Impact of Cardiac Troponin T N-Terminal Deletion and Phosphorylation on Myofilament Function
Impact of Cardiac Troponin T N-Terminal Deletion and Phosphorylation on Myofilament Function
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DOI：
10.1021/bi900516n
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发表时间：
2009-08-18
期刊：
影响因子：
2.9
通讯作者：
Homsher, Earl
中科院分区：
文献类型：
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作者：
Sumandea, Marius P.;Vahebi, Susan;Homsher, Earl
心肌肌钙蛋白T（cTnT）是一种磷蛋白，通过与邻近的细丝蛋白广泛而多样的相互作用来调节心肌收缩。它的N-末端一半是将肌钙蛋白复合物锚定到原肌球蛋白-肌动蛋白的“胶水”。到目前为止，旨在研究N-末端尾区的作用的研究尚未考虑磷酸化的影响。为了更好地理解磷酸化cTnT N-末端尾区的调节作用，我们研究了N-末端缺失（氨基酸1-91）和磷酸化油Ca（2+）依赖性对肌丝等长收缩力产生、等长ATP酶速率和细丝滑动速度的功能影响。在洗涤剂透化纤维制备物中评估化学机械特征，其中在存在野生型cTn 1和cTnC的情况下，将天然肌钙蛋白（cTnI）与经改造以含有修饰cTnT（截短的、磷酸化的）的重组cTnI交换。去除cTnT N-末端氨基酸1-91（cTnT-del）可增强肌丝对非饱和Ca（2+）水平（心肌细胞的生理范围）的反应性。然而，在饱和Ca（2+）水平下，等长张力和ATP酶速率降低。一方面，cTnT-del的磷酸化减弱了由N-末端尾截短诱导的敏化效应，将肌丝Ca（2+）反应性“重置”回到对照水平。另一方面，它损害等长张力发展和ATP酶速率。有趣的是，cTnT的磷酸化（cTnT-P）差异调节张力成本（跨桥循环速率的所有指标）：cTn-del-P增加，而完整的cTn-wt-P减少。与等距纤维数据一样，与cTn-wt相比，cTn-del调节的细丝滑动速度对Ca（2+）更敏感。cTnT的磷酸化（无论是cTnT-del还是-wt）抑制了滑动速度，并且与细丝滑动速度的Ca（2+）脱敏有关。
Cardiac troponin T (cTnT) is a phosphoprotein that modulates cardiac muscle contraction through its extensive and diverse interactions with neighboring thin filament proteins. Its N-terminal half is the "glue" that anchors the troponin complex to tropomyosin-actin. Until now, studies aimed at investigating the role of the N-terminal tail region have not considered the effects of phosphorylation. To understand better the regulatory role of the N-terminal tail region of phosphorylated cTnT, we investigated the functional effects of N-terminal deletion (amino acids 1-91) and phosphorylation oil Ca(2+) dependence of myofilament isometric force production, isometric ATPase rate, and thin filament sliding speed. Chemomechanical profiles were assessed in detergent permeabilized fiber preparations where the native troponin (cTn) was exchanged with recombinant cTn engineered to contain modified cTnT (truncated, phosphorylated) in the presence of wild-type cTn1 and cTnC. Removal of the cTnT N-terminal amino acids 1-91 (cTnT-del) enhances myofilament responsiveness to nonsaturating Ca(2+) levels (the physiological range in cardiac myocytes). However, at saturating Ca(2+) levels, there is a reduction in isometric tension and ATPase rate. On one hand, phosphorylation of cTnT-del attenuates the sensitizing effect induced by truncation of the N-terminal tail, "resetting" myofilament Ca(2+) responsiveness back to control levels. On the other hand, it impairs isometric tension development and ATPase rate. Interestingly, phosphorylation of cTnT (cTnT-P) differentially regulates tension cost (all index of cross-bridge cycling rate): increased by cTn-del-P and decreased by intact cTn-wt-P. Like the isometric fiber data, sliding speed of thin filaments regulated by cTn-del is more sensitive to Ca(2+) compared with cTn-wt. Phosphorylation of cTnT (whether cTnT-del or -wt) depresses sliding speed and is associated with Ca(2+) desensitization of thin filament sliding speed.