MOLECULAR PHYSIOLOGY OF MYOCARDIAL TROPONIN I VARIANTS

描述（申请人的逐字描述）：心脏收缩 通过调节肌丝蛋白的相互作用发生， 增加细胞间钙浓度。收缩功能障碍 与心肌疾病相关的是与改变的反应， 收缩蛋白质转化为钙。肌钙蛋白肌钙蛋白的抑制性亚单位，肌钙蛋白 I是一种关键的调节蛋白，其基于其对收缩性的调节而调节收缩性。 磷酸化状态。最近，肌钙蛋白I的修饰和突变体 与缺血性损伤、心力衰竭和心肌病有关。的 该提案的目标是了解疾病相关的改变如何 肌钙蛋白I改变其功能并在心肌疾病中发挥中心作用 states.提出了一个全面的方法来界定这两个 功能改变的病理生理学和分子机制 心脏中的肌钙蛋白I变体。具体的变化将是 特征在于a）截短的变体（I - 193），缺失16个氨基酸， 来自羧基末端的残基，其概括了截短形式 由顿抑心肌中的钙依赖性蛋白水解产生，和B） 在蛋白激酶A中具有定点突变的肌钙蛋白I变体， 蛋白激酶C磷酸化位点，以确定这些位点在 内源性收缩，预适应，缺血/再灌注与心脏 失败和最后c）具有单个氨基酸突变的肌钙蛋白I变体 在抑制区复制最近描述的突变体中发现， 家族性肥厚型心肌病方法包括测量 转基因小鼠的心室力学， 电导微压计导管，稳态力研究-钙 在来自这些小鼠的Fura-2负载的完整骨小梁和体外骨小梁中， 用重组蛋白和合成肽进行实验， 改变了这些肌钙蛋白I变体的生化特性。这项工作将 确定心肌顿抑的分子机制，肌钙蛋白I的作用 体内磷酸化和一种形式的肥大性 心肌病预计所获得的信息也将 最终为开发新的治疗方法提供了合理的基础， 心功能不全

DESCRIPTION (the applicant's description verbatim): Contraction of the heart occurs through regulated interactions of the myofilament proteins in response to increasing intercellular calcium concentrations. Contractile dysfunction associated with myocardial disease is linked to an altered response of contractile proteins to calcium. The inhibitory subunit of troponin, troponin I, is a key regulatory protein which modulates contractility based on its phosphorylation status. Recently, modifications and mutants of troponin I have been associated with ischemic injury, heart failure and cardiomyopathy. The goal of this proposal is to understand how disease related alterations to troponin I modify its function and play a central role in myocardial disease states. A comprehensive approach is proposed to delineate the both the pathophysiology and molecular mechanisms of alteration of function produced by troponin I variants in the heart . Specific variants which will be characterized are a) truncated variant (I - 193), a loss of 16 amino acid residues from the carboxy-terminus, which recapitulates the truncated form produced by calcium dependent proteolysis in stunned myocardium, and b) troponin I variants with site-directed mutations in protein kinase A and protein kinase C phosphorylation sites to determine the role of these sites in intrinsic contractility, preconditioning, ischemia/reperfusion and heart failure and finally c) a troponin I variant with a single amino acid mutation in the inhibitory region reproducing a recently described mutant found in a familial hypertrophic cardiomyopathy. Methods include measurements of ventricular mechanics in transgenic mice using a miniaturized conductance-micromanometer catheter, studies of steady state force-calcium relationships in fura-2 loaded intact trabeculae from these mice and in vitro experiments with recombinant protein and synthesized peptides to dissect the altered biochemical properties of these troponin I variants. This work will determine a molecular mechanism of myocardial stunning, the role of troponin I phosphorylation in vivo and the pathophysiology of one form of hypertrophic cardiomyopathy. It is also anticipated that the information gained will ultimately provide a rational basis for the development of novel therapies for cardiac dysfunction.