Regulation of Evi-1 function via the corepressor CtBP

通过辅阻遏物 CtBP 调节 Evi-1 功能

• 批准号: 7101781
• 负责人: QINGHONG ZHANG
• 金额: $ 16.11万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-08-01 至 2007-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7101781
• 关键词: DNA binding protein; carboxyl group; fibroblasts; fluorescence microscopy; gene induction /repression; genetic techniques; laboratory rat; liver cells; nicotinamide adenine dinucleotide; nucleoside inhibitor; oncoproteins; oxidation reduction reaction; protein protein interaction; protein sequence; protein structure function; radiotracer; tissue /cell culture; transcription factor

DESCRIPTION (provided by applicant): The transcriptional corepressor CtBP participates in pathways important for development, cell cycle regulation, and transformation. One such pathway involves the oncoprotein Evi-1, which has been linked to human acute myelogenous leukemia. Recently, the principal investigator demonstrated that CtBP is regulated by the nicotinamide adenine dinucleotides, NAD and NADH, in a manner that regulates its ability to interact with a variety of transcriptional repressors. This work suggested a completely novel mechanism for transcriptional regulation, in which the formation of a repressor:corepressor complex is regulated by changes in the nuclear levels of free NAD/NADH, with CtBP serving as a redox sensor for transcription. The main thrust of the current project is to extend this model by measuring the levels of free nicotinamide adenine dinucleotides in the nucleus, establishing the impact of this regulatory mechanism on the properties of the human transforming protein Evi-1, and elucidating the mechanism of CtBP-mediated transcriptional repression. Three specific aims are proposed: 1) Determine whether the free nuclear concentrations of nicotinamide adenine dinucleotides are appropriate for their proposed role in CtBP regulation. The affinities of NAD and NADH for CtBP will be measured in vitro and compared to their free nuclear concentrations, determined in vivo using two-photon fluorescence microscopy, 2) Establish whether the ability of CtBP to sense the nuclear redox state regulates the function of human oncoprotein Evi-1 and test whether blockers of the NAD/NADH binding site in CtBP inhibit Evi-1 function, 3) Identify components of the human cellular CtBP complex and determine the contributions of these components to transcriptional repression. Regulation of Evi-1 function through the proposed NAD/NADH-induced allosteric effect on CtBP is likely to affect the course of cell transformation. Blockers of the NAD/NADH-induced change may therefore be therapeutically useful in certain types of leukemia. Through a mentored career development plan, the principal investigator will gain proficiency in the field of gene regulation as it relates to molecular oncology. The mentor's laboratory and institution are rich in intellectual and physical resources that provide an excellent environment for facilitating the principal investigator's transition to independent scientist.

描述(由申请人提供)：转录辅阻遏子CtBP参与对发育、细胞周期调节和转化至关重要的途径。其中一个途径涉及癌蛋白Evi-1，它与人类急性髓系白血病有关。最近，首席研究人员证明了CtBP受烟酰胺腺嘌呤二核苷酸NAD和NADH的调控，这种方式调节了它与各种转录抑制物相互作用的能力。这项工作提出了一种全新的转录调控机制，其中抑制物：辅抑制子复合体的形成受游离NAD/NADH核水平的调节，CtBP作为转录的氧化还原传感器。本项目的主要目的是通过测量细胞核中游离烟酰胺腺嘌呤二核苷酸的水平来扩展这一模型，建立这一调控机制对人转化蛋白Evi-1性质的影响，并阐明CtBP介导的转录抑制的机制。提出了三个具体的目标：1)确定烟酰胺腺嘌呤二核苷酸的游离核浓度是否适合它们在CtBP调节中的作用。我们将在体外测量NAD和NADH与CtBP的亲和力，并将其与体内使用双光子荧光显微镜测定的游离核浓度进行比较；2)确定CtBP感知核氧化还原状态的能力是否调节人癌蛋白Evi-1的功能，并测试CtBP中NAD/NADH结合位点的阻断剂是否抑制Evi-1功能；3)鉴定人类细胞CtBP复合体的成分，并确定这些成分在转录抑制中的作用。通过NAD/NADH对CtBP的变构效应来调节Evi-1的功能可能会影响细胞转化的过程。因此，NAD/NADH诱导的改变的阻断剂可能对某些类型的白血病有治疗作用。通过有指导的职业发展计划，首席研究员将在与分子肿瘤学相关的基因调控领域获得熟练程度。导师的实验室和机构拥有丰富的智力和物质资源，为首席研究员向独立科学家的转变提供了良好的环境。