Role of Ras in signal transduction from the nerve growth factor receptor: relationship to protein kinase C, calcium and cyclic AMP.
A dominant inhibitory ras mutant (Ha-ras Asn-17) has been used to investigate the role of Ras in nerve growth factor (NGF)-mediated signal transduction in PC12 cells. Expression of Ha-Ras Asn-17 blocks neuronal differentiation of these cells in response to NGF treatment. The Ha-Ras Asn-17 block was bypassed by treatment with NGF plus dibutyryl cAMP or NGF plus the Ca2+ ionophore ionomycin, but not by NGF plus 12-O-tetradecanoyl phorbol acetate (TPA). Direct stimulation of the cAMP or Ca2+ pathways thus appeared to act synergistically with a Ras-independent NGF signaling pathway. This Ras-independent pathway was also distinct from protein kinase C, since its activity was not affected by protein kinase C down-regulation. It thus appears that NGF stimulation generates a Ras-independent intracellular signal that contributes to neuronal differentiation independently of the cAMP, Ca2+ or protein kinase C second messenger systems. Since TPA did not bypass the Ha-Ras Asn-17 block to differentiation, protein kinase C also did not appear to be sufficient for Ras-dependent pathways mediating NGF-induced differentiation. Down-regulation experiments further indicated that protein kinase C was not required for NGF induction of early response genes via either Ras-dependent or Ras-independent pathways. Moreover, the formation of inositol phosphates and mobilization of intracellular calcium in response to NGF was not inhibited in PC12 cells expressing the Ha-Ras Asn-17 protein. Therefore, although calcium was able to bypass the Ha-Ras Asn-17 block to PC12 differentiation, Ras activity was not required for activation of phospholipase C in response to NGF. It thus appears that both Ras-dependent and Ras-independent signaling pathways contribute to NGF-induced PC12 cell differentiation independently of the cAMP, calcium and protein kinase C second messenger systems.
Szeberényi
J
J
Dana-Farber Cancer Institute, Boston, Massachusetts.
Erhardt
P
P
Cai
H
H
Cooper
G M
GM
D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000502:physiology*; D002454:Cell Differentiation; D000242:Cyclic AMP / Q000502:physiology*; D005786:Gene Expression Regulation; D016762:Genes, fos; D011905:Genes, ras; D007295:Inositol Phosphates / Q000378:metabolism; D015759:Ionomycin / Q000494:pharmacology; D008969:Molecular Sequence Data; D009414:Nerve Growth Factors / Q000494:pharmacology; D011493:Protein Kinase C / Q000502:physiology*; D051381:Rats; D017475:Receptors, Nerve Growth Factor / Q000502:physiology*; D015398:Signal Transduction; D014407:Tumor Cells, Cultured
D007295:Inositol Phosphates; D009414:Nerve Growth Factors; D017475:Receptors, Nerve Growth Factor; D015759:Ionomycin; D000242:Cyclic AMP; D011493:Protein Kinase C; D002118:Calcium