Agonist and cyclic AMP-mediated regulation of beta 1-adrenergic receptor mRNA and gene transcription in rat C6 glioma cells.
Exposure of rat C6 glioma cells to either agonists or agents that increase cyclic AMP levels leads to down-regulation of beta 1-adrenergic receptors (beta 1 AR) as measured by loss of radioligand binding sites. The present study examines the influence of isoproterenol and forskolin treatment on levels of beta 1 AR mRNA, mRNA stability, and gene transcription rate. Isoproterenol treatment of C6 cells altered beta 1 AR mRNA levels in a biphasic manner; i.e., short-term exposure (30-60 min) increased by 50%, whereas longer exposure (2-6 h) decreased by 50% the levels of beta 1 AR mRNA. The extent of both the up- and down-regulation was dependent on agonist concentration. Similar regulation of beta 1 AR mRNA was observed in forskolin-treated cells. Pretreatment of the cells with Pseudomonas exotoxin A, a potent inhibitor of protein synthesis, completely blocked isoproterenol- and forskolin-mediated down-regulation of beta 1 AR mRNA, and thereby potentiated the increase in receptor mRNA up to fourfold over the 6-h time course. The mechanisms underlying beta 1 AR mRNA down-regulation were examined. The half-life of beta 1 AR mRNA was slightly increased (from 61 to 77 min) after a 2-h exposure of the cells to either isoproterenol or forskolin. Nuclear run-on analysis demonstrated that the rate of beta 1 AR gene transcription was increased after isoproterenol incubation for 60 min, but then decreased after 90-240 min, consistent with the time course for up- and down-regulation of beta 1 AR mRNA. Isoproterenol treatment (120 min) also decreased the level of beta 1 AR nascent transcripts, purified by affinity chromatography of RNA isolated from 4-thiouridine-pulsed cells. The results demonstrate that beta 1 AR mRNA has a relatively short half-life and that agonist regulation of beta 1 AR mRNA is mediated by activation of the cyclic AMP system. Moreover, the results indicate that agonist regulation of beta 1 AR mRNA occurs at the level of beta 1 AR gene transcription, not mRNA stability. Finally, the observed requirement for protein synthesis indicates that beta 1 AR mRNA down-regulation may be mediated by the induction of a repressor of the beta 1 AR gene.
Hosoda
K
K
Department of Psychiatry, Yale University School of Medicine, New Haven, Connecticut.
Feussner
G K
GK
Rydelek-Fitzgerald
L
L
Fishman
P H
PH
Duman
R S
RS
D058665:Adrenergic beta-1 Receptor Agonists; D000818:Animals; D005576:Colforsin / Q000494:pharmacology*; D000242:Cyclic AMP / Q000819:agonists* / Q000494:pharmacology* / Q000502:physiology; D015536:Down-Regulation; D004731:Endotoxins / Q000494:pharmacology; D005910:Glioma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology*; D007545:Isoproterenol / Q000494:pharmacology*; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D018342:Receptors, Adrenergic, beta-1 / Q000235:genetics* / Q000502:physiology; D013997:Time Factors; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D058665:Adrenergic beta-1 Receptor Agonists; D004731:Endotoxins; D012333:RNA, Messenger; D018342:Receptors, Adrenergic, beta-1; D005576:Colforsin; D000242:Cyclic AMP; D007545:Isoproterenol