Analysis of Mutant Platelet-Derived Growth Factor Receptors Expressed in PC12 Cells Identifies Signals Governing Sodium Channel Induction during Neuronal Differentiation

The mechanisms governing neuronal differentiation, including the signals underlying the induction of voltage-dependent sodium (Na+) channel expression by neurotrophic factors, which occurs independent of Ras activity, are not well understood. Therefore, Na+channel induction was analyzed in sublines of PC12 cells stably expressing platelet-derived growth factor (PDGF) β receptors with mutations that eliminate activation of specific signalling molecules. Mutations eliminating activation of phosphatidylinositol 3-kinase (PI3K), phospholipase Cϒ(PLCϒ), the GTPase-activating protein (GAP), and Syp phosphatase failed to diminish the induction of type II Na+channel alpha-subunit mRNA and functional Na+channel expression by PDGF, as determined by RNase protection assays and whole-cell patch clamp recording. However, mutation of juxtamembrane tyrosines that bind members of the Src family of kinases upon receptor activation inhibited the induction of functional Na+channels while leaving the induction of type II α-subunit mRNA intact. Mutation of juxtamembrane tyrosines in combination with mutations eliminating activation of PI3K, PLCϒ, GAP, and Syp abolished the induction of type II α-subunit mRNA, suggesting that at least partially redundant signaling mechanisms mediate this induction. The differential effects of the receptor mutations on Na+channel expression did not reflect global changes in receptor signaling capabilities, as in all of the mutant receptors analyzed, the induction of c-fosand transin mRNAs still occurred. The results reveal an important role for the Src family in the induction of Na+channel expression and highlight the multiplicity and combinatorial nature of the signaling mechanisms governing neuronal differentiation.