Regulation of Stat Function in Breast Cancer

乳腺癌中 Stat 功能的调节

• 批准号: 6678088
• 负责人: Charles V Clevenger
• 金额: $ 28.21万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-07-01 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6678088
• 关键词: MCF7 cell; athymic mouse; binding proteins; breast neoplasms; chromatin; gene deletion mutation; gene expression; hormone receptor; immunoprecipitation; microarray technology; molecular oncology; peptidylprolyl isomerase; phosphorylation; point mutation; prolactin; protein localization; protein protein interaction; protein structure function; protooncogene; small interfering RNA; transcription factor

DESCRIPTION (provided by applicant): The Stat family of transcription factors is necessary for the growth and differentiation of mammary tissues. In mammary tissues, a significant means of Stat activation is the binding of the neuroendocrine hormone prolactin (PRL) to its receptor, the PRLr. PRL/PRLr binding results in the activation of receptor-associated signaling networks, such as the Jak/Stat pathway and the endocytosis and nuclear retrotransport of a complex between PRL and the prolyl isomerase cyclophilin B (CypB). The intranuclear PRL/CypB complex regulates Stat5-mediated gene expression and DNA binding. Using multiple approaches including yeast two-hybrid and transcription factor array analysis, we have demonstrated that the PRL/CypB complex regulates the interaction of Stat5 with multiple intranuclear proteins including the peptide inhibitor of activated Stat, PIAS3, the small ubiquitin-related modifier, SUMO2, and the transcription factor and proto-oncogene, c-Myb. It is the central hypothesis of this proposal that the temporal and spatial interaction of Stat5 with these proteins, as modulated by the PRL/CypB complex, regulates Stat5 function. This hypothesis will be tested by three specific aims using breast cancer and PRL-responsive cell lines both in vitro and in vivo. First, the structural basis for Stat5/PIAS3 interaction and the functional significance of this interaction will be mapped using complementary mutagenesis/overexpression and knockdown strategies. Second, the functional consequences of Stat5 sumolyation will be evaluated by cellular and biochemical approaches. Third, the association between Stat5 and co-activators/repressors, such as c-Myb will be assessed by targeted mutagenesis, small-interfering RNA (SiRNA), and chromatin immunoprecipitation (ChlP)-based methodologies. Given that our preliminary evidence indicates that the manipulation of these interactions results in the alteration of Stat5-mediated gene expression and the survival of breast cancer cells, our proposed studies are directly relevant to a better understanding of the pathophysiology of: human breast cancer, and as such, may provide the basis for novel therapeutic strategies aimed at modifying Stat function in this disease.

描述（由申请人提供）：转录因子 Stat 家族对于乳腺组织的生长和分化是必需的。在乳腺组织中，Stat 激活的一个重要方式是神经内分泌激素催乳素 (PRL) 与其受体 PRLr 的结合。 PRL/PRLr 结合导致受体相关信号网络的激活，例如 Jak/Stat 通路以及 PRL 和脯氨酰异构酶亲环蛋白 B (CypB) 之间复合物的内吞作用和核逆转录。核内 PRL/CypB 复合物调节 Stat5 介导的基因表达和 DNA 结合。使用酵母双杂交和转录因子阵列分析等多种方法，我们证明 PRL/CypB 复合物调节 Stat5 与多种核内蛋白的相互作用，包括激活 Stat 的肽抑制剂 PIAS3、小泛素相关修饰剂 SUMO2 以及转录因子和原癌基因 c-Myb。该提议的中心假设是 Stat5 与这些蛋白质的时间和空间相互作用，在 PRL/CypB 复合物的调节下，调节 Stat5 的功能。该假设将通过三个具体目标使用乳腺癌和 PRL 响应细胞系在体外和体内进行测试。首先，将使用互补诱变/过表达和敲低策略来绘制 Stat5/PIAS3 相互作用的结构基础以及这种相互作用的功能意义。其次，Stat5 sumolyation 的功能后果将通过细胞和生化方法进行评估。第三，将通过靶向诱变、小干扰RNA (SiRNA) 和基于染色质免疫沉淀 (ChlP) 的方法来评估 Stat5 和共激活子/阻遏物（例如 c-Myb）之间的关联。鉴于我们的初步证据表明，操纵这些相互作用会导致 Stat5 介导的基因表达的改变和乳腺癌细胞的存活，因此我们提出的研究与更好地了解人类乳腺癌的病理生理学直接相关，因此，可能为旨在改变该疾病中 Stat 功能的新治疗策略提供基础。