Role of protein kinase C in the phosphorylation of cardiac myosin light chain 2.
Role of protein kinase C in the phosphorylation of cardiac myosin light chain 2.
复制标题
蛋白激酶C在心肌肌球蛋白轻链2磷酸化中的作用。
DOI：
10.1042/bj2940401
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Kuo,JF
中科院分区：
文献类型：
--
作者：
Venema,RC;Raynor,RL;NolandJr,TA;Kuo,JF
研究了蛋白激酶C(PKC)在成年大鼠心肌细胞肌球蛋白轻链2(MLC2)磷酸化中的作用。在体外成年大鼠心肌原纤维中，PKC介导的MLC2的磷酸化具有与肌球蛋白轻链激酶(MLCK)相似的化学计量比(0.7mol磷酸/摩尔蛋白质)。经PKC或MLCK体外磷酸化后的MLC2的二维胰酶磷酸肽图谱显示，每个蛋白激酶的主要磷酸肽是相同的。当分离的心肌细胞与[32P]P(I)孵育时，这些位置也被32P原位标记。心肌细胞中MLC2的~(32)P标记在与磷酸酶抑制剂Calyculin A孵育后10分钟内增加5倍，表明在这些细胞中存在MLC2磷酸化的快速翻转成分。当心肌细胞暴露在2，3-丁二酮单肟中时，~(32)P标记从MLC2中完全去掉。2，3-丁二酮单肟是一种已知的心脏收缩抑制剂，可使肌丝对钙激活不敏感。在PKC选择性抑制剂Calphostin C和Chelerythine作用下，MLC2的32P标记也减少了50%-100%，这表明PKC而不是MLCK主要负责将迅速转化的磷酸盐原位结合到MLC2中。综上所述，这些数据表明PKC参与了心肌细胞MLC2的磷酸化，并支持了心脏MLC2的磷酸化在决定肌原纤维钙敏感性中的作用的假说。
The role of protein kinase C (PKC) in the phosphorylation of myosin light chain 2 (MLC2) in adult rat heart cells has been investigated. PKC-mediated phosphorylation of MLC2 in adult rat cardiac myofibrils in vitro occurs with a stoichiometry (0.7 mol of phosphate/mol of protein) similar to that mediated by myosin light chain kinase (MLCK). Two-dimensional tryptic phosphopeptide mapping of MLC2 following phosphorylation by PKC or MLCK in vitro yields the same major phosphopeptides for each protein kinase. These sites are also 32P-labelled in situ when isolated cardiomyocytes are incubated with [32P]P(i). 32P labelling of MLC2 in cardiomyocytes is increased by 5-fold in 10 min upon incubation with the phosphatase inhibitor calyculin A, demonstrating the existence of a rapidly turning over component of MLC2 phosphorylation in these cells. 32P label is completely removed from MLC2 when myocytes are exposed to 2,3-butanedione monoxime, an inhibitor of cardiac contraction known to desensitize the myofilaments to activation by Ca2+. 32P labelling of MLC2 is also decreased by 50-100% following exposure to the PKC-selective inhibitors calphostin C and chelerythrine, suggesting that PKC, and not MLCK, is primarily responsible for incorporation of rapidly turning over phosphate into MLC2 in situ. Taken together, these data implicate PKC in the phosphorylation of MLC2 in heart cells and support the hypothesis that phosphorylation of cardiac MLC2 has a role in determining myofibrillar Ca2+ sensitivity.