Catecholamine-induced desensitization of adenylate cyclase coupled beta-adrenergic receptors in turkey erythrocytes: evidence for a two-step mechanism.
Preincubation of turkey erythrocytes with isoproterenol is associated with (1) 50-60% attenuation of agonist-stimulated adenylate cyclase activity, (2) altered mobility of the beta-adrenergic receptor on sodium dodecyl sulfate-polyacrylamide gels, and (3) increased phosphorylation of the beta-adrenergic receptor. Using a low-cross-linked polyacrylamide gel, the beta-adrenergic receptor protein from isoproterenol-desensitized cells, labeled with 32P or with the photoaffinity label 125I-(p-azidobenzyl)carazolol, can be resolved into a doublet (Mr congruent to 37,000 and Mr congruent to 41,000) as compared to a single Mr congruent to 37,000 beta-adrenergic receptor protein from control erythrocytes. The appearance of the doublet was dependent on the concentration of agonist used to desensitize the cells. Incubation of erythrocytes with dibutyryl-cAMP did not promote formation of the doublet but decreased agonist-stimulated adenylate cyclase activity 40-50%. Limited-digestion peptide maps of 32P-labeled beta-adrenergic receptors using papain revealed a unique phosphopeptide in the larger molecular weight band (Mr congruent to 41,000) of the doublet from the agonist-desensitized preparation that was absent in the peptide maps of the smaller band (Mr congruent to 37,000), as well as control or dibutyryl-cAMP-desensitized receptor. These data provide evidence that maximal agonist-induced desensitization of adenylate cyclase coupled beta-adrenergic receptors in turkey erythrocytes occurs by a two-step mechanism.
Stadel
J M
JM
Department of Molecular Pharmacology, Smith Kline and French Laboratories, Philadelphia, Pennsylvania 19101.
Rebar
R
R
Crooke
S T
ST
D000262:Adenylyl Cyclases / Q000097:blood*; D000345:Affinity Labels / Q000378:metabolism; D000818:Animals; D001386:Azides / Q000378:metabolism*; D003994:Bucladesine / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000187:drug effects / Q000378:metabolism*; D007545:Isoproterenol / Q000494:pharmacology*; D008970:Molecular Weight; D010206:Papain; D010449:Peptide Mapping; D011412:Propanolamines / Q000378:metabolism*; D011943:Receptors, Adrenergic, beta / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D014422:Turkeys
D000345:Affinity Labels; D001386:Azides; D011412:Propanolamines; D011943:Receptors, Adrenergic, beta; C054348:(4-azido-3-iodobenzyl)carazolol; D003994:Bucladesine; D010206:Papain; D000262:Adenylyl Cyclases; D007545:Isoproterenol