Desensitization of the mammalian beta-adrenergic receptor: analysis of receptor redistribution on nonlinear sucrose gradients.
The distribution of beta-adrenergic receptors in lysates from several mammalian cell lines was analyzed on nonlinear sucrose density gradients before and after desensitization by isoproterenol. On the nonlinear gradients, the receptors in lysates from untreated HeLa, A431, S49 cyc- and C6 cells were well resolved into light and heavy density membrane fractions. In contrast, with the former three cell lines, there was very poor or no separation of the two peaks of receptors on linear sucrose gradients. With C6 cells, resolution was better on the nonlinear than on the linear gradient. In all cases, successful separation of the two density fractions of the receptor was achieved only when cells had been treated with concanavalin A prior to lysis. Adenylate cyclase activity cosedimented with the heavy membrane fraction of the receptor, and no activity was detected with the light fraction. After desensitization of adenylate cyclase by isoproterenol, there was a redistribution of the receptors to the light density fraction. This shift of receptors, but not desensitization, was prevented when cells were pretreated at 37 degrees C with concanavalin A prior to exposure to isoproterenol. Thus, sequestration of beta-adrenergic receptors away from the plasma membrane and adenylate cyclase to a lighter density membrane fraction appears to accompany, but may not be a prerequisite for desensitization in mammalian cells. This receptor redistribution, however, can be readily detected on nonlinear sucrose gradients.
D000262:Adenylyl Cyclases / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D003208:Concanavalin A / Q000494:pharmacology; D005910:Glioma / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D011943:Receptors, Adrenergic, beta / Q000378:metabolism*; D013395:Sucrose
D011943:Receptors, Adrenergic, beta; D003208:Concanavalin A; D013395:Sucrose; D000262:Adenylyl Cyclases