RETINOIDS--RECEPTOR ACTIVATION AND BINDING KINETICS

类维生素A——受体激活和结合动力学

• 批准号: 6377046
• 负责人: BRAHMA P SANI
• 金额: $ 35.19万
• 依托单位: SOUTHERN RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-06-01 至 2004-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6377046
• 关键词: DNA binding protein; chemical binding; chemical kinetics; genetic regulatory element; high performance liquid chromatography; intermolecular interaction; ligands; nuclear receptors; protein isoforms; receptor binding; retinoid binding proteins; retinoids; stoichiometry; tissue /cell culture; transcription factor

The effects of natural retinoic acids [RA, 9-cis-RA, 3,4-didehydroRA (DDRA) and 4-oxo-RA] in the regulation of discrete sets of target gens are produced through a cascade of events mediated by the retinoid family of nuclear receptors (RARs and RXRs). Ligand binding, receptor dimerization and transcriptional complex formation are three main steps in the display of the biological activity of these retinoids. Following our development of an analytical procedure to study the homo- and hetero- dimerization of RARs/RXRs and complexing with RA response elements (RARE), we propose to unravel the intricate mechanisms of receptor- mediated molecular events by determining in a quantitative manner the receptor subtype selectivity and ligand specificity in ligand/receptor, receptor protein-protein, and protein-DNA interactions. The selectivity in binding produced by single ligands and combinations of ligands for dimers of multiple receptors isoforms will be elucidated and further verified by transcriptional activation assays in CV-1 cells. We will examine whether the optimal physical conditions established in the above interactions are physiological relevant with respect to the kinetics of binding of Ras to RARs/RXRs under monomer, dimer, and dimer-RE forming conditions. Since DDRA (vitamin A2 acid) and 4-oxo-RA (the major cellular metabolite of RA) display selective biological activities and bind RARs/RXRs, we will elucidate, through separate set of experiments, their specific participating roles in the central scheme of RA/9-cis-RA-RAR/RXR-RE mediated transcriptional complex formation and gene activation. These studies will help unravel the key steps in the receptor-mediated molecular mechanism of action of retinoids in reference to the discrete genomic functions of natural Ras in the control of cell differentiation and tumorigenesis.

天然视黄酸[RA，9-CIS-RA，3,4-二羟基氢（DDRA）和4-氧RA]在调节离散靶性质集合中的影响是通过由维氏核受体家族（RARS和RXRS）介导的一系列事件产生的。配体结合，受体二聚化和转录复合物的形成是这些类维生素类似的生物学活性的三个主要步骤。遵循我们开发了一种分析程序，以研究RARS/RXR的同质和异构化以及与RA响应元件的络合（罕见），我们建议通过以定量的方式确定受体亚型选择性和Ligand特异性的LIGAND特异性和受体蛋白酶，蛋白酶和蛋白酶，通过定量方式来阐明受体介导的分子事件的复杂机制。单个配体产生的结合和配体组合对多种受体同工型的二聚体的组合将得到阐明，并通过CV-1细胞中的转录激活测定法进一步验证。我们将研究上述相互作用中建立的最佳物理条件是否与RAS在单体，二聚体和二聚体形成条件下ras/rxR的结合的动力学相关。由于DDRA（维生素A2酸）和4-oxo-RA（RA的主要细胞代谢产物）表现出选择性的生物学活动并结合RARS/RXR，我们将通过单独的实验集阐明它们在RA/9-CIS-RA/RXR/RXR介导的介导的转录复合物和依次的特定参与方案中。这些研究将有助于阐明类视视感素在控制细胞分化和肿瘤发生中天然RA的离散基因组功能方面受体介导的分子作用分子机理的关键步骤。