Szallasi Z, Denning M F, Smith C B, Dlugosz A A, Yuspa S H, Pettit G R, Blumberg P M
Laboratory of Cellular Carcinogenesis, National Cancer Institute, Bethesda, Maryland 20892.
Mol Pharmacol. 1994 Nov;46(5):840-50.
Bryostatin 1 and phorbol-12-myristate-13-acetate (PMA) are both potent activators of protein kinase C (PKC), although in primary mouse keratinocytes bryostatin 1 does not induce differentiation and blocks PMA-induced differentiation. We report here that in primary mouse keratinocytes PMA caused translocation of PKC-epsilon to the Triton X-100-soluble fraction with an approximately 2-order of magnitude higher potency, compared with translocation of PKC-alpha and PKC-delta. The kinetics of translocation were fastest for PKC-epsilon, slower for PKC-alpha, and slowest for PKC-delta. At 5-20 min bryostatin 1 showed potency similar to that of PMA for translocating PKC-alpha, higher potency for translocating PKC-delta, and lower potency for translocating PKC-epsilon. At a later time (6 hr), bryostatin 1 was 1-2 orders magnitude more potent than PMA for causing loss of PKC-alpha, -delta, and -epsilon from the soluble fraction. Bryostatin 1 was 40-fold more potent than PMA for down-regulating PKC-alpha and showed a biphasic dose-response curve for down-regulating PKC-delta. Bryostatin 1 at 0.1-1 nM down-regulated PKC-delta to a similar extent as did PMA. Bryostatin 1 at 100 nM to 1 microM, on the other hand, failed to induce down-regulation, and these high (100 nM to 1 microM) doses of bryostatin 1 showed noncompetitive inhibition of PKC-delta down-regulation by 1 microM PMA after coapplication. This protected portion of PKC-delta retained kinase activity. The dose-response curve for bryostatin 1 protection of PKC-delta from down-regulation by PMA correlated with bryostatin 1 inhibition of the effects of PMA on cornified envelope formation (a marker of differentiation) and epidermal growth factor binding. Although PKC-epsilon was readily translocated by both PMA and bryostatin 1, the PKC-epsilon originally associated with the particulate fraction showed no down-regulation by either of these agents. We hypothesize that differential regulation of PKC isozymes by PMA and bryostatin 1 may contribute to the different patterns of biological responses that they induce.
苔藓抑素1和佛波醇-12-肉豆蔻酸酯-13-乙酸酯（PMA）都是蛋白激酶C（PKC）的有效激活剂，尽管在原代小鼠角质形成细胞中，苔藓抑素1不会诱导分化，反而会阻断PMA诱导的分化。我们在此报告，在原代小鼠角质形成细胞中，与PKC-α和PKC-δ的转位相比，PMA导致PKC-ε转位至Triton X-100可溶性组分的效力高约2个数量级。转位动力学方面，PKC-ε最快，PKC-α较慢，PKC-δ最慢。在5 - 20分钟时，苔藓抑素1在使PKC-α转位方面显示出与PMA相似的效力，使PKC-δ转位的效力更高，而使PKC-ε转位的效力较低。在稍后的时间点（6小时），苔藓抑素1在使PKC-α、-δ和-ε从可溶性组分中丢失方面比PMA强1 - 2个数量级。苔藓抑素1在下调PKC-α方面比PMA强40倍，并且在下调PKC-δ方面呈现双相剂量反应曲线。0.1 - 1 nM的苔藓抑素1下调PKC-δ的程度与PMA相似。另一方面，100 nM至1 μM的苔藓抑素1未能诱导下调，并且在共同应用后，这些高剂量（100 nM至1 μM）的苔藓抑素1对1 μM PMA下调PKC-δ表现出非竞争性抑制。PKC-δ的这部分受保护区域保留了激酶活性。苔藓抑素1保护PKC-δ不被PMA下调的剂量反应曲线与苔藓抑素1抑制PMA对角质包膜形成（分化标志物）和表皮生长因子结合的作用相关。尽管PKC-ε很容易被PMA和苔藓抑素1转位，但最初与颗粒组分相关的PKC-ε不会被这两种试剂下调。我们推测，PMA和苔藓抑素1对PKC同工酶的差异调节可能导致它们诱导的不同生物学反应模式。
