Structural Basis Of Dilated Cardiomyopathy

描述(申请人提供)：我的研究计划的总体目标是确定扩张型心肌病(DCM)的分子和结构决定因素，扩张型心肌病是全球心力衰竭(HF)的主要原因。DCM涉及三类主要的心脏蛋白：细胞骨架蛋白、核蛋白和肌节蛋白。我们小组专注于研究磷蛋白(PLN)中发生的突变，磷蛋白是一种参与肌浆网(SR)钙运输的膜蛋白。PLN结合并抑制SR Ca-ATPase(SERCA)，调节心脏的舒张性。PLN在Ser16位被蛋白激酶A(PKA)磷酸化，逆转了这种抑制效应，构成了心脏对b-肾上腺素能刺激的主要反应。磷酸化循环中的中断会进展到HF。R9C突变(PLNR9C)和R14缺失(PLNR14del)均位于PLN细胞质结构域，通过未知的机制阻止PLN的磷酸化并导致DCM。利用一系列生化、分子生物学和光谱方法，我们将表征这些突变在PKA/PLN复合体形成过程中的结构和动态影响，并将它们与其心脏毒性联系起来。具体地说，我们计划在存在和不存在这些致命突变的情况下，通过激酶破译PLN识别和磷酸化机制。此外，我们将通过肉豆蔻酰化来阐明脂膜和空间定位在磷酸化过程中的作用。这些研究将为理解心脏中磷酸化信号的生理学和病理生理学奠定基础。 公共卫生相关性：扩张型心肌病(DCM)是全球心力衰竭(HF)的主要原因。这项建议试图用生物物理、生化和分子生物学的方法来阐明DCM的分子决定因素。在原子水平上了解这种疾病的起源将有助于将这一知识转化为对抗心力衰竭的新治疗方法的分子设计。

DESCRIPTION (provided by applicant): The overall goal of my research program is to determine the molecular and structural determinants of dilated cardiomyopathy (DCM), the leading cause for heart failure (HF) worldwide. Three major groups of cardiac proteins are involved in DCM: cytoskeletal, nuclear, and sarcomeric. Our group focuses on the study of mutations occurring in phospholamban (PLN), a membrane protein involved in calcium transport in the sarcoplasmic reticulum (SR). PLN binds and inhibits the SR Ca-ATPase (SERCA), regulating heart diastole. PLN phosphorylation at Ser16 by protein kinase A (PKA) reverses the inhibitory effects, constituting the primary response to b-adrenergic stimulation in the heart. Disruptions in the phosphorylation cycle progress to HF. The R9C mutation (PLNR9C) and R14 deletion (PLNR14del), both located in the PLN cytoplasmic domain, prevent PLN phosphorylation and lead to DCM via an unknown mechanism. Using a battery of biochemical, molecular biology, and spectroscopic methods, we will characterize the structural and dynamic effects of these mutations in the formation of the PKA/PLN complex and link them to their cardiotoxicity. Specifically, we plan to decipher the PLN recognition and phosphorylation mechanisms by the kinase in the presence and absence of these deadly mutations. Moreover, we will elucidate the role of lipid membranes and spatial localization via myristoylation in the phosphorylation process. These studies will set the groundwork for understanding the physiology and pathophysiology of phosphorylation signaling in the heart. PUBLIC HEALTH RELEVANCE: Dilated cardiomyopathy (DCM) is the leading cause for heart failure (HF) worldwide. This proposal seeks to elucidate the molecular determinants for DCM using biophysical, biochemical, and molecular biology approaches. Understanding the genesis of the disease at the atomic level will help translate this knowledge into molecular design of new therapeutic approaches to counteract HF.