CDF-1 Regulation of Zinc Homeostasis

CDF-1 锌稳态的调节

• 批准号: 6898862
• 负责人: Kerry Kornfeld
• 金额: $ 29.61万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-06-01 至 2007-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6898862
• 关键词: Caenorhabditis elegans; atomic absorption spectrometry; biological transport; dietary trace element; fluorescence microscopy; genetic regulation; genetic screening; homeostasis; mass spectrometry; messenger RNA; metal metabolism; molecular cloning; protein binding; protein kinase; transcription factor; yeast two hybrid system; zinc

DESCRIPTION (provided by applicant): The long-term objective of this proposal is to characterize the mechanisms of zinc metabolism and the regulation of Zn2+-homeostasis in multicellular animals. Zn2+ metabolism has important implications for human health because zinc deficiencies caused by inadequate diet or inborn errors of metabolism result in many pathologies. Zn2+ also regulates important processes such as cell proliferation, and Zn2+ metabolism may affect diseases such as cancer. The first specific aim is to characterize the role of the C. elegans cdf-1 gene in Zn2+ metabolism in an intact animal. cdf-1 encodes a cation diffusion facilitator protein that promotes Zn2+ efflux across the plasma membrane. The research design and methods include developing assays of zinc content, distribution, uptake and excretion in nematode worms. These assays will be used to determine how changes in dietary zinc affect Zn2+-homeostasis. The role of cdf-1 in Zn2+ metabolism will be determined by analyzing mutants that lack CDF-1 activity, overexpress CDF-1, or express CDF-1 in specific tissues. The regulation of CDF-1 activity will be characterized by analyzing cdf-1 mRNA and protein in intact animals and determining how these products are regulated by dietary zinC. The final part of the first specific aim is to characterize the biochemical mechanism of action of CDF-1 in promoting Zn2+ transport across the plasma membrane. The research design and methods include developing assays of Zn2+ transport using purified components or cellular systems and characterizing the role of CDF-1 in Zn 2+ transport. Proteins that bind to CDF-1 will be identified using the yeast two-hybrid system and the role of CDF-l-interacting proteins in Zn2+ transport will be analyzed. The role of CDF-1-interacting proteins in intact animals will be investigated using genetic approaches. The second specific aim is to identify a network of genes that regulate Zn2+- homeostasis. The research design and methods include conducting genetic screens for mutations that affect C. elegans Zn2+ metabolism. Genetic methods will be used to determine the specific role of newly identified genes in Zn2+ metabolism. The role of these genes in cell fate specification will be determined to characterize the relationship between Zn2+ metabolism and Ras-mediated signaling. Molecular approaches will be used to clone the affected genes and reveal the mechanisms used by these proteins to regulate Zn2+ metabolism. These studies are likely to provide significant new insights into Zn2+ metabolism by establishing the role of CDF-1 in an intact animal, elucidating the biochemical mechanism of CDF-1, and identifying and characterizing new proteins that regulate Zn2+.

描述（由申请人提供）：该提案的长期目标是表征多细胞动物中锌代谢的机制和Zn2+-Homeostasis的调节。 Zn2+代谢对人类健康具有重要意义，因为饮食不足或代谢的天生错误引起的锌不足会导致许多病理。 Zn2+还调节重要过程，例如细胞增殖，Zn2+代谢可能会影响诸如癌症之类的疾病。 第一个具体目的是表征秀丽隐杆线虫CDF-1基因在完整动物中代谢中的作用。 CDF-1编码阳离子扩散促进蛋白促进Zn2+外排跨质膜。 研究设计和方法包括在线虫蠕虫中开发锌含量，分布，摄取和排泄的测定法。 这些分析将用于确定饮食锌的变化如何影响Zn2+-Homeostasis。 CDF-1在Zn2+代谢中的作用将通过分析缺乏CDF-1活性，过表达CDF-1或表达CDF-1的突变体在特定组织中确定。 CDF-1活性的调节将以分析完整动物中的CDF-1 mRNA和蛋白质的特征，并确定如何通过饮食锌调节这些产物。 第一个特定目的的最后部分是表征CDF-1在促进Zn2+转运跨质膜中的生化机理。 研究设计和方法包括使用纯化的组件或细胞系统开发Zn2+传输的测定，并表征CDF-1在Zn 2+传输中的作用。 将使用酵母两杂交系统鉴定与CDF-1结合的蛋白质，并将分析CDF-L交互蛋白在Zn2+转运中的作用。 CDF-1相互作用蛋白在完整动物中的作用将使用遗传方法研究。 第二个具体目的是确定调节Zn2+稳态的基因网络。 研究设计和方法包括用于影响秀丽隐杆线虫Zn2+代谢的突变的遗传筛选。 遗传方法将用于确定新鉴定的基因在Zn2+代谢中的特定作用。 这些基因在细胞命运规范中的作用将被确定以表征Zn2+代谢与RAS介导的信号传导之间的关系。 分子方法将用于克隆受影响的基因，并揭示这些蛋白质用于调节Zn2+代谢的机制。 这些研究可能通过确定CDF-1在完整动物中的作用，阐明CDF-1的生化机制，并识别和表征调节Zn2+的新蛋白质，从而为Zn2+代谢提供重要的新见解。