Protein kinase C regulates dopamine D4 receptor-mediated phospholipid methylation

Dopamine D4 receptors (D4 receptors) mediate dopamine-stimulated, folate-dependent phospholipid methylation. To investigate possible regulation of this multi-step D4 receptor-mediated phospholipid methylation cycle by protein kinases, specific kinase activators and inhibitors were studied in SK-N-MC human neuroblastoma cells, using [14C] formate to label folate-derived single-carbon groups. Phorbol dibutyrate (PDB), an activator of protein kinase C, stimulated basal phospholipid methylation and also shifted the dose-response curve for dopamine-stimulated phospholipid methylation to the right by more than an order of magnitude. Calphostin C, an inhibitor of protein kinase C, had little effect on basal phospholipid methylation but significantly inhibited dopamine-stimulated phospholipid methylation and also blocked the stimulatory response to PDB. Chelerythrine, which inhibits protein kinase C and other kinases, strongly inhibited both basal and dopamine-stimulated phospholipid methylation. Forskolin, an activator of protein kinase A, inhibited basal and dopamine-stimulated phospholipid methylation, but only at high concentrations while Rp-cAMP, an inhibitor of protein kinase A, did not block this effect. Inhibition of protein kinase G produced a modest decrease in dopamine-stimulated phospholipid methylation, but neither sodium nitroprusside, which increases nitric oxide (NO) production and activates protein kinase G, nor the NO synthase inhibitor N-nitro-L-arginine had any effect on basal or dopamine-stimulated phospholipid methylation. These observations indicate that protein kinase C is an important regulator of basal and D4 receptor-mediated folate-dependent phospholipid methylation, whereas protein kinase A and protein kinase G have a lesser or minimal role.