Studies on the Ah receptor signaling mechanism

Ah受体信号传导机制研究

• 批准号: 6504601
• 负责人: WILLIAM K CHAN
• 金额: $ 11.82万
• 依托单位: UNIVERSITY OF THE PACIFIC-STOCKTON
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-06 至 2004-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6504601
• 关键词: Baculoviridae; aromatic hydrocarbon receptor; biological signal transduction; dioxins; environmental contamination; gene expression; ligands; protein protein interaction; protein purification; protein structure function; receptor binding; transcription factor; western blottings

DESCRIPTION (provided by applicant): Dioxins, generated both commercially and naturally, are chlorinated polycyclic aromatic hydrocarbons that are highly toxic environmental contaminants. These agents are known to be potent rodent carcinogens and suspected human carcinogens. The best-known prototype of this group of agents is 3,4,7,8-tetrachlorodibenzo-p-dioxin (TCDD). It has been well documented that most, if not all, of the TCDD effects are mediated through the Ah receptor (AhR). Thus, in an effort to better understand the mechanism of dioxins action, I propose to investigate the molecular mechanism of the AhR signaling pathway using the overexpressed AhR and Arnt proteins. My working hypothesis is as follows: Upon ligand binding, the AhR undergoes conformational changes resulting in alteration of a number of AhR-protein interactions, which subsequently leads to a cascade of biologic events to occur. In addition to AhR-protein interactions, other interactions involving Arnt and other proteins also contribute to the action of dioxins mediated by the AhR. This proposal contains specific aims investigating the mechanism for protein factors involvement in the AhR signaling. Not only do these studies provide important information and reagents to further the study of the PAH action mediated by the AhR, but they also provide mechanistic insights on how the AhR regulates its target genes by understanding what other proteins are involved in the AhR signaling pathway. Five specific aims have been proposed: Aim 1) Identify the protein factors in rabbit reticulocyte lysate (RRL) and the Sf9 soluble extract which are responsible for the AhR/Arnt/DRE complex formation; Aim 2) Identity interacting proteins of the AhR and Arnt and examine their effect on the AhR/Arnt/DRE complex formation; Aim 3) Determine whether different forms of the AhR/Arnt heterodimer are present in cell lines which contribute to the different levels of ligand dependent transactivation; Aim 4) Identify ARA9-interacting proteins which are responsible for the ARA9-mediated AhR/Arnt/DRE complex formation and Aim 5) Determine how CyP4O enhances the AhR/Arnt/DRE complex formation.

说明（由申请方提供）：商业和天然产生的二恶英是氯化多环芳烃，是剧毒环境污染物。已知这些物质是啮齿类动物的强效致癌物和疑似人类致癌物。这组药剂中最著名的原型是3，4，7，8-四氯二苯并对二恶英（TCDD）。它已被充分证明，大多数，如果不是全部，TCDD的影响是通过介导的Ah受体（AhR）。因此，在努力更好地了解二恶英的作用机制，我建议调查的AhR信号通路的分子机制，使用过表达的AhR和Arnt蛋白。我的工作假设如下：在配体结合后，AhR经历构象变化，导致许多AhR-蛋白质相互作用的改变，随后导致一连串生物事件的发生。除了AhR-蛋白质相互作用之外，涉及Arnt和其他蛋白质的其他相互作用也有助于AhR介导的二恶英的作用。该提案包含了研究蛋白质因子参与AhR信号传导的机制的具体目标。这些研究不仅为进一步研究AhR介导的PAH作用提供了重要的信息和试剂，而且还通过了解AhR信号通路中涉及的其他蛋白质，提供了关于AhR如何调节其靶基因的机制见解。目的1）鉴定兔网织红细胞裂解物（RRL）和Sf 9可溶性提取物中与AhR/Arnt/DRE复合物形成有关的蛋白因子：目的2）鉴定AhR和Arnt相互作用的蛋白，并研究它们对AhR/Arnt/DRE复合物形成的影响;目的3）确定细胞系中是否存在不同形式的AhR/Arnt异源二聚体，其有助于不同水平的配体依赖性反式激活;目的4）鉴定负责ARA 9介导的AhR/Arnt/DRE复合物形成的ARA 9相互作用蛋白，目的5）确定CyP 4 O如何增强AhR/Arnt/DRE复合物形成。