Dixon B S, Breckon R, Burke C, Anderson R J
Department of Medicine, Veterans Administration Hospital, Denver, Colorado.
Am J Physiol. 1988 Jan;254(1 Pt 1):C183-91. doi: 10.1152/ajpcell.1988.254.1.C183.
Activators of protein kinase C, a calcium- and phospholipid-dependent protein kinase, inhibit vasopressin-stimulated water flow in toad bladder. To determine the biochemical mechanisms of this inhibition, we examined the effects of activators of protein kinase C on arginine vasopressin (AVP)-stimulated adenylate cyclase activity in cultured rabbit cortical collecting tubular cells. The phorbol ester, 4 beta-phorbol 12-myristate 13-acetate (PMA), the diacylglycerol, 1-oleyl-2-acetyl glycerol (OAG), and the diacylglycerol kinase inhibitor, R59022, all rapidly activate protein kinase C in collecting tubular cells. Pretreatment with PMA produces a delayed inhibition (greater than or equal to 4 h) of AVP-stimulated adenylate cyclase activity. The 4-h time lag suggests that the effects of protein kinase C are mediated indirectly, possibly as a consequence of stimulating cell proliferation. PMA does not inhibit cholera toxin- or forskolin-stimulated adenylate cyclase activity, suggesting an effect on the vasopressin receptor or coupling of the receptor to the stimulatory guanine nucleotide regulatory protein. Neither prostaglandins nor the inhibitory guanine nucleotide regulatory protein appear to mediate this effect. In contrast, treatment with either OAG or R59022 produces a rapid inhibition of both AVP- and forskolin-stimulated adenylate cyclase activity suggesting a prominent distal site of action, presumably at the catalytic subunit of adenylate cyclase. The results demonstrate that different activators of protein kinase C inhibit AVP-stimulated adenylate cyclase activity by distinctly different mechanisms possibly by altering the substrate specificity or activating multiple forms of the kinase. These results have important implications when using different activators to study the biological effects of protein kinase C.
蛋白激酶C（一种钙和磷脂依赖性蛋白激酶）的激活剂可抑制蟾蜍膀胱中血管加压素刺激的水流动。为了确定这种抑制作用的生化机制，我们研究了蛋白激酶C激活剂对培养的兔皮质集合管细胞中精氨酸血管加压素（AVP）刺激的腺苷酸环化酶活性的影响。佛波酯4β-佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）、二酰基甘油1-油酰基-2-乙酰甘油（OAG）和二酰基甘油激酶抑制剂R59022，均可快速激活集合管细胞中的蛋白激酶C。用PMA预处理会对AVP刺激的腺苷酸环化酶活性产生延迟抑制（大于或等于4小时）。4小时的时间延迟表明蛋白激酶C的作用是间接介导的，可能是刺激细胞增殖的结果。PMA不会抑制霍乱毒素或福斯高林刺激的腺苷酸环化酶活性，这表明其对血管加压素受体或受体与刺激性鸟嘌呤核苷酸调节蛋白的偶联有影响。前列腺素和抑制性鸟嘌呤核苷酸调节蛋白似乎均不介导这种作用。相比之下，用OAG或R59022处理会对AVP和福斯高林刺激的腺苷酸环化酶活性产生快速抑制，这表明作用位点在远端，可能是在腺苷酸环化酶的催化亚基上。结果表明，蛋白激酶C的不同激活剂通过明显不同的机制抑制AVP刺激的腺苷酸环化酶活性，可能是通过改变底物特异性或激活激酶的多种形式。这些结果对于使用不同激活剂研究蛋白激酶C的生物学效应具有重要意义。
