Kirchhefer Uwe, Brekle Christiane, Eskandar John, Isensee Gunnar, Kučerová Dana, Müller Frank U, Pinet Florence, Schulte Jan S, Seidl Matthias D, Boknik Peter
From the Institut für Pharmakologie und Toxikologie, Universitätsklinikum Münster, D-48149 Münster, Germany and
From the Institut für Pharmakologie und Toxikologie, Universitätsklinikum Münster, D-48149 Münster, Germany and.
J Biol Chem. 2014 Dec 5;289(49):33862-73. doi: 10.1074/jbc.M114.598938. Epub 2014 Oct 15.
Dephosphorylation of important myocardial proteins is regulated by protein phosphatase 2A (PP2A), representing a heterotrimer that is comprised of catalytic, scaffolding, and regulatory (B) subunits. There is a multitude of B subunit family members directing the PP2A holoenzyme to different myocellular compartments. To gain a better understanding of how these B subunits contribute to the regulation of cardiac performance, we generated transgenic (TG) mice with cardiomyocyte-directed overexpression of B56α, a phosphoprotein of the PP2A-B56 family. The 2-fold overexpression of B56α was associated with an enhanced PP2A activity that was localized mainly in the cytoplasm and myofilament fraction. Contractility was enhanced both at the whole heart level and in isolated cardiomyocytes of TG compared with WT mice. However, peak amplitude of [Ca]i did not differ between TG and WT cardiomyocytes. The basal phosphorylation of cardiac troponin inhibitor (cTnI) and the myosin-binding protein C was reduced by 26 and 35%, respectively, in TG compared with WT hearts. The stimulation of β-adrenergic receptors by isoproterenol (ISO) resulted in an impaired contractile response of TG hearts. At a depolarizing potential of -5 mV, the ICa,L current density was decreased by 28% after administration of ISO in TG cardiomyocytes. In addition, the ISO-stimulated phosphorylation of phospholamban at Ser(16) was reduced by 27% in TG hearts. Thus, the increased PP2A-B56α activity in TG hearts is localized to specific subcellular sites leading to the dephosphorylation of important contractile proteins. This may result in higher myofilament Ca(2+) sensitivity and increased basal contractility in TG hearts. These effects were reversed by β-adrenergic stimulation.
重要心肌蛋白的去磷酸化由蛋白磷酸酶2A（PP2A）调节，PP2A是一种异源三聚体，由催化亚基、支架亚基和调节（B）亚基组成。有众多B亚基家族成员将PP2A全酶导向不同的心肌细胞区室。为了更好地理解这些B亚基如何对心脏功能调节产生影响，我们构建了转基因（TG）小鼠，使其在心肌细胞中定向过表达PP2A - B56家族的磷蛋白B56α。B56α的2倍过表达与PP2A活性增强相关，该活性主要定位于细胞质和肌丝部分。与野生型（WT）小鼠相比，TG小鼠的全心水平和分离的心肌细胞的收缩力均增强。然而，TG和WT心肌细胞之间的[Ca]i峰值幅度没有差异。与WT心脏相比，TG心脏中心肌肌钙蛋白抑制因子（cTnI）和肌球蛋白结合蛋白C的基础磷酸化分别降低了26%和35%。异丙肾上腺素（ISO）刺激β - 肾上腺素能受体导致TG心脏的收缩反应受损。在 - 5 mV的去极化电位下，TG心肌细胞给予ISO后L型钙电流（ICa,L）密度降低了28%。此外，TG心脏中ISO刺激的受磷蛋白在Ser(16)位点的磷酸化降低了27%。因此，TG心脏中增加的PP2A - B56α活性定位于特定的亚细胞位点，导致重要收缩蛋白的去磷酸化。这可能导致TG心脏中肌丝对Ca(2+)的敏感性更高以及基础收缩力增加。这些效应通过β - 肾上腺素能刺激得以逆转。
