Characterization:novel genotoxic stress-regulated gene

表征：新型遗传毒性应激调节基因

• 批准号: 7030099
• 负责人: M. SAEED SHEIKH
• 金额: $ 22.8万
• 依托单位: UPSTATE MEDICAL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-01 至 2008-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7030099
• 关键词: DNA damage; RNA interference; SDS polyacrylamide gel electrophoresis; apoptosis; biological signal transduction; calcineurin; calmodulin; cell adhesion molecules; cell growth regulation; colon neoplasms; environment related neoplasm /cancer; gene environment interaction; gene induction /repression; genetic markers; genetic promoter element; genetic transcription; green fluorescent proteins; human tissue; messenger RNA; molecular cloning; neoplasm /cancer genetics; neoplastic process; p53 gene /protein; protein binding; protein protein interaction; protein structure function; small interfering RNA; western blottings

DESCRIPTION (provided by applicant): Cells are constantly exposed to a variety of environmental agents some of which induce DNA damage i.e. genotoxic stress. Cellular responses to genotoxic stress are complex and therefore, more studies are needed to better understand the mechanisms that control such responses. Here we propose to characterize a novel gene, which we have named PIQ (p53-regulated IQ protein) that codes for an IQ motif protein. IQ motifs promote protein interactions with calmodulin (CaM). CaM is a calcium-binding protein that controls several important signaling events. Evidence presented here show that PIQ does indeed bind to CaM. Furthermore, PIQ mRNA is overexpressed in primary colon tumors when compared with matching normal tissues and that PIQ is down-regulated in response to genotoxic stress and p53 activation. PUMA is a key mediator of p53- dependent genotoxic stress-induced apoptosis and our results indicate that PIQ down-regulates PUMA promoter expression. Importantly, PUMA promoter harbors regulatory elements that are predicted to be regulated by CaM-dependent signaling pathways. We, therefore, propose that in unstressed cells, PIQ may negatively regulate PUMA expression by interfering with CaM-dependent signaling events and PIQ downregulation following genotoxic stress serves to promote PUMA upregulation partly via CaM-dependent pathways. Our overall hypothesis is that PIQ is a negative modulator of apoptosis and that higher levels of PIQ expression noted in colon cancer may alter the threshold of apoptosis and consequently contribute in part to colon tumorigenesis. Three specific aims are proposed here to further characterize PIQ in order to elucidate its role in cellular response to genotoxic stress in general and in the development and/or progression of colon cancer in particular. Specific Aim 1 is to investigate the role of PIQ during p53 and genotoxic stress-induced apoptosis. Specific Aim 2 is to investigate the mechanism by which PIQ negatively regulates PUMA promoter expression. Specific Aim 3 is to analyze PIQ expression at the mRNA and protein levels in a larger pool of primary colon tumors and their matching normal tissues and to evaluate a relationship between tumor PIQ-status and the clinicopathologic features.

描述（由申请人提供）：细胞不断暴露于多种环境药物，其中一些会诱导DNA损伤，即遗传毒性应激。细胞对遗传毒性应激的反应很复杂，因此需要更多的研究来更好地了解控制此类反应的机制。在这里，我们建议表征一个新型基因，我们将其命名为PIQ（P53调节的IQ蛋白），该基因编码为IQ基序蛋白。智商基序可促进与钙调蛋白（CAM）的蛋白质相互作用。 CAM是一种控制几个重要信号事件的钙结合蛋白。这里提供的证据表明，PIQ确实与CAM结合了。此外，与匹配正常组织相比，PIQ mRNA在原发性结肠肿瘤中过表达，并且该PIQ响应于遗传毒性应激和p53激活而下调。 PUMA是p53依赖性遗传毒性应激诱导的细胞凋亡的关键介质，我们的结果表明PIQ下调了PUMA启动子的表达。重要的是，PUMA启动子具有预测由CAM依赖性信号通路调节的调节元件。因此，我们提出，在无重力的细胞中，PIQ可能通过干扰CAM依赖性信号传导事件和遗传毒性应激后的PIQ下调来负调节PUMA表达，以部分通过CAM依赖性途径促进PUMA上调。我们的总体假设是，PIQ是凋亡的负调节剂，在结肠癌中指出的较高水平的PIQ表达可能会改变凋亡的阈值，因此部分促进了结肠肿瘤发生。这里提出了三个特定的目的，以进一步表征PIQ，以阐明其在一般和尤其是结肠癌的发展和/或进展中对遗传毒性应激的作用。具体目的1是研究PIQ在p53和遗传毒性应激诱导的凋亡过程中的作用。具体目的2是研究PIQ负调节PUMA启动子表达的机制。具体目的3是分析较大的原发性结肠肿瘤及其匹配的正常组织中的mRNA和蛋白质水平的PIQ表达，并评估肿瘤PIQ-STATUS与临床病理学特征之间的关系。