Regulation of retinoic acid receptor alpha by protein kinase C in B16 mouse melanoma cells.
We have previously found that retinoic acid stimulates the expression of protein kinase C alpha (PKC) in B16 mouse melanoma cells. Because it has been reported that PKC can phosphorylate retinoic acid receptor (RAR) and alter its function, we determined whether changes in the level and/or activity of PKC could affect the expression or function of the RAR in B16 melanoma. Using in vivo phosphorylation and band shift techniques, we could not demonstrate that altering PKC activity and/or protein level changed the in vivo phosphorylation of RAR alpha. However activation of PKC resulted in increased RAR alpha protein. Increased receptor protein correlated with a phorbol dibutyrate-stimulated increase in receptor activation function-2 (AF-2)-dependent transcriptional activity. Use of enzyme inhibitors and dominant-negative PKCs indicated that enzyme activity was required for elevation in the RAR alpha. The PKC-mediated increase in RAR alpha was due to a 2.5-fold increase in the half-life of this protein. In contrast, the down-regulation of PKC diminished RAR alpha protein half-life and markedly inhibited AF-2-dependent transcriptional activity. The down-regulation of PKC also inhibited the binding of RAR to a retinoic acid response element and the retinoic acid induction of RAR beta expression. These findings suggest that PKC can influence retinoic acid signaling by altering the stability of RAR protein without directly phosphorylating this receptor.
Boskovic
Goran
G
Department of Biochemistry and Molecular Biology, Joan C. Edwards School of Medicine, Marshall University, Huntington, West Virginia 25704, USA.
Desai
Dinakar
D
Niles
Richard M
RM
D000818:Animals; D015153:Blotting, Western; D015536:Down-Regulation; D004789:Enzyme Activation; D007527:Isoenzymes / Q000378:metabolism*; D008545:Melanoma / Q000378:metabolism*; D051379:Mice; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D018168:Receptors, Retinoic Acid / Q000378:metabolism*; D000072482:Retinoic Acid Receptor alpha; D015398:Signal Transduction; D014162:Transfection; D014407:Tumor Cells, Cultured
D007527:Isoenzymes; C000607072:Rara protein, mouse; D018168:Receptors, Retinoic Acid; D000072482:Retinoic Acid Receptor alpha; C082925:retinoic acid receptor beta; D015240:Phorbol 12,13-Dibutyrate; C496630:Prkca protein, mouse; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha