Chen S G, Murakami K
Department of Biochemical Pharmacology, School of Pharmacy, State University of New York at Buffalo 14260, USA.
Neuroscience. 1995 Oct;68(4):1017-26. doi: 10.1016/0306-4522(95)00175-i.
cis-Unsaturated fatty acid, which activates protein kinase C in vitro, stimulates protein phosphorylation in intact hippocampal slices. Two protein bands (44,000 and 47,000 mol. wt) are particularly sensitive to cis-fatty acid and are phosphorylated in a dose- and time-dependent manner. The cis-fatty acid-stimulated protein phosphorylation can be further potentiated with diacylglycerol or 12-O-tetradecanoylphorbol 13-acetate. Several lines of evidence indicate that the cis-fatty acid-stimulated phosphorylation of these proteins is mediated by protein kinase C. First, the cis-fatty acid effect is mimicked by other protein kinase C activators such as diacylglycerol. Second, the stimulation of the phosphorylation by these activators can be blocked by staurosporine, which potently inhibits protein kinase C. Third, a concomitant application of cis-fatty acid and diacylglycerol or 12-O-tetradecanoylphorbol 13-acetate enhances the 44,000 and 47,000 mol. wt phosphorylation in a synergistic manner, which is a novel activation mode for protein kinase C. Fourth, they can be phosphorylated by purified protein kinase C (type III: alpha). Moreover, the synergistic activation of purified protein kinase C by cis-fatty acid and diacylglycerol leads to a drastic increase in the phosphorylation of these two protein bands. Two-dimensional gel electrophoresis and immunoblot analysis revealed that they are both acidic proteins. The 47,000 mol. wt band consists of two protein components; one is found to be F1/growth-associated protein-43 (pI = 4.5), and the other 47,000 mol, wt protein has broad pI ranging from 4.6 to 4.9. The 44,000 mol. wt component is a major phosphoprotein with pI of 4.8-5.1. Our results strongly indicate that cis-fatty acid can act as a regulator of endogenous protein kinase C in concert with diacylglycerol, and stimulate protein phosphorylation of its substrates such as F1/growth-associated protein-43 in the hippocampus.
顺式不饱和脂肪酸在体外可激活蛋白激酶C，它能刺激完整海马切片中的蛋白质磷酸化。两条蛋白带（分子量分别为44,000和47,000）对顺式脂肪酸特别敏感，并以剂量和时间依赖的方式被磷酸化。顺式脂肪酸刺激的蛋白质磷酸化可被二酰甘油或12 - O - 十四烷酰佛波醇13 - 乙酸酯进一步增强。几条证据表明，顺式脂肪酸刺激的这些蛋白质的磷酸化是由蛋白激酶C介导的。首先，顺式脂肪酸的作用可被其他蛋白激酶C激活剂如二酰甘油模拟。其次，这些激活剂对磷酸化的刺激可被能有效抑制蛋白激酶C的星形孢菌素阻断。第三，顺式脂肪酸与二酰甘油或12 - O - 十四烷酰佛波醇13 - 乙酸酯同时应用，以协同方式增强了分子量为44,000和47,000的蛋白的磷酸化，这是蛋白激酶C的一种新的激活模式。第四，它们可被纯化蛋白激酶C（III型：α）磷酸化。此外，顺式脂肪酸和二酰甘油对纯化蛋白激酶C的协同激活导致这两条蛋白带的磷酸化急剧增加。二维凝胶电泳和免疫印迹分析表明它们都是酸性蛋白。分子量为47,000的条带由两种蛋白质成分组成；一种被发现是F1/生长相关蛋白 - 43（pI = 4.5），另一种分子量为47,000的蛋白质的pI范围较宽，为4.6至4.9。分子量为44,000的成分是一种主要的磷蛋白，pI为4.8 - 5.1。我们的结果有力地表明，顺式脂肪酸可与二酰甘油协同作为内源性蛋白激酶C的调节剂，并刺激其底物如海马中的F1/生长相关蛋白 - 43的蛋白质磷酸化。
