Activation of distinct second messenger systems in anterior pituitary corticotrophic tumor cells alters the phosphorylation states of both shared and distinct cytosolic proteins.
The purpose of the present study was to investigate the effects of activation of different second messenger systems on protein phosphorylation in pituitary corticotrophic tumor cells (AtT-20/D16-16). Using two-dimensional gel analysis of cytosolic extracts from AtT-20 cells, several phosphoproteins exhibited alterations in 32P incorporation in response to stimulation of the cells with either forskolin--an activator of adenylate cyclase--or 12-O-tetradecanoyl phorbol-13-acetate (TPA)--a tumor promoting phorbol ester linked to protein kinase C activation. Alterations in phosphorylation levels were seen for phosphoproteins of the following apparent molecular weights and pIs: 87 kDa (pI 4.4-4.6), 67 kDa (pI 4.7-4.9), 43 kDa (pI 4.8-5.0), 39 kDa (pI 4.9-5.1), 33 kDa (pI 4.8-5.0), 19.5 kDa (pI 5.7-5.9), 19 kDa (pI 5.8-6.0), 16 kDa (pI 5.2-5.4) and 14 kDa (pI 5.1-5.3). For individual phosphoproteins, 32P incorporation varied over time and was also modulated by concentrations of Ca2+ and Mg2+ in the incubation medium. Treatment of the cells with forskolin led to statistically significant changes in the phosphorylation states of the 19.5 and 14 kDa proteins. Treatment of the cells with TPA also produced statistically significant changes in the 19.5 and 14 kDa proteins but, in addition, the 87 kDa, the 39 kDa and the 16 kDa phosphoproteins also exhibited significant changes. Alterations in the phosphorylation states of the 19.5 and the 14 kDa proteins were significantly correlated with alterations in beta-endorphin release from the cells. The primary finding of the present study was that activation of distinct second messenger systems can lead to alterations in the phosphorylation states of both shared and distinct phosphoproteins.
D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D004723:Endorphins / Q000378:metabolism; D004789:Enzyme Activation; D007700:Kinetics; D051379:Mice; D009363:Neoplasm Proteins / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D010911:Pituitary Neoplasms / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D001615:beta-Endorphin
D004723:Endorphins; D009363:Neoplasm Proteins; D010761:Phosphorus Radioisotopes; D005576:Colforsin; D001615:beta-Endorphin; D011493:Protein Kinase C; D000262:Adenylyl Cyclases; D013755:Tetradecanoylphorbol Acetate