College of Pharmacy, Ohio State University, Columbus, Ohio, USA.
Department of Physiology and Cell Biology, Ohio State University, Columbus, Ohio, USA.
J Biol Chem. 2020 Aug 14;295(33):11720-11728. doi: 10.1074/jbc.RA120.014054. Epub 2020 Jun 24.
Post-translational modifications of proteins involved in calcium handling in myocytes, such as the cardiac ryanodine receptor (RyR2), critically regulate cardiac contractility. Recent studies have suggested that phosphorylation of RyR2 by protein kinase G (PKG) might contribute to the cardioprotective effects of cholinergic stimulation. However, the specific mechanisms underlying these effects remain unclear. Here, using murine ventricular myocytes, immunoblotting, proximity ligation as-says, and nitric oxide imaging, we report that phosphorylation of Ser-2808 in RyR2 induced by the muscarinic receptor agonist carbachol is mediated by a signaling axis comprising phosphoinositide 3-phosphate kinase, Akt Ser/Thr kinase, nitric oxide synthase 1, nitric oxide, soluble guanylate cyclase, cyclic GMP (cGMP), and PKG. We found that this signaling pathway is compartmentalized in myocytes, as it was distinct from atrial natriuretic peptide receptor-cGMP-PKG-RyR2 Ser-2808 signaling and independent of muscarinic-induced phosphorylation of Ser-239 in vasodilator-stimulated phosphoprotein. These results provide detailed insights into muscarinic-induced PKG signaling and the mediators that regulate cardiac RyR2 phosphorylation critical for cardiovascular function.
肌细胞钙处理蛋白的翻译后修饰，如心脏兰尼碱受体（RyR2），对心肌收缩力有重要的调节作用。最近的研究表明，蛋白激酶 G（PKG）对 RyR2 的磷酸化可能有助于胆碱能刺激的心脏保护作用。然而，这些作用的具体机制尚不清楚。在这里，我们使用鼠心室肌细胞，通过免疫印迹、接近连接分析和一氧化氮成像，报告了由毒蕈碱受体激动剂卡巴胆碱诱导的 RyR2 的 Ser-2808 磷酸化是由包括磷酸肌醇 3-激酶、Akt Ser/Thr 激酶、一氧化氮合酶 1、一氧化氮、可溶性鸟苷酸环化酶、环鸟苷酸（cGMP）和 PKG 组成的信号轴介导的。我们发现，该信号通路在肌细胞中是分隔的，因为它与心房利钠肽受体-cGMP-PKG-RyR2 Ser-2808 信号通路不同，并且不依赖于毒蕈碱诱导的血管扩张刺激磷蛋白 Ser-239 磷酸化。这些结果为毒蕈碱诱导的 PKG 信号转导以及调节心脏 RyR2 磷酸化的介质提供了详细的见解，这对心血管功能至关重要。
