Ueno E, Rosenberg P
Department of Pharmaceutical Sciences, University of Connecticut, School of Pharmacy, Storrs 06269, USA.
Biochem Pharmacol. 1996 Oct 25;52(8):1287-93. doi: 10.1016/0006-2952(96)00484-4.
The phospholipase A2 (PLA2) neurotoxin, beta-bungarotoxin (beta-BuTX), presynaptically alters acetylcholine release. We previously found that beta-BuTX inhibits protein phosphorylation in rat brain synaptosomes. This inhibition was not due to the inhibition of ATP synthesis, the action of arachidonic acid (AA) metabolites, or the stimulation of phosphatase activities. A typical PLA2 enzyme from Naja naja atra (N. n. atra) venom also inhibited phosphorylation but with lesser potency than that of beta-BuTX. We now report the effects of beta-BuTX and N. n. atra PLA2 on the activities of protein kinases. Treatments of synaptic plasma membrane or cytosol with N. n. atra PLA2 stimulated the activities of cAMP-dependent kinase, Ca2+/calmodulin-dependent kinase II, and protein kinase C (PKC), whereas beta-BuTX had no effect on these kinases. Calyculin A, a phosphatase-1 and -2A inhibitor, increased the stimulation of phosphorylation by N. n. atra PLA2, indicating that the stimulation is not due to an inhibition of phosphatase activities. The stimulation of PKC by N. n. atra PLA2 appears to be mediated by free fatty acids (FFAs) resulting from phospholipid hydrolysis by PLA2, since (1) treatment of either synaptic plasma membrane or cytosol with N. n. atra PLA2 produced large amounts of FFAs, and (2) AA, an exogenous FFA, stimulated PKC activity to an extent similar to that caused by N. n. atra PLA2. Thus, the mechanisms of action of beta-BuTX and N. n. atra PLA2 appear quite different from each other although both agents inhibit phosphorylation in intact synaptosomes.
磷脂酶A2（PLA2）神经毒素β-银环蛇毒素（β-BuTX）可在突触前改变乙酰胆碱的释放。我们之前发现β-BuTX可抑制大鼠脑突触体中的蛋白质磷酸化。这种抑制并非由于ATP合成的抑制、花生四烯酸（AA）代谢产物的作用或磷酸酶活性的刺激。来自眼镜蛇毒的一种典型PLA2酶也能抑制磷酸化，但效力低于β-BuTX。我们现在报告β-BuTX和眼镜蛇PLA2对蛋白激酶活性的影响。用眼镜蛇PLA2处理突触质膜或胞质溶胶可刺激环磷酸腺苷（cAMP）依赖性激酶、Ca2+/钙调蛋白依赖性激酶II和蛋白激酶C（PKC）的活性，而β-BuTX对这些激酶没有影响。磷酸酶-1和-2A抑制剂冈田酸可增强眼镜蛇PLA2对磷酸化的刺激作用，这表明这种刺激并非由于磷酸酶活性的抑制。眼镜蛇PLA2对PKC的刺激作用似乎是由PLA2水解磷脂产生的游离脂肪酸（FFA）介导的，因为：（1）用眼镜蛇PLA2处理突触质膜或胞质溶胶会产生大量FFA；（2）外源性FFA花生四烯酸刺激PKC活性的程度与眼镜蛇PLA2引起的程度相似。因此，尽管β-BuTX和眼镜蛇PLA2在完整突触体中均抑制磷酸化，但其作用机制似乎彼此有很大不同。
