片段，鉴定了 与特定启动子序列复杂的反式作用因子将是 已经成立了。这些实验将补充和扩展那些拟议的实验 在第一个目标中。3)很难获得过多的细胞系- 表达BRCA1以研究相关的表型和分子事件 带着蛋白质。为了克服这个问题，四环素 已建立了可诱导的细胞系。BRCA1在测试下-可诱导 启动子将被引入这些细胞中以研究表型效应 BRCA1过度表达包括细胞周期参数、细胞凋亡、 对DNA损伤的反应，以及对分化试剂的反应。4)使用 这些同样的细胞系，由BRCA1表达诱导的基因将是 使用代表性差异分析(RDA)进行识别。从 这些基因的启动子元件，常见的BRCA1反应元件将是 杰出的。

Identification of the hereditary breast and ovarian cancer susceptibility gene, BRCA1, has led to increased awareness regarding the disease, increased surveillance in women with a family history, and the advent of genetic testing for mutations. However, our knowledge regarding the role of this gene in sporadic cancers and its function in normal and neoplastic growth and development is sparse. This proposal is a continuation of studies performed over the past two years designed to investigate these issues. The following experiments are outlined in this proposal: 1) Since BRCA1 is rarely a target for somatic mutation breast cancer, is it inactivated by loss of expression? Using microdissected normal epithelium, carcinoma in situ, and invasive cancers, both expression and promoter methylation will be evaluated employing PCR based methodology. 2) BRCA1 mRNA expression is tightly regulated through the cell cycle. Alterations in expression, either the levels of temporal nature, may find its function. Therefore, the basal and cell cycle regulated promoter elements will be identified. Using the minimal promoter fragment, the identity of trans-acting factors that complex with specific promoter sequences will be established. These experiments will complement and extend those proposed in the first aim. 3) It has been difficult to derive cell lines that over- express BRCA1 in order to study phenotypic and molecular events associated with the protein. In order to overcome this problem, tetracycline inducible cell lines have been established. BRCA1 under the test-inducible promoter will be introduced into these cells to study phenotypic effects of BRCA1 over-expression including cell cycle parameters, apoptosis, response to DNA damage, and response to differentiating agents. 4) Using these same cell lines, genes that are induced by BRCA1 expression will be identified using representational difference analysis (RDA). From the promoter elements of these genes, common BRCA1 response elements will be distinguished.