title: Cross-regulation between G-protein-mediated pathways. Acute activation of the inhibitory pathway of adenylylcyclase reduces beta 2-adrenergic receptor phosphorylation and increases beta-adrenergic responsiveness.
abstract: Cross-regulation from the stimulatory to the inhibitory adenylylcyclase pathways has been described (Hadcock, J. R., Ros, M., Watkins, D. C., and Malbon, C. C. (1990) J. Biol. Chem. 265, 14784-14790). More recently, persistent activation (48 h) of the inhibitory adenylylcyclase pathway has been shown to cross-regulate the stimulatory pathway (i) enhancing the maximal response of beta-adrenergic agonits, (ii) increasing the expression of beta-adrenergic receptor, and (iii) reducing the ED50 for the isoproterenol-stimulated response by 50-fold (Hadcock, J. R., Port, J. D., and Malbon, C. C. (1991) J. Biol. Chem. 266, 11915-11922). Here, we report that short term activation (60 min) of the inhibitory adenylylcyclase pathway of hamster smooth muscle DDT1MF-2 cells with the A1-adenosine receptor agonist N6-phenylisopropyladenosine (PIA) likewise enhances the stimulatory adenylylcyclase response to the beta-adrenergic agonist isoproterenol. The PIA effect was exerted at the level of the receptor, i.e., the beta-adrenergic receptor-mediated response was enhanced, whereas the guanosine 5'-O-(thiotriphosphate)- and forskolin-stimulated adenylylcyclase activities were largely unaffected. In contrast to longer term persistent activation of the inhibitory pathway, receptor number and affinity for 125I-labeled cyanopindolol were unaffected. Metabolic labeling of cells with [32P]orthophosphate and immuneprecipitation of beta-adrenergic receptors detected phosphorylation of the receptor in unstimulated cells and marked phosphorylation in cells challenged with epinephrine. When cells were challenged short term with PIA, the basal state of beta-adrenergic receptor phosphorylation was reduced by 75%. Treating cells with PIA in combination with the cAMP analog 8-(4-chlorophenylthio)adenosine cyclic AMP attenuated the enhanced receptor-mediated adenylylcyclase response observed in cells treated with PIA alone. These data suggest that short term cross-regulation from the inhibitory to stimulatory adenylylcyclase pathways results in the following: (i) decreased intracellular cAMP levels and protein kinase A activity, (ii) reduced phosphorylation of the beta 2-adrenergic receptor in the "basal" (i.e. unstimulated) state, and (iii) enhanced receptor-mediated activation of Gs.
authors.affiliation: Department of Pharmacology, School of Medicine, State University of New York, Stony Brook 11794-8651.
authors.forename: J D
authors.initials: JD
authors.lastname: Port
authors.forename: J R
authors.initials: JR
authors.lastname: Hadcock
authors.forename: C C
authors.initials: CC
authors.lastname: Malbon
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D006224:Cricetinae; D000242:Cyclic AMP / Q000494:pharmacology; D004789:Enzyme Activation; D004837:Epinephrine / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D011869:Radioligand Assay; D011943:Receptors, Adrenergic, beta / Q000378:metabolism*
chemical_list: D011943:Receptors, Adrenergic, beta; D005576:Colforsin; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D000242:Cyclic AMP; D011494:Protein Kinases; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases; D004837:Epinephrine