ROLE OF PROTEIN-KINASE-C IN MEDIATING NGF EFFECT ON NEUROPEPTIDE-Y EXPRESSION IN PC12 CELLS

Neuropeptide Y (NPY) is a 36 amino acid peptide present in the central and peripheral nervous systems. Treatment with Nerve Growth Factor (NGF) induces an increase in NPY mRNA in PC12 cells, a rat pheochromocytoma cell line extensively used as a model of neuronal differentiation. Stimulators of both cAMP and calcium-phospholipid dependent protein kinases (PKA and PKC respectively) increase NPY mRNA levels in a similar way to NGF. Nevertheless, H-89, a specific inhibitor of PKA failed to block NGF stimulated NPY mRNA accumulation. Furthermore, direct measurement of PKA activity in cell extracts showed no increase following NGF, in contrast to forskolin. H7, an inhibitor of both PKC and PKA systems completely abolished the NGF induced increase in NPY mRNA, suggesting that PKC is necessary for NGF induction of the NPY gene. NGF also increased PKC activity in cell extracts in a similar way to phorbol myristate acetate (PMA). Use of a reporter function, chloramphenicol acetyl transferase, controlled by 700 base pairs of the 5' flanking region of the NPY gene demonstrated that NGF and phorbol ester stimulated transcription of the NPY gene. This stimulation could be blocked by pre-incubating PC12 cells with calphostin C, a specific inhibitor of PKC. Our results indicate that NGF induces NPY gene expression via activation of PKC system. Although an increase in adenylate cyclase activity affects the expression of the NPY gene, activation of PKA appears not to be involved in mediating the NGF effects.