Proteome analysis of phosphorylation signaling pathways involved in maxillofacial development

P38MAPK通路是一种细胞内信号转导通路，通过磷酸化转导途径参与细胞发育和分化过程中细胞凋亡的调控。我们已经建立了一个细胞培养体系，在该体系中，通过p38途径的激活剂ASK1的表达，选择性地和结构性地激活p38MAPK，从而诱导PC12细胞的神经元分化。在这里，我们已经确定癌蛋白18(OP18)/stathmin是p38级联反应的一个新的潜在靶点。双向凝胶电泳法发现，在PC12细胞中，OP18/stathmin的磷酸化水平随着ASK1表达的增加而增加。SB2 0 35 80可减弱ASK1依赖性的OP18/STATHMIN的磷酸化，提示p38α和/或p38β参与了OP18/STATHMIN的磷酸化。我们一致地发现，在体外，p38的所有四种亚型都直接磷酸化OP18/stathmin，主要是在丝氨酸25处。结合定量逆转录聚合酶链式反应结果表明，p38α是PC12细胞中主要表达的异构体，在这些细胞中，ask1诱导的op18/stathmin的磷酸化似乎主要是通过p38α介导的。鉴于OP18/stathmin的微管破坏活性受其磷酸化的调节，ASK1-p38级联反应可能通过OP18/stathmin调节微管的动力学。

p38 MAP kinase pathway is an intracellular signal transduction pathway using a phosphorylation relay and is involved in the regulation of apoptosis during development and differentiation. We have established a cell culture system, in which neuronal differentiation of PC12 cells is induced by the selective and constitutive activation of p38 MAP kinase through the expression of constitutively active ASK1, an activator of the p38 pathway. Here we have identified oncoprotein 18 (Op18)/stathmin as a novel potential target of the p38 cascade. By two-dimensional electrophoresis, phosphorylation of Op18/stathmin was found to be increased upon the expression of constitutively active ASK1 in PC12 cells. The ASK1-dependent increase in the phosphorylation of Op18/stathmin was attenuated by the treatment with SB203580, suggesting that p38α and/or p38β contribute to the phosphorylation of Op18/stathmin. Consistently, we found that all four isoforms of p38 directly phosphorylated Op18/stathmin primarily at serine 25 in vitro. Taken together with the quantitative RT-PCR data indicating that p38α was the dominantly expressed isoform in PC12 cells, ASK1-induced phosphorylation of Op18/stathmin appears to be mediated mainly through p38α in these cells. Given that the microtubule-destabilizing activity of Op18/stathmin is regulated by its phosphorylation, the ASK1-p38 cascade may regulate microtubule dynamics through Op18/stathmin.