MODULATION OF CALCIUM CONTROL OF CARDIAC MYOFIBRILS

描述（改编自研究者的申请）：申请人的 目的是了解肌丝对Ca 2+的反应如何以及何时改变 调节机械和生物化学活动的动力学和水平， 心肌 受磷蛋白的体外观察和研究 基因敲除小鼠显示出重要作用，但迄今为止还没有实验 特别确定了改变肌丝对Ca 2+反应的影响， 对肌细胞和心脏的活动和动力学的影响。 为了解决这个问题 已经产生过表达同种型和突变体的转基因小鼠 肌钙蛋白I（TnI）和原肌球蛋白（Tm）的调节蛋白， 激活肌动蛋白-肌球蛋白反应。 过度表达的蛋白质， 与天然对应物交换，包括β-Tm和慢骨骼肌TnI- 胎儿亚型 其他的转基因小鼠被认为过度表达 缺乏蛋白激酶C（PKC）和PKA位点的突变TnI和突变Tm 不同的收费。 目标1和 2：Tm同工型转换、错义突变和 磷酸化对心肌收缩和舒张的影响 存在差异 在Tn与细丝的结合中起作用吗 函数的大小 一个单位的细丝改变了？ 目标3：TnI是决定 心脏和快/慢骨骼肌之间pH效应的差异 肌节长度对肌丝对Ca 2+？ 目标4和5：什么 是TnI磷酸化在其独特的PKA和PKC中的意义 关于心肌收缩和舒张的网站 机械和 细胞内测量是在几个层次的组织， 单细胞和多细胞制备物用于原位跳动的心脏。 的 研究人员的实验提供：1）洞察的重要性， 在生理环境中改变肌丝对Ca 2+的原位反应 二是对这一监管机制的重要性有了更清晰的认识 在心脏病理学，特别是家族性肥大性肌病中。

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.