SARCOPLASMIC RETICULUM FUNCTION IN NORMAL AND FAILING HEARTS

在心肌兴奋收缩偶联过程中，肌浆网(SR)对胞浆内游离钙离子浓度的调节起着至关重要的作用。肌浆网有三个主要功能：a)胞浆中钙离子的摄取，导致肌松弛；b)SR腔内的钙储存；c)这些功能分别是：钙离子转运ATPase(SERCA2)、钙储存蛋白Calequestrin和钙释放通道或兰诺定受体。磷蛋白(PLB)是另一种SR蛋白，在钙-ATPase活性和心肌收缩能力的调节中起着关键作用。在这个项目中，我们建议进一步研究PLB在哺乳动物心脏中的调节作用，并确定PLB和钙泵之间的化学计量偶联比率，这似乎是心脏收缩参数的关键决定因素。我们还建议通过抑制物-1调节其磷酸酶活性来阐明PLB的磷酸化状态在基础和β-激动剂条件下的收缩控制中的作用。此外，我们建议将我们的研究扩展到临床领域，并：a)筛选心力衰竭患者PLB和SR钙泵之间相互作用区域的点突变，这可能会改变这两种蛋白质相互作用的性质或程度，导致病理生理后果；以及b)评估PLB和SR Ca2+泵的水平以及PLB在人类衰竭心脏中的磷酸化程度。我们提出的研究将促进我们对正常和衰竭心脏中SR功能调节钙稳态的潜在机制的了解。他们还将提供有价值的见解，以了解不同的SR钙处理蛋白之间的串扰及其对心肌收缩能力的调节作用。

In cardiac excitation-contraction coupling, the sarcoplasmic reticulum (SR) plays an essential role in the regulation of the cytosolic free Ca2+ concentration. There are three major functions of the SR: a) Ca2+-uptake from the cytosol into the SR lumen resulting in muscle relaxation; b) Ca2+ storage in the SR lumen; and c) for these functions are: the Ca2+- transport ATPase (SERCA2), the Ca2+ storage protein calsequestrin and the Ca2+ release channel or ryanodine receptor, respectively. phospholamban (PLB) is another SR protein, which plays a crucial role in the regulation of the Ca2+-ATPase activity and myocardial contractility. In this project, we propose further studies on elucidating the regulatory role of PLB in the mammalian heart and defining the stoichiometric coupling ratio between PLB and the Ca2+-pump, which appears to be a key determinant of cardiac contractile parameters. We also propose to elucidate the role of the PLB phosphorylation status, through regulation of its phosphatase activity by inhibitor-1, in the control of contractility under basal and beta-agonist conditions. Furthermore, we propose to extend our studies to the clinical arena and: a) screen patients with heart failure for point mutations in the areas of interaction between PLB and the SR Ca2+-pump, which may modify the nature or degree of interaction of these two proteins, resulting in pathophysiological consequences; and b) assess the levels of PLB and the SR CA2+ pump as well as the degree of PLB phosphorylation in human failing hearts. Our proposed studies will advance our knowledge on the mechanisms underlying regulation of Ca2+ homeostasis by the SR function in the normal and failing heart. They will also provide valuable insights into the crosstalk between the various SR Ca2+ handling proteins and their regulatory effects on cardiac contractility.