Coordinate Regulation Of Nuclear DGK-theta and PLD

描述（由申请人提供）：本提案侧重于一个新兴的信号转导范式，该范式反驳了主流观点，即核是质膜产生的信号的被动接受者。我们的实验室和其他人已经证明，细胞表面的受体激活导致核酶的调节，核酶产生和修饰有效的脂质信号分子。虽然这一领域引起了越来越多的兴趣，但我们仍然不了解调控的分子方面，也不了解诱导脂质代谢的确切生理作用。提出的研究旨在阐明这一信号转导机制的关键要素及其功能后果。我们发现小分子量gtp结合蛋白RhoA在响应成纤维细胞的α -凝血酶刺激时，在细胞核中相互调节两种信号酶，二酰基甘油激酶-和磷脂酶D。我们将这一规定称为“RhoA开关”。我们预测，由此产生的二酰基甘油和磷脂酸水平和分子组成的调节具有重要的生理后果，如核定位和蛋白激酶c - α活性的调节。两种异三聚体gtp结合蛋白Gq和G13以及脂质激酶PI 3-激酶介导凝血酶激活这一途径。我们假设；α -凝血酶在启动核二酰基甘油激酶-theta活性的早期Gq依赖性增加后，刺激G13/PI 3-激酶依赖性的RhoA核易位，该易位作为一个时间“开关”，在激活核磷脂酶d的同时抑制核二酰基甘油激酶-theta。我们将检验Gq对核二酰基甘油激酶-theta活性的诱导增加所必需的假设。Aim II将通过测试PI 3-激酶激活是否负责核RhoA易位来确定RhoA开关的分子机制。此外，我们将确定RhoA、二酰基甘油激酶和磷脂酶d之间的分子相互作用。RhoA开关的生理后果将在Aim III中进行研究。我们将重点关注开关对核蛋白激酶c - α活性、核磷脂酸组成和生长的影响。我们相信对核脂质信号的理解是我们干预诸如伤口修复和组织重塑等事件的关键。

DESCRIPTION (provided by applicant): This proposal focuses on an emerging signal transduction paradigm which counters the prevailing view that the nucleus is a passive recipient of signals produced at the plasma membrane. Our laboratory and others have shown that receptor activation at the cell surface results in regulation of nuclear enzymes that produce and modify potent lipid signaling molecules. While this area has garnered increasing interest, we still understand neither the molecular aspects of the regulation nor the precise physiological roles of the induced lipid metabolism. The proposed studies aim at elucidating key elements of this signal transduction mechanism and its functional consequences. We find that the small molecular weight GTP-binding protein RhoA reciprocally regulates two signaling enzymes, diacylglycerol kinase-theta and phospholipase D, at the nucleus in response to alpha-thrombin stimulation of fibroblasts. We have termed this regulation the "RhoA Switch". We predict that the resulting modulation of diacylglycerol and phosphatidic acid levels and molecular composition have important physiological consequences such as the regulation of nuclear localization and activity of protein kinase C-alpha. Two heterotrimeric GTP-binding proteins, Gq and G13, and the lipid kinase PI 3-kinase mediate thrombin activation of this pathway. We hypothesize that; alpha-thrombin, after initiating an early Gq-dependent increase in nuclear diacylglycerol kinase-theta activity, stimulates a G13/PI 3-kinase-dependent nuclear translocation of RhoA that acts as a temporal "switch" to inhibit nuclear diacylglycerol kinase-theta while activating nuclear phospholipase D. We will test the hypothesis that Gq is essential for the induced increase in nuclear diacylglycerol kinase-theta activity, and identify the diacylglycerol-theta domains essential for its nuclear translocation in Aim I. Aim II will determine the molecular mechanism of the RhoA Switch by testing whether PI 3-kinase activation is responsible for nuclear RhoA translocation. Further, we will determine the molecular interactions between RhoA, diacylglycerol kinase-theta and phospholipase D. The physiological consequences of the RhoA Switch will be examined in Aim III. We will focus on the effect of the Switch on nuclear protein kinase C-alpha activity, composition of nuclear phosphatidic acids, and growth. We believe that an understanding of nuclear lipid signaling is key to our ability to intervene in events such as wound repair and tissue remodeling.