Department of Pediatrics/Division of Cardiology, Johns Hopkins University School of Medicine, Ross Bldg 1144/720 Rutland Avenue, Baltimore, MD 21205, USA.
J Mol Cell Cardiol. 2010 May;48(5):943-53. doi: 10.1016/j.yjmcc.2010.01.004. Epub 2010 Jan 18.
Transgenic models with pseudo phosphorylation mutants of troponin I, PKA sites at Ser 22 and 23 (cTnIDD(22,23) mice) or PKC sites at Ser 42 and 44 (cTnIAD(22,23)DD(42,44)) displayed differential force-frequency relationships and afterload relaxation delay in vivo. We hypothesized that cTnI PKA and PKC phosphomimics impact cardiac muscle rate-related developed twitch force and relaxation kinetics in opposite directions. cTnIDD(22,23) transgenic mice produce a force frequency relationship (FFR) equivalent to control NTG albeit at lower peak Ca(2+), while cTnIAD(22,23)DD(42,44) TG mice had a flat FFR with normal peak systolic Ca(2+), thus suggestive of diminished responsiveness to Ca(2+) at higher frequencies. Force-Ca(2+) hysteresis analysis revealed that cTnIDD(22,23) mice have a combined enhanced myofilament calcium peak response with an enhanced slope of force development and decline per unit of Ca(2+), whereas cTnIAD(22,23)DD(42,44) transgenic mice showed the opposite. The computational ECME model predicts that the TG lines may be distinct from each other due to different rate constants for association/dissociation of Ca(2+) at the regulatory site of cTnC. Our data indicate that cTnI phosphorylation at PKA sites plays a critical role in the FFR by increasing relative myofilament responsiveness, and results in a distinctive transition between activation and relaxation, as displayed by force-Ca(2+) hysteresis loops. These findings may have important implications for understanding the specific contribution of cTnI to beta-adrenergic inotropy and lusitropy and to adverse contractile effects of PKC activation, which is relevant during heart failure development.
肌钙蛋白 I 的假磷酸化突变体(PKA 位点丝氨酸 22 和 23 处的 cTnIDD(22,23)小鼠或 PKC 位点丝氨酸 42 和 44 处的 cTnIAD(22,23)DD(42,44))的转基因模型在体内表现出不同的力频率关系和后负荷松弛延迟。我们假设肌钙蛋白 I 的 PKA 和 PKC 磷酸模拟物以相反的方向影响心肌与速率相关的收缩力和松弛动力学。cTnIDD(22,23)转基因小鼠产生与对照 NTG 相当的力频率关系(FFR),尽管峰值 Ca(2+)较低,而 cTnIAD(22,23)DD(42,44)TG 小鼠的 FFR 平坦,峰值收缩期 Ca(2+)正常,因此提示在较高频率时对 Ca(2+)的反应性降低。力-Ca(2+)滞后分析显示,cTnIDD(22,23)小鼠具有增强的肌球蛋白钙峰反应,同时具有增强的力发展斜率和单位 Ca(2+)的力下降,而 cTnIAD(22,23)DD(42,44)转基因小鼠则表现出相反的情况。ECME 计算模型预测,由于 cTnC 调节位点上 Ca(2+)的缔合/解离速率常数不同,TG 系可能彼此不同。我们的数据表明,PKA 位点肌钙蛋白 I 的磷酸化在 FFR 中起着关键作用,通过增加相对肌球蛋白反应性,导致激活和松弛之间的独特转变,如力-Ca(2+)滞后环所示。这些发现对于理解肌钙蛋白 I 对β肾上腺素能变力性和变松弛性的特定贡献以及 PKC 激活的不利收缩效应具有重要意义,这在心力衰竭发展过程中是相关的。