P19, A hormonally regulated phosphoprotein of peptide hormone-producing cells: Secretagogue-induced phosphorylation in AtT-20 mouse pituitary tumor cells and in rat and hamster insulinoma cells

P19, a group of 19,000 mol wt cytosolic proteins, with apparent isoelectric points of pi 5.9, pi 5.7, and pi 5.4, respectively, was identified in three peptide hormone-producing cell types: AtT20 mouse pituitary tumor cells, RIN-1122 rat insulinoma cells, and hamster insulinoma cells. Secretagoguedependent phosphorylation of P19 was analyzed in ³²P-labeled cells by two-dimensional electrophoresis and autoradiography. The results were quantitated by computer-assisted derisitometry. Cellular levels of cAMP and hormone release were measured in parallel incubations. In addition to stimulating ACTH release, CRF raised the cellular level, of cAMP and increased the ³²P labeling of all three 19,000 mol wt proteins in AtT20 cells. Other agents known to act through cAMP, which included isoproterenol, forskolin, and 8-bromo-cAMP, mimicked the effect of CRF on both ACTH release and phosphorylation of P19.12-O-Tetradecanoylphorbol- 13-acetate, a tumor-promoting phorbol ester, also stimulated both ACTH release and phosphorylation of P19. In contrast, although 40 mM K⁺ promoted ACTH release, it did not affect the phosphorylation of P19. Analogous findings were observed in insulinoma cells. Glucagon stimulated insulin release, increased cellular cAMP and promoted phosphorylation of P19 in RIN 1122 cells. 12-0-Tetradecanoylphorbol-13-acetate also enhanced insulin release and the phosphorylation of P19 in these cells. The results obtained with hamster insulinoma cells closely resembled the observations in RIN-1122 cells. In conclusion, P19, an apparently homologous set of cytosolic proteins, undergoes phosphorylation in three peptide hormoneproducing cells in response to two groups of secretagogues, the effect of which is probably mediated, in one case, by cAMPdependent protein kinase and, in the other, by protein kinase C. The data suggest the possibility that P19 participates in a secretory pathway activated by these two effector systems.