title: Modulation of adenylylcyclase by protein kinase C in human neurotumor SK-N-MC cells: evidence that the alpha isozyme mediates both potentiation and desensitization.
abstract: Exposure of human SK-N-MC neurotumor cells to 4 beta-phorbol 12-myristate 13-acetate (PMA) increased isoproterenol stimulation of cyclic AMP levels by severalfold. This potentiation was blocked by inhibitors of protein kinase C (PKC) and did not occur in cells in which PKC had been down-regulated. PMA treatment also enhanced the stimulation by dopamine, cholera toxin, and forskolin. Thus, the effect of PMA on the adenylylcyclase system was postreceptor and involved either the guanine nucleotide binding regulatory (G) proteins or the cyclase itself. As PMA treatment did not impair the inhibition of isoproterenol stimulation by neuropeptide Y, an involvement of the inhibitory G protein Gi was unlikely. Cholate extracts of membranes from control and PMA-treated cells were equally effective in the reconstitution of adenylylcyclase activity in S49 cyc- membranes, which lack the stimulatory G protein subunit Gs alpha; thus, Gs did not appear to be the target of PMA action. Membranes from PMA-treated cells exhibited increased adenylylcyclase activity to all stimulators including Mn2+ and Mn2+ plus forskolin. In addition, activity was increased when control membranes were incubated with ATP and purified PKC from rat brain. This is consistent with a direct effect of PKC on the adenylylcyclase catalyst in SK-N-MC cells. PMA treatment also resulted in a shift to less sensitivity in the K(act) for isoproterenol but not for dopamine or CGP-12177 (a beta 3-adrenergic agonist) stimulation. Thus, the beta 1 but not the D1 or beta 3 receptors were being desensitized by PKC activation. Analysis of SK-N-MC cells by western blotting with antibodies against different PKC isozymes revealed that both the alpha and zeta isozymes were present in these cells. Whereas PKC-alpha was activated and translocated from cytosol to membrane by phorbol esters, the zeta isozyme was not. Thus, PKC-alpha, which has been implicated in desensitization in other cell lines, also appears to potentiate adenylylcyclase activity.
authors.affiliation: Laboratory of Molecular and Cellular Neurobiology, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland.
authors.forename: X M
authors.initials: XM
authors.lastname: Zhou
authors.forename: P
authors.initials: P
authors.lastname: Curran
authors.forename: J
authors.initials: J
authors.lastname: Baumgold
authors.forename: P H
authors.initials: PH
authors.lastname: Fishman
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism*; D000319:Adrenergic beta-Antagonists / Q000494:pharmacology; D000470:Alkaloids / Q000494:pharmacology; D017607:Aluminum Compounds / Q000494:pharmacology; D000818:Animals; D001921:Brain / Q000201:enzymology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002772:Cholera Toxin / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism*; D004298:Dopamine / Q000494:pharmacology; D004357:Drug Synergism; D005459:Fluorides / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D007545:Isoproterenol / Q000494:pharmacology*; D007700:Kinetics; D008345:Manganese / Q000494:pharmacology; D009447:Neuroblastoma; D009478:Neuropeptide Y / Q000494:pharmacology; D011412:Propanolamines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D013110:Sphingosine / Q000494:pharmacology; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
chemical_list: D000319:Adrenergic beta-Antagonists; D000470:Alkaloids; D017607:Aluminum Compounds; D007527:Isoenzymes; D009478:Neuropeptide Y; D011412:Propanolamines; D005576:Colforsin; C050992:tetrafluoroaluminate; D008345:Manganese; D002772:Cholera Toxin; D000242:Cyclic AMP; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases; D019311:Staurosporine; D007545:Isoproterenol; D013110:Sphingosine; D013755:Tetradecanoylphorbol Acetate; D005459:Fluorides; C033394:CGP 12177; D004298:Dopamine