Engel Patti L, Kobayashi Tomoyoshi, Biesiadecki Brandon, Davis Jonathan, Tikunova Svetlana, Wu Steven, Solaro R John
Center for Cardiovascular Research, Department of Physiology and Biophysics, University of Illinois College of Medicine, Chicago, Illinois 60612, USA.
J Biol Chem. 2007 Jan 5;282(1):183-93. doi: 10.1074/jbc.M512337200. Epub 2006 Nov 12.
Force generating strong cross-bridges are required to fully activate cardiac thin filaments, but the molecular signaling mechanism remains unclear. Evidence demonstrating differential extents of cross-bridge-dependent activation of force, especially at acidic pH, in myofilaments in which slow skeletal troponin I (ssTnI) replaced cardiac TnI (cTnI) indicates the significance of a His in ssTnI that is an homologous Ala in cTnI. We compared cross-bridge-dependent activation in myofilaments regulated by cTnI, ssTnI, cTnI(A66H), or ssTnI(H34A). A drop from pH 7.0 to 6.5 induced enhanced cross-bridge-dependent activation in cTnI myofilaments, but depressed activation in cTnI(A66H) myofilaments. This same drop in pH depressed cross-bridge-dependent activation in both ssTnI myofilaments and ssTnI(H34A) myofilaments. Compared with controls, cTnI(A66H) myofilaments were desensitized to Ca(2+), whereas there was no difference in the Ca(2+)-force relationship between ssTnI and ssTnI(H34A) myofilaments. The mutations in cTnI and ssTnI did not affect Ca(2+) dissociation rates from cTnC at pH 7.0 or 6.5. However, at pH 6.5, cTnI(A66H) had lower affinity for cTnT than cTnI. We also probed cross-bridge-dependent activation in myofilaments regulated by cTnI(Q56A). Myofilaments containing cTnI(Q56A) demonstrated cross-bridge-dependent activation that was similar to controls containing cTnI at pH 7.0 and an enhanced cross-bridge-dependent activation at pH 6.5. We conclude that a localized N-terminal region of TnI comprised of amino acids 33-80, which interacts with C-terminal regions of cTnC and cTnT, is of particular significance in transducing signaling of thin filament activation by strong cross-bridges.
产生强力横桥对于完全激活心肌细肌丝是必需的，但分子信号传导机制仍不清楚。有证据表明，在慢肌骨骼肌肌钙蛋白I（ssTnI）取代心肌肌钙蛋白I（cTnI）的肌丝中，横桥依赖性力激活的程度存在差异，尤其是在酸性pH值下，这表明ssTnI中的一个组氨酸（His）具有重要意义，该组氨酸在cTnI中是一个同源的丙氨酸（Ala）。我们比较了由cTnI、ssTnI、cTnI（A66H）或ssTnI（H34A）调节的肌丝中横桥依赖性激活情况。pH值从7.0降至6.5会导致cTnI肌丝中横桥依赖性激活增强，但cTnI（A66H）肌丝中的激活受到抑制。相同的pH值下降会抑制ssTnI肌丝和ssTnI（H34A）肌丝中的横桥依赖性激活。与对照组相比，cTnI（A66H）肌丝对Ca(2+)脱敏，而ssTnI和ssTnI（H34A）肌丝之间的Ca(2+) - 力关系没有差异。cTnI和ssTnI中的突变在pH值为7.0或6.5时不影响Ca(2+)从cTnC的解离速率。然而，在pH值为6.5时，cTnI（A66H）对cTnT的亲和力低于cTnI。我们还探究了由cTnI（Q56A）调节的肌丝中横桥依赖性激活情况。含有cTnI（Q56A）的肌丝在pH值为7.0时表现出与含有cTnI的对照组相似的横桥依赖性激活，在pH值为6.5时表现出增强的横桥依赖性激活。我们得出结论，TnI的一个由氨基酸33 - 80组成的局部N端区域，它与cTnC和cTnT的C端区域相互作用，在通过强力横桥转导细肌丝激活信号方面具有特别重要的意义。
