MODULATION OF CALCIUM CONTROL OF CARDIAC MYOFIBRILS

描述(根据调查员的申请改编)：申请人的 目的是了解肌丝对钙离子的反应如何以及何时发生改变。 调节机械和生化活性的动态和水平 心肌。磷脂蛋白的体外观察与研究 基因敲除的老鼠表明了一个重要的作用，但到目前为止还没有实验表明 特异地确定了改变的肌丝对钙的反应的影响 关于心肌细胞和心脏的活动和动力学。要解决这个问题 已经产生了过量表达异构体和突变体的转基因小鼠 肌钙蛋白I(TnI)和原肌球蛋白(TM)--触发 并激活肌动蛋白-肌球蛋白反应。过度表达的蛋白质，显示出 与本地同行的交流，包括β-TM和慢骨骼TnI- 胎儿的异构体。其他转基因小鼠被认为是过度表达 突变型TnI缺乏蛋白激酶C(PKC)和PKA位点及突变型TM‘s 用不同的罪名。目标涉及以下问题：目标1和 2：TM亚型转换、错义突变和 磷酸化对心肌收缩和松弛的影响？是不同吗？ 与TN与细丝的结合有关吗？是函数的大小 细丝单位改变了吗？目标3：TNI是否是决定 心肌和快/慢骨骼肌在pH效应上的差异 肌节长度对肌丝对钙离子反应的影响？目标4和目标5：什么 TnI在其独特的PKA和PKC处的磷酸化的意义 心脏肌肉收缩和放松的部位？机械和 从以下几个组织级别进行细胞内测量 心脏原位跳动的单细胞和多细胞制剂。这个 研究人员的实验提供了：1)洞察 生理环境中肌丝对原位钙离子反应的改变 2)对这一监管机制的重要性有更清晰的认识 心脏病理学，特别是家族性肥厚性肌病。

DESCRIPTION (Adapted from the investigator's application): The applicant's objective is to know how and when altered myofilament response to Ca2+ regulates dynamics and level of mechanical and biochemical activity of cardiac muscle. In vitro observations and studies on the phospholamban knock-out mouse indicate an important role, but as yet no experiments have specifically determined the impact of altered myofilament response to Ca2+ on activity and dynamics of myocytes and hearts. To approach this problem transgenic mice have been generated which over express isoforms and mutants of troponin I (TnI) and tropomyosin (Tm)-regulatory proteins that trigger and activate the actin-myosin reaction. Overexpressed proteins, shown to exchange with native counterparts, include beta-Tm and slow skeletal TnI- the fetal isoforms. Other transgenic mice are proposed that over-express mutant TnI's lacking protein kinase C (PKC) and PKA sites and mutant Tm's with different charge. The objectives address these questions: Aims 1 and 2: What is the impact of Tm isoform switching, missense mutations and phosphorylation on heart muscle contraction and relaxation? Are differences in binding of Tn to thin filaments involved? Is the size of the functional unit of thin filaments altered? Aim 3: Is TnI the key element determining differences between cardiac and fast/slow skeletal muscle in effects of pH and sarcomere length on myofilament response to Ca2+? Aims 4 and 5: What is the significance of phosphorylation of TnI at its unique PKA and PKC sites on heart muscle contraction and relaxation? Mechanical and intracellular measurements are made at several levels of organization from single and multi-cellular preparations to hearts beating in situ. The investigators' experiments provide: 1) insight into the importance of altered myofilament response to Ca2+ in situ in the physiological context and 2) clearer understanding of the importance of this regulatory mechanism in cardiac pathology especially familial hypertrophic myopathies.