Sibley D R, Jeffs R A, Daniel K, Nambi P, Lefkowitz R J
Arch Biochem Biophys. 1986 Jan;244(1):373-81. doi: 10.1016/0003-9861(86)90126-8.
Incubation of intact frog erythrocytes with 12-O-tetradecanoyl phorbol-13-acetate (TPA), a tumor-promoting phorbol diester which activates protein kinase C, results in an approximate two- to threefold increase in subsequently tested beta-adrenergic agonist-stimulated adenylate cyclase activity. This increase is due to an elevation in the Vmax of the enzyme rather than to a change in affinity for the agonist. TPA treatment of frog erythrocytes does not alter the affinity (KD) or the binding capacity (Bmax) for the beta-adrenergic antagonist [125I]cyanopindolol. In addition, agonist/[125I]cyanopindolol competition curves are not affected by TPA pretreatment nor is their sensitivity to guanine nucleotides. Incubation of frog erythrocyte membranes alone with TPA does not promote sensitization or activation of adenylate cyclase activity. Pretreatment of intact frog erythrocytes with TPA also produces approximately two- to threefold increases in basal, guanine nucleotide-, prostaglandin E1-, forskolin-, NaF-, and MnCl2-stimulated adenylate cyclase activities in frog erythrocyte membranes. This enhancement of adenylate cyclase activity by TPA is induced rapidly (t1/2 approximately equal to 5 min) and with an EC50 of about 10(-7) to 10(-6) M. Other tumor-promoting phorbol diesters or phorbol diester-like compounds including 4 beta-phorbol 12,13-dibutyrate, 4 beta-phorbol 12,13-didecanoate, and mezerein are effective in promoting enhanced adenylate cyclase activity. In contrast, phorbols such as 4 beta-phorbol, 4 alpha-phorbol 12,13-didecanoate, and 4-O-methylphorbol 12-myristate 13-acetate, which are inactive in tumor promotion and which do not activate protein kinase C, do not affect frog erythrocyte adenylate cyclase activity. These data are suggestive of a protein kinase C-mediated phosphorylation of one of the adenylate cyclase components that is distal to the receptor, i.e., the nucleotide regulatory and/or catalytic components.
用12 - O - 十四烷酰佛波醇 - 13 - 乙酸酯（TPA）孵育完整的青蛙红细胞，TPA是一种能激活蛋白激酶C的促肿瘤佛波醇二酯，结果导致随后检测的β - 肾上腺素能激动剂刺激的腺苷酸环化酶活性增加约两到三倍。这种增加是由于该酶的Vmax升高，而不是对激动剂的亲和力发生变化。用TPA处理青蛙红细胞不会改变对β - 肾上腺素能拮抗剂[125I]氰胍心安的亲和力（KD）或结合能力（Bmax）。此外，激动剂/[125I]氰胍心安竞争曲线不受TPA预处理的影响，对鸟嘌呤核苷酸的敏感性也不受影响。单独用TPA孵育青蛙红细胞膜不会促进腺苷酸环化酶活性致敏或激活。用TPA预处理完整的青蛙红细胞还会使青蛙红细胞膜中基础的、鸟嘌呤核苷酸、前列腺素E1、福斯可林、氟化钠和氯化锰刺激的腺苷酸环化酶活性增加约两到三倍。TPA对腺苷酸环化酶活性的这种增强作用诱导迅速（半衰期约等于5分钟），EC50约为10（-7）至10（-6）M。其他促肿瘤佛波醇二酯或佛波醇二酯样化合物，包括4β - 佛波醇12,13 - 二丁酸酯、4β - 佛波醇12,13 - 二癸酸酯和大戟二萜醇，在促进腺苷酸环化酶活性增强方面是有效的。相比之下，诸如4β - 佛波醇、4α - 佛波醇12,13 - 二癸酸酯和4 - O - 甲基佛波醇12 - 肉豆蔻酸酯13 - 乙酸酯等佛波醇，它们在肿瘤促进方面无活性且不激活蛋白激酶C，不会影响青蛙红细胞腺苷酸环化酶活性。这些数据提示蛋白激酶C介导了受体远端的腺苷酸环化酶成分之一的磷酸化，即核苷酸调节和/或催化成分。
