Effects and regulation of phosphatase inhibitor-1 in the heart and its therapeutic potential in heart failure

磷酸酶抑制剂-1对心脏的作用和调节及其治疗心力衰竭的潜力

• 批准号: 13327559
• 负责人: Professor Dr. Ali El-Armouche
• 金额: --
• 依托单位: Institut für Pharmakologie und Toxikologie
• 依托单位国家: 德国
• 项目类别: Research Units
• 财政年份: 2005
• 资助国家: 德国
• 起止时间: 2004-12-31 至 2011-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/13327559?language=en
• 关键词: Effects; regulation; phosphatase; inhibitor; heart

Control of protein phosphorylation/dephosphorylation is an integral component of intracellular signal transduction. Phosphatase inhibitor-1 (I-1) is a prototypical mediator of cross-talk between kinases and phosphatases. Activated cAMP-dependent protein kinase (PKA) phosphorylates I-1 at Thr-35, converting it into a potent inhibitor of phosphatase-1. Inhibition of phosphatase-1 subsequently enhances PKA-mediated protein phosphorylation. We showed that adenovirally overexpression of I-1 does indeed amplify ß-agonist responses in adult rat cardiac myocytes. This was associated with increased phosphorylation of phospholamban (PLB), a key regulator of cardiac function. Conversely, in samples from failing human hearts we observed that downregulation and dephosphorylation of I-1 at Thr-35 was associated with decreased phosphorylation of PLB. In a recent study, phosphorylation of I-1 at Ser-67 has been reported to reduce its ability to inhibit phosphatase-1. Interestingly, a rise in intracellular Ca2+ could trigger the dephosphorylation of both phosphorylation sites by the same Ca2+-activated phosphatase 2B (calcineurin). The aim of this project is (i) to test whether overexpression of I-1 has different effects on phosphorylation of various PKA substrates (e.g. PLB as opposed to the ryanodine receptor), (ii) to evaluate I-1´s regulation by kinases/phosphatases in cardiac myocytes, (iii) to characterize I-1 overexpressing mice (cardiac specific I-1 TG), (iv) to generate and to characterize transgenic mice overexpressing mutated I-1 (Ser-67 replaced by Ala on an I-1 knockout background) under control of a regulatable promoter (Tet-Off system), and (v) to cross-breed I-1 TG mice with mice models with contractile dysfunction. The results are expected to extend our understanding of I-1´s role in cardiac myocyte signaling and contribute to answering the question whether I-1 may constitute a potential target in treating the depressed function of failing hearts.

蛋白质磷酸化/去磷酸化的控制是细胞内信号转导的组成部分。磷酸酶抑制剂-1（I-1）是激酶和磷酸酶之间的典型交互作用介质。活化的cAMP依赖性蛋白激酶（PKA）在Thr-35磷酸化I-1，将其转化为磷酸酶-1的强效抑制剂。磷酸酶-1的抑制随后增强PKA介导的蛋白磷酸化。我们发现，腺病毒过表达I-1确实放大了成年大鼠心肌细胞的β激动剂反应。这与受磷蛋白（PLB）的磷酸化增加有关，受磷蛋白是心脏功能的关键调节因子。相反，在衰竭的人类心脏样本中，我们观察到I-1在Thr-35的下调和去磷酸化与PLB磷酸化的降低相关。在最近的一项研究中，据报道I-1在Ser-67的磷酸化降低了其抑制磷酸酶-1的能力。有趣的是，细胞内Ca 2+的升高可以通过相同的Ca 2+激活的磷酸酶2B（钙调神经磷酸酶）触发两个磷酸化位点的去磷酸化。本项目的目的是（i）检测I-1的过表达是否对不同PKA底物的磷酸化有不同的影响（例如PLB，而不是ryanodine受体），（ii）评估心肌细胞中激酶/磷酸酶对I-1的调节，（iii）表征I-1过表达小鼠（心脏特异性I-1 TG），（iv）产生和表征过表达突变的I-1的转基因小鼠（在I-1敲除背景下Ser-67被Ala替代），以及（v）将I-1 TG小鼠与具有收缩功能障碍的小鼠模型杂交。这些结果有望扩展我们对I-1在心肌细胞信号传导中作用的理解，并有助于回答I-1是否可能成为治疗衰竭心脏功能低下的潜在靶点的问题。