Protein kinase C activation mimics but does not mediate thyrotropin-induced desensitization of adenylyl cyclase in cultured dog thyroid cells.
The mechanism and site(s) of the defect responsible for desensitization to hormone stimulation of adenylyl cyclase (AC) vary with cell type. Plasma membrane preparations were assayed after treatment of primary cultured dog thyroid cells to determine the role of the TSH receptor, stimulatory and inhibitory guanine nucleotide binding proteins (Gs and Gi), and catalytic unit in AC desensitization. Exposure of cells to TSH or the phorbol ester, 12-O-tetradecanoyl-phorbol-13-acetate (TPA), caused time dependent decreases in TSH-stimulated AC and [125I]TSH binding with approximately 50% decreases seen after 18 h; Bt2cAMP was unable to reproduce the TSH effect. Whereas TSH treatment caused concomitant decreases (approximately 25%) in both cyclase activity and [125I]TSH binding after 2 h, TPA treatment decreased AC activity after 6 h and binding only after 18 h. The protein kinase C inhibitor, H-7, prevented TPA-induced but not TSH-induced effects on AC and hormone binding. Membrane AC activation by cholera toxin or forskolin was not altered by 18 h pretreatment of cells with TSH or TPA, indicating that these agents had no apparent effect on intrinsic functionality of either Gs or the catalytic unit. TSH or TPA pretreatment of cells reduced subsequent toxin-mediated AD[32P]-ribosylation of Gs and Gi in isolated membranes. However, the TSH- and TPA-induced decreases in AD[32P]-ribosylation and desensitization do not appear to be due to endogenous ribosylation of G proteins, since treatment of cells with pertussis toxin, for example, to endogenously ribosylate Gi, both increased TSH-stimulated AC activity and failed to affect the ability of TSH or TPA to desensitize. Thus, in this system, although specific hormone-induced AC desensitization and receptor down-regulation conform to several aspects of classic homologous processes, similar effects are also induced by a nonreceptor (phorbol ester) pathway; desensitization, however, can precede down-regulation, possibly due to receptor-Gs uncoupling.
Deery
W J
WJ
Department of Medicine, Baylor College of Medicine, Houston, Texas 77030.
Rani
C S
CS
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D037361:Adenylate Cyclase Toxin; D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D002772:Cholera Toxin / Q000494:pharmacology; D004285:Dogs; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism; D007546:Isoquinolines / Q000494:pharmacology; D007700:Kinetics; D037342:Pertussis Toxin; D010879:Piperazines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013961:Thyroid Gland / Q000166:cytology / Q000201:enzymology*; D013972:Thyrotropin / Q000494:pharmacology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D037361:Adenylate Cyclase Toxin; D007546:Isoquinolines; D010879:Piperazines; D010566:Virulence Factors, Bordetella; D000246:Adenosine Diphosphate Ribose; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D013972:Thyrotropin; D002772:Cholera Toxin; D037342:Pertussis Toxin; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases; D013755:Tetradecanoylphorbol Acetate