Choi D Y, Toledo-Aral J J, Segal R, Halegoua S
Department of Neurobiology & Behavior, State University of New York at Stony Brook, Stony Brook, New York 11794-5230, USA.
Mol Cell Biol. 2001 Apr;21(8):2695-705. doi: 10.1128/MCB.21.8.2695-2705.2001.
In contrast to conventional signaling by growth factors that requires their continual presence, a 1-min pulse of nerve growth factor (NGF) is sufficient to induce electrical excitability in PC12 cells due to induction of the peripheral nerve type 1 (PN1) sodium channel gene. We have investigated the mechanism for this triggered signaling pathway by NGF in PC12 cells. Mutation of TrkA at key autophosphorylation sites indicates an essential role for the phospholipase C-gamma (PLC-gamma) binding site, but not the Shc binding site, for NGF-triggered induction of PN1. In concordance with results with Trk mutants, drug-mediated inhibition of PLC-gamma activity also blocks PN1 induction by NGF. Examination of the kinetics of TrkA autophosphorylation indicates that triggered signaling does not result from sustained activation and autophosphorylation of the TrkA receptor kinase, whose phosphorylation state declines rapidly after NGF removal. Rather, TrkA triggers an unexpectedly prolonged phosphorylation and activation of PLC-gamma signaling that is sustained for up to 2 h. Prevention of the elevation of intracellular Ca2+ levels using BAPTA-AM results in a block of PN1 induction by NGF. Sustained signaling by PLC-gamma provides a means for differential neuronal gene induction after transient exposure to NGF.
与需要生长因子持续存在的传统信号传导不同,由于外周神经1型(PN1)钠通道基因的诱导,1分钟的神经生长因子(NGF)脉冲足以诱导PC12细胞产生电兴奋性。我们研究了PC12细胞中NGF触发的这条信号通路的机制。TrkA关键自磷酸化位点的突变表明,磷脂酶C-γ(PLC-γ)结合位点而非Shc结合位点对于NGF触发的PN1诱导至关重要。与Trk突变体的结果一致,药物介导的PLC-γ活性抑制也会阻断NGF对PN1的诱导。对TrkA自磷酸化动力学的研究表明,触发信号并非源于TrkA受体激酶的持续激活和自磷酸化,在去除NGF后其磷酸化状态迅速下降。相反,TrkA触发了PLC-γ信号的意外长时间磷酸化和激活,这种激活可持续长达2小时。使用BAPTA-AM防止细胞内Ca2+水平升高会导致NGF对PN1诱导的阻断。PLC-γ的持续信号传导为短暂暴露于NGF后的差异性神经元基因诱导提供了一种方式。