Fanger G R, Vaillancourt R R, Heasley L E, Montmayeur J P, Johnson G L, Maue R A
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755, USA.
Mol Cell Biol. 1997 Jan;17(1):89-99. doi: 10.1128/MCB.17.1.89.
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) beta receptors with mutations that eliminate activation of specific signalling molecules. Mutations eliminating activation of phosphatidylinositol 3-kinase (PI3K), phospholipase C gamma (PLC gamma), 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 alpha-subunit mRNA intact. Mutation of juxtamembrane tyrosines in combination with mutations eliminating activation of PI3K, PLC gamma, GAP, and Syp abolished the induction of type II alpha-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-fos and 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.
目前对于神经元分化的调控机制，包括神经营养因子诱导电压依赖性钠（Na+）通道表达的信号传导机制（该过程独立于Ras活性发生），我们还了解得不够透彻。因此，我们对稳定表达血小板衍生生长因子（PDGF）β受体的PC12细胞亚系进行了分析，这些受体带有消除特定信号分子激活的突变。通过核糖核酸酶保护试验和全细胞膜片钳记录测定，消除磷脂酰肌醇3激酶（PI3K）、磷脂酶Cγ（PLCγ）、GTP酶激活蛋白（GAP）和Syp磷酸酶激活的突变未能减少PDGF对II型Na+通道α亚基mRNA的诱导以及功能性Na+通道的表达。然而，受体激活时与Src激酶家族成员结合的近膜酪氨酸突变抑制了功能性Na+通道的诱导，而II型α亚基mRNA的诱导却不受影响。近膜酪氨酸突变与消除PI3K、PLCγ、GAP和Syp激活的突变相结合，消除了II型α亚基mRNA的诱导，这表明至少部分冗余的信号传导机制介导了这种诱导。受体突变对Na+通道表达的不同影响并不反映受体信号传导能力的整体变化，因为在所有分析的突变受体中，c-fos和转胶酶mRNA的诱导仍然会发生。这些结果揭示了Src家族在Na+通道表达诱导中的重要作用，并突出了调控神经元分化的信号传导机制的多样性和组合性质。
