Reynolds N J, Talwar H S, Baldassare J J, Henderson P A, Elder J T, Voorhees J J, Fisher G J
Department of Dermatology, University of Michigan, Ann Arbor 48109.
Biochem J. 1993 Sep 1;294 ( Pt 2)(Pt 2):535-44. doi: 10.1042/bj2940535.
We have investigated coupling between the epidermal growth factor (EGF) receptor and the phospholipase C (PLC)/protein kinase C (PKC) signal-transduction system in normal skin fibroblasts and keratinocytes, for which EGF and transforming growth factor alpha (TGF-alpha) are mitogenic. EGF and TGF-alpha induced a rapid increase in tyrosine phosphorylation of the EGF receptor, in both fibroblasts and keratinocytes, but failed to induce tyrosine phosphorylation of PLC-gamma 1 or detectable phosphoinositide hydrolysis, as measured by two sensitive assays. In fibroblasts, EGF induced phosphatidylcholine (PC) hydrolysis, resulting in increased diacylglycerol (DAG). In contrast, in keratinocytes, there was no detectable PC hydrolysis or elevation of DAG in response to EGF or TGF-alpha. EGF and TGF-alpha activated PKC in fibroblasts, as evidenced by increased phosphorylation of a specific cellular PKC substrate (myristoylated alanine-rich C-kinase substrate, 'MARCKS'). In keratinocytes, TGF-alpha and EGF induced only a modest increase in MARCKS protein phosphorylation. This apparent modest activation of PKC, in the absence of detectable DAG formation, may have been mediated by arachidonic acid, which was released from keratinocytes in response to TGF-alpha, and has been shown to stimulate PKC activity in vitro. These data demonstrate that (1) in dermal fibroblasts and keratinocytes, which express normal levels of EGF receptors, EGF receptor activation is not coupled to tyrosine phosphorylation of PLC-gamma 1 or PtdIns hydrolysis, suggesting that these events are not required for the mitogenic activity of EGF or TGF-alpha in these cells, (2) coupling of EGF receptor to PC hydrolysis is cell-type specific, and (3) in skin fibroblasts, DAG, formed through EGF-induced PC hydrolysis, is capable of activating PKC.
我们研究了正常皮肤成纤维细胞和角质形成细胞中表皮生长因子（EGF）受体与磷脂酶C（PLC）/蛋白激酶C（PKC）信号转导系统之间的偶联，其中EGF和转化生长因子α（TGF-α）具有促有丝分裂作用。EGF和TGF-α在成纤维细胞和角质形成细胞中均能迅速诱导EGF受体的酪氨酸磷酸化增加，但未能诱导PLC-γ1的酪氨酸磷酸化或通过两种灵敏检测方法测得的可检测到的磷酸肌醇水解。在成纤维细胞中，EGF诱导磷脂酰胆碱（PC）水解，导致二酰基甘油（DAG）增加。相反，在角质形成细胞中，未检测到PC水解或对EGF或TGF-α的DAG升高。EGF和TGF-α激活了成纤维细胞中的PKC，这通过特定细胞PKC底物（肉豆蔻酰化富含丙氨酸的C激酶底物，“MARCKS”）磷酸化增加得以证明。在角质形成细胞中，TGF-α和EGF仅诱导MARCKS蛋白磷酸化适度增加。在未检测到DAG形成的情况下，PKC这种明显的适度激活可能是由花生四烯酸介导的，花生四烯酸是角质形成细胞响应TGF-α释放的，并且已证明在体外可刺激PKC活性。这些数据表明：（1）在表达正常水平EGF受体的真皮成纤维细胞和角质形成细胞中，EGF受体激活与PLC-γ1的酪氨酸磷酸化或磷脂酰肌醇水解不偶联，这表明这些事件对于这些细胞中EGF或TGF-α的促有丝分裂活性不是必需的；（2）EGF受体与PC水解的偶联具有细胞类型特异性；（3）在皮肤成纤维细胞中，通过EGF诱导的PC水解形成的DAG能够激活PKC。
