Proteome analysis of phosphorylation signaling pathways involved in maxillofacial development

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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.