PKC-EHN-PKD signal complex in cardiac remodeling

描述(申请人提供)：心脏重塑是在机械应激和各种神经体液因素的反应下触发的。我们最近发现，在心肌细胞中研究很少的蛋白激酶D(PKD)在Gq偶联受体(GqPCR)-蛋白激酶C(PKC)信号的下游对心肌肥大和重构起着重要的调节作用。此外，我们还发现了一种肌肉限制性的PDZ-3LIM蛋白ENH1，它具有选择性地直接与PKC和PKD结合的独特功能，并将该激酶复合体与心脏Z-Disc分离，后者被认为是机械应激信号的中枢机制。基于这些观察结果，我们推测PKCRENH1-PKD1复合体通过整合GqPCR信号和机械应激信号在心肌肥厚和重构的调节中发挥关键作用。在这项提议中，我们将使用转基因小鼠模型和体内生理分析的组合策略来检验这一假设。本研究侧重于体内分析机械应激诱导的心脏重构与PKC-ENH1-PKD1信号复合体之间的关系。因此，我们的具体目标是：1)在心肌肥厚和衰竭的动物模型中表征PKD在心脏重塑过程中的活体激活；2)建立和鉴定PKD的结构性活性突变和显性阴性突变体的心脏转基因小鼠；3)检测ENH缺失的心脏和心肌细胞，以确定ENH在心脏机械应激信号中的生理作用；以及4)检测PKC-ENH-PKD信号通路在心脏重塑中的潜在下游靶分子。我们预计，这些研究的结果将有助于我们更好地理解决定应激状态下心肌细胞命运的分子机制，并有助于设计新的药物来诱导病变心脏的有利重构。

DESCRIPTION (provided by applicant): Cardiac remodeling is triggered in response to mechanical stress and various neurohumoral factors. We have recently discovered that protein kinase D (PKD), which has been scarcely studied in cardiomyocytes, plays a significant role to regulate cardiac hypertrophy and remodeling in the downstream of Gq-coupling receptor (GqPCR) -protein kinase C (PKC) signaling. Furthermore, we have identified that a muscle restricted PDZ-3LIM protein, ENH1, has a unique function to selectively and directly bind to PKC and PKD and segregate this kinase complex to cardiac Z-disc, which has been thought as a central machinery of mechanical stress signaling. Based on these observations, we hypothesize that the PKCr ENH1-PKD1 complex plays a critical role in the regulation of cardiac hypertrophy and remodeling by integrating GqPCR signaling and mechanical stress signaling. In this proposal, we will test this hypothesis using a combinatory strategy of genetically modified mouse models and in vivo physiological analysis. The research focuses on in vivo analysis of the relationship between the mechanical stress-induced cardiac remodeling and the PKC-ENH1-PKD1 signal complex. Accordingly, our specific aims are: 1) to characterize in vivo activation of PKD in the process of cardiac remodeling in animal models of cardiac hypertrophy and failure, 2) to generate and characterize cardiac transgenic mice with constitutive active and dominant negative mutants of PKD, 3) to test ENH-null hearts and cardiomyocytes to define the physiological roles of ENH in cardiac mechanical stress signaling, and 4) to examine potential downstream target molecules of PKC-ENH-PKD signaling in cardiac remodeling. We anticipate that the outcome of these studies will assist our better understanding of molecular mechanisms that determine the fate of stressed cardiomyocytes and help to design new drug to induce favorable remodeling in diseased hearts.