Intact Ras function is required for sustained activation and nuclear translocation of extracellular signal-regulated kinases in nerve growth factor-stimulated PC12 cells.

PC12 pheochromocytoma cell lines expressing the dominant negative Ha-Ras Asn-17 protein at different levels were used in this study to analyze the relationship between nerve growth factor (NGF)-induced activation of members of the mitogen-activated protein kinase (MAPK) family, and neuritogenesis. In wild-type PC12 cells, NGF rapidly stimulated the extracellular signal-regulated kinases (ERKs). Kinase activation was sustained and was followed by the translocation of ERK 1 and ERK 2 into the nucleus ultimately leading to neurite outgrowth. In cells expressing relatively high levels of the inhibitory Ras protein, NGF stimulation of ERK 1 and ERK 2 as well as nuclear translocation of these protein kinases were greatly inhibited. In contrast, in PC12 subclones expressing low amounts of Ha-Ras Asn-17 the peak of ERK activation was only slightly reduced, but became transient in nature and was not followed by nuclear translocation of ERKs 1 and 2. Since all PC12 subclones expressing detectable levels of the dominant inhibitory Ras protein are resistant to NGF induction of neurite formation, our observations support the notion that sustained activation and translocation of ERKs into the nucleus are essential for NGF-induced neuronal differentiation of PC12 cells.