Wen-Sheng Wu, Jun-Ming Huang
Department of Medical Technology, TZU CHI University, Taiwan.
J Biomed Sci. 2005;12(2):289-96. doi: 10.1007/s11373-005-1210-5.
The signaling mechanisms for most of the antiproliferative processes are not fully understood. We have demonstrated that ERK(MAPK) signaling was involved in the induction of both p15(INK4b)and p16(INK4a) CDK inhibitors and growth inhibition of hepatoma cell HepG2 triggered by the tumor promoter tetradecanoyl phorbol acetate (TPA). In this study, the upstream signal mechanism for TPA-induced ERK(MAPK) activation was investigated. In HepG2 cells only one of the cPKC isozymes, PKCalpha, but not cPKCbetaII, nPKCepsilon or aPKCzeta was activated by TPA as demonstrated by its membrane translocation within 10-30 min and down-regulation at 24 h after TPA treatment. Pretreatment of 0.2-2.0 microM Bisindolylmaleimides, an inhibitor of PKC, attenuated the TPA-induced phosphorylation of ERK, gene expressions of p15(INK4b) and p16(INK4a), and growth inhibition of HepG2 cell in a dose-dependent manner. Consistently, transfection of HepG2 with 1.0-3.0 microM antisense (AS) PKCalpha, but not (AS) PKCbetaII, or nPKCepsilon oligonucleotides (ODN), for 36 h prior to TPA treatment also prevented the TPA-induced molecular and cellular effects described above. Taken together, we concluded that PKCalpha is specifically required for TPA-induced ERK(MAPK) signaling to trigger gene expressions of p15(INK4b) and p16(INK4a) leading to HepG2 growth inhibition.
大多数抗增殖过程的信号传导机制尚未完全明确。我们已经证明，ERK（丝裂原活化蛋白激酶）信号传导参与了肿瘤启动子十四酰佛波醇乙酸酯（TPA）诱导的p15（INK4b）和p16（INK4a）细胞周期蛋白依赖性激酶抑制剂的产生以及肝癌细胞HepG2的生长抑制。在本研究中，我们对TPA诱导的ERK（MAPK）激活的上游信号机制进行了研究。在HepG2细胞中，只有一种经典蛋白激酶C同工酶PKCalpha可被TPA激活，而不是经典蛋白激酶CbetaII、新型蛋白激酶Cepsilon或非典型蛋白激酶Czeta，这可通过其在TPA处理后10 - 30分钟内的膜转位以及24小时后的下调得以证明。用0.2 - 2.0微摩尔的双吲哚马来酰胺（一种蛋白激酶C抑制剂）预处理，可呈剂量依赖性地减弱TPA诱导的ERK磷酸化、p15（INK4b）和p16（INK4a）的基因表达以及HepG2细胞的生长抑制。同样，在TPA处理前36小时，用1.0 - 3.0微摩尔的反义（AS）PKCalpha而非（AS）PKCbetaII或新型蛋白激酶Cepsilon寡核苷酸（ODN）转染HepG2细胞，也可阻止上述TPA诱导的分子和细胞效应。综上所述，我们得出结论，PKCalpha是TPA诱导的ERK（MAPK）信号传导所特需的，该信号传导可触发p15（INK4b）和p16（INK4a）的基因表达，从而导致HepG2细胞生长抑制。
