Mechanisms for glucocorticoid vs. progesterone receptor-specific gene regulation

糖皮质激素与孕酮受体特异性基因调控的机制

• 批准号: 8349788
• 负责人: S S SIMONS
• 金额: $ 37.75万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8349788
• 关键词: Accounting; Agonist; Binding; Biochemical; Biochemistry; Biological; Biological Assay; Biological Process; C-terminal; Cells; Complex; Development; Enzyme Kinetics; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic Epistasis; Genetic Transcription; Glucocorticoid Receptor; Glucocorticoids; Goals; Human; Joints; Modeling; Molecular; N-terminal; Osteogenesis; Physiological; Physiology; Progesterone Receptors; Progestins; Property; Reaction; Relative (related person); Reporter Genes; Repression; Residual state; Role; Site; Steroid Receptors; Steroids; Structure; Theoretical model; Transactivation; analog; base; bone; cofactor; gene induction; hormone response element; in vivo; inhibitor/antagonist; novel; receptor; research study; response

The corepressors NCoR and SMRT have been documented to have opposite effects on the EC50 of agonists, and the PAA, for GR and PR induction of the same gene in the same cells. These inverted responses depend upon the joint actions of the N- and C-terminal domains of each receptor (Song et al., 2001, J. Biol. Chem., 276, 24806-24816). These results are consistent with the demonstration that corepressors interact with N-terminal regions of both GRs and PRs (Wang et al., 2007, Biochemistry, 48, 8036-8049; Wang and Simons Jr., 2005, Mol. Endo., 19, 1483-1500) in addition to the initially defined sites in the C-terminal sequences of receptors. Similarly, four other factors known to modulate GR activity (GME, GMEB2, Ubc9, and STAMP) were recently found either to differentially alter several induction parameters of GRs vs. PRs under otherwise identical conditions or to require different regions of each receptor for their activities (Szapary et al., 2008, Mol Cell Endocrinol, 283, 114-126). The objective of this study is to determine whether the mechanisms of action change for those factors known to differentially modulate the EC50, PAA, and Amax of GR- vs. PR-regulated gene transcription. This task has just become tractable with our development of a theoretical model of steroid receptor action (see DK DK057800-20). Three novel features of this model, and its associated graphical analysis, permit an unprecedented level of mechanistic information regarding steroid receptor-regulated gene transactivation. First, it is now possible to determine the kinetically-defined type of action being displayed by the factor (competitive inhibitor, uncompetitive inhibitor, coactivator, etc.). Second, it is usually possible to define where the factor acts relative to a reference point called the concentration limiting step (CLS), which is the steady state analog of the rate limiting step of enzyme kinetics. Third, the model and its graphical analysis have recently been extended to the analysis of two competing factors in the same assay. As opposed to making the situation more obscure, it actually yields greater mechanistic information. Not only can such competition assays determine how and where each factor acts, relative to the CLS, but the site of action of the two factors relative to each other is revealed. Thus, one can now assemble an ordered sequence of reactions based on the biological function of cofactors, much as in epistasis analysis, even when the biochemical properties of the cofactors are not known. Initial experiments with a few cofactors have not exposed any major differences between GR and PR action. However, we expect that continued studies, especially with endogenous genes, will identifiy some mechanistic differences between GR and PR interactions with cofactors. This type of information is not now available by any other means. Therefore, these studies will greatly contribute to our long-term goal of defining the action of GRs vs. PRs at a molecular level and of understanding their role in human physiology.

已证明辅阻遏物NCoR和SMRT对激动剂的EC 50和PAA具有相反的作用，用于相同细胞中相同基因的GR和PR诱导。 这些反向响应取决于每种受体的N-和C-末端结构域的联合作用（Song等人，2001，J. Biol. Chem.，276，24806-24816）。 这些结果与辅阻遏物与GR和PR的N-末端区域相互作用的证明一致（Wang等人，2007，Biochemistry，48，8036-8049; Wang和Simons Jr.，2005，Mol.远藤，19，1483-1500）。 类似地，最近发现已知调节GR活性的四种其他因子（GME、GMEB 2、Ubc 9和STAMP）在其他相同条件下差异性地改变GR相对于PR的几个诱导参数，或者需要每种受体的不同区域用于其活性（Szapary等人，2008，Mol Cell Endocrinol，283，114-126）。 本研究的目的是确定已知差异调节GR与PR调节基因转录的EC 50、PAA和Amax的那些因子的作用机制是否发生变化。 随着我们开发类固醇受体作用的理论模型（参见DK DK 057800 -20），这项任务刚刚变得容易处理。 该模型的三个新特征及其相关的图形分析，允许类固醇受体调节的基因反式激活的机制信息的前所未有的水平。 首先，现在可以确定因子所显示的动力学定义的作用类型（竞争性抑制剂、非竞争性抑制剂、共激活剂等）。 其次，通常可以定义因子相对于称为浓度限制步骤（CLS）的参考点的作用位置，浓度限制步骤是酶动力学速率限制步骤的稳态模拟。 第三，该模型及其图形分析最近已扩展到同一测定中两个竞争因素的分析。 与使情况更加模糊相反，它实际上产生了更多的机械信息。 这样的竞争测定不仅可以确定每个因子相对于CLS如何和在何处起作用，而且可以揭示两个因子相对于彼此的作用位点。 因此，人们现在可以根据辅因子的生物学功能组装有序的反应序列，就像上位性分析一样，即使辅因子的生物化学性质是未知的。 用几个辅因子进行的初步实验没有发现GR和PR作用之间的任何重大差异。 然而，我们期望继续的研究，特别是内源性基因的研究，将确定GR和PR与辅因子相互作用之间的一些机制差异。 这类信息目前无法通过任何其他途径获得。 因此，这些研究将大大有助于我们在分子水平上定义GR与PR的作用以及了解它们在人体生理学中的作用的长期目标。