Regulation of alpha 1B-adrenergic receptor half-life: protein synthesis dependence and effect of norepinephrine.
Agonist-mediated down-regulation of alpha 1B-adrenergic receptors (AAR) may involve a decrease in synthesis, an increase in degradation, or a combination of the two mechanisms. Norepinephrine (NE) causes downregulation of AAR in rabbit aortic smooth muscle cells (RbSMC). To study the role of receptor degradation in this phenomenon, we studied the half-life (t1/2) of the AAR under basal conditions and during exposure to NE. We first determined the disappearance rate of AAR in the presence of cycloheximide, a commonly used method for measuring receptor t1/2. By this approach, the basal t1/2 was surprisingly long, 172 +/- 20 h. In contrast, the t1/2 in NE-treated cells was 11.8 +/- 0.3 h, suggesting that either NE decreased the t1/2 of AAR or cycloheximide increased the basal t1/2. To distinguish these possibilities, basal receptor t1/2 was determined by a second, independent method based on the repopulation of AAR after irreversible alkylation with chloroethylclonidine. This approach indicated a basal t1/2 (7.4 +/- 0.2 h) that was similar to the t1/2 with NE but more than 20-fold shorter than the t1/2 with cycloheximide. We conclude that in RbSMC NE-induced downregulation of AAR occurs without a decrease in receptor t1/2. The unexpected finding that cycloheximide markedly increases the basal t1/2 of the AAR further indicates that the t1/2 of the AAR is regulated, at least in part, by a short-lived protein that may regulate and/or mediate receptor degradation.
Izzo
N J
NJ
Department of Medicine, Brigham and Women's Hospital, Boston, Massachusetts 02115.
Colucci
W S
WS
D000477:Alkylating Agents / Q000494:pharmacology; D000818:Animals; D001011:Aorta / Q000166:cytology / Q000378:metabolism*; D002478:Cells, Cultured; D003000:Clonidine / Q000031:analogs & derivatives / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D015536:Down-Regulation; D006207:Half-Life; D009124:Muscle Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D009638:Norepinephrine / Q000494:pharmacology*; D011817:Rabbits; D011943:Receptors, Adrenergic, beta / Q000187:drug effects / Q000378:metabolism*
D000477:Alkylating Agents; D009124:Muscle Proteins; D011943:Receptors, Adrenergic, beta; C051731:chlorethylclonidine; D003513:Cycloheximide; D003000:Clonidine; D009638:Norepinephrine