Denning M F, Dlugosz A A, Cheng C, Dempsey P J, Coffey R J, Threadgill D W, Magnuson T, Yuspa S H
Laboratory of Cellular Carcinogenesis and Tumor Promotion, National Cancer Institute, Bethesda, MD, USA.
Exp Dermatol. 2000 Jun;9(3):192-9. doi: 10.1034/j.1600-0625.2000.009003192.x.
The induction of epidermal differentiation by extracellular Ca2+ involves activation of both tyrosine kinase and protein kinase C (PKC) signaling cascades. To determine if the differentiation-dependent activation of tyrosine kinase signaling can influence the PKC pathway, we examined the tyrosine phosphorylation status of PKC isoforms in primary mouse keratinocytes stimulated to terminally differentiate with Ca2+. Elevation of extracellular Ca2+ induced tyrosine phosphorylation of PKC-delta, but not the other keratinocyte PKC isoforms (alpha, epsilon, eta, zeta). We have previously demonstrated that activation of the epidermal growth factor receptor (EGFR) pathway induces PKC-delta tyrosine phosphorylation in basal keratinocytes (Denning M F, Dlugosz A A, Threadgill D W, Magnuson T, Yuspa S H (1996) J Biol Chem 271: 5325-5331). When basal keratinocytes were stimulated to differentiate by Ca2+, the level of cell-associated transforming growth factor-alpha (TGF-alpha) increased 30-fold, while no increase in secreted TGF-alpha was detected. Furthermore, Ca2+-induced tyrosine phosphorylation of PKC-delta and phosphotyrosine-association of the receptor adapter protein Shc was diminished in EGFR -/- keratinocytes, suggesting that EGFR activation may occur during keratinocyte differentiation. Tyrosine phosphorylated PKC-delta was also detected in mouse epidermis, suggesting that this differentiation-associated signaling pathway is physiological. These results establish a requirement for the EGFR in Ca2+-induced tyrosine phosphorylation of PKC-delta, and document the production of cell-associated TGF-alpha in differentiated keratinocytes which may function independent of its usual mitogenic effects.
细胞外钙离子诱导表皮分化涉及酪氨酸激酶和蛋白激酶C（PKC）信号级联的激活。为了确定酪氨酸激酶信号的分化依赖性激活是否会影响PKC途径，我们检测了用钙离子刺激终末分化的原代小鼠角质形成细胞中PKC同工型的酪氨酸磷酸化状态。细胞外钙离子升高诱导了PKC-δ的酪氨酸磷酸化，但未诱导其他角质形成细胞PKC同工型（α、ε、η、ζ）的酪氨酸磷酸化。我们之前已经证明，表皮生长因子受体（EGFR）途径的激活可诱导基底角质形成细胞中PKC-δ的酪氨酸磷酸化（丹宁MF、德鲁戈什AA、思雷德吉尔DW、马格努森T、尤斯帕SH（1996年）《生物化学杂志》271:5325 - 5331）。当基底角质形成细胞被钙离子刺激分化时，细胞相关的转化生长因子-α（TGF-α）水平增加了30倍，而未检测到分泌的TGF-α增加。此外，在EGFR基因敲除的角质形成细胞中，钙离子诱导的PKC-δ酪氨酸磷酸化和受体衔接蛋白Shc的磷酸酪氨酸结合减少，这表明EGFR激活可能发生在角质形成细胞分化过程中。在小鼠表皮中也检测到了酪氨酸磷酸化的PKC-δ，这表明这种与分化相关的信号通路具有生理学意义。这些结果确定了EGFR在钙离子诱导的PKC-δ酪氨酸磷酸化中的必要性，并证明了在分化的角质形成细胞中产生的细胞相关TGF-α可能独立于其通常的促有丝分裂作用发挥功能。
