Dembinsky A, Rubin H, Ravid S
Department of Biochemistry, Hadassah Medical School, The Hebrew University, Jerusalem, Israel.
J Biol Chem. 1997 Jan 10;272(2):828-34. doi: 10.1074/jbc.272.2.828.
Myosin II heavy chain (MHC)-specific protein kinase C (MHC-PKC) isolated from the ameba, Dictyostelium discoideum, regulates myosin II assembly and localization in response to the chemoattractant cAMP. cAMP stimulation of Dictyostelium cells leads to translocation of MHC-PKC from the cytosol to the membrane fraction, as well as causing an increase in both MHC-PKC phosphorylation and its kinase activity. MHC-PKC undergoes autophosphorylation with each mole of kinase incorporating about 20 mol of phosphate. The MHC-PKC autophosphorylation sites are thought to be located within a domain at the COOH-terminal region of MHC-PKC that contains a cluster of 21 serine and threonine residues. Here we report that deletion of this domain abolished the ability of the enzyme to undergo autophosphorylation in vitro. Furthermore, after this deletion, cAMP-dependent autophosphorylation of MHC-PKC as well as cAMP-dependent increases in kinase activity and subcellular localization were also abolished. These results provide evidence for the role of autophosphorylation in the regulation of MHC-PKC and indicate that this MHC-PKC autophosphorylation is required for the kinase activation in response to cAMP and for subcellular localization.
从变形虫盘基网柄菌中分离出的肌球蛋白II重链（MHC）特异性蛋白激酶C（MHC-PKC），可响应趋化因子cAMP调节肌球蛋白II的组装和定位。对盘基网柄菌细胞进行cAMP刺激会导致MHC-PKC从细胞质转移至膜部分，同时导致MHC-PKC磷酸化及其激酶活性增加。每摩尔激酶中，MHC-PKC会发生自身磷酸化，结合约20摩尔的磷酸盐。MHC-PKC自身磷酸化位点被认为位于MHC-PKC羧基末端区域的一个结构域内，该结构域包含一簇21个丝氨酸和苏氨酸残基。在此我们报告，该结构域的缺失消除了该酶在体外进行自身磷酸化的能力。此外，在此缺失之后，MHC-PKC的cAMP依赖性自身磷酸化以及激酶活性和亚细胞定位的cAMP依赖性增加也被消除。这些结果为自身磷酸化在MHC-PKC调节中的作用提供了证据，并表明这种MHC-PKC自身磷酸化是响应cAMP激活激酶以及亚细胞定位所必需的。
