Strader C D, Sibley D R, Lefkowitz R J
Life Sci. 1984 Oct 8;35(15):1601-10. doi: 10.1016/0024-3205(84)90359-x.
Chronic exposure of frog erythrocytes to beta-adrenergic agonists leads to desensitization of the responsiveness of adenylate cyclase to isoproterenol and is accompanied by "down-regulation", a decrease in the number of beta-adrenergic receptors on the cell surface. When frog erythrocyte plasma membranes are prepared by osmotic lysis of cells, the receptors lost from the cell surface during desensitization can be recovered in a "light membrane fraction", obtained by centrifuging the cell cytosol at 158,000 X g for 1 hr. These receptors are sequestered away from the plasma membrane fraction which contains the adenylate cyclase and the guanine nucleotide regulatory protein. If desensitized frog erythrocytes are disrupted by gentler freeze/thaw procedures, however, the sequestered beta-adrenergic receptors can be demonstrated to be physically associated with the plasma membrane. Typically, plasma membranes prepared in this fashion do not demonstrate a significant down regulation despite attenuation of isoproterenol-stimulated adenylate cyclase activity. Under these conditions, beta-adrenergic receptors from control and desensitized preparations co-migrate on sucrose density gradients in exactly the same place as the plasma membrane marker, adenylate cyclase. In contrast, when membranes from osmotically lysed desensitized cells are fractionated on sucrose gradients the down regulated receptors are sequestered in a light membrane fraction which barely enters the gradient and which is physically separated from adenylate cyclase activity. The data are consistent with a novel mechanism of receptor down-regulation which appears to involve the sequestration of the beta-adrenergic receptors away from the cell surface into a membrane compartment which remains physically associated with the plasma membrane.
将青蛙红细胞长期暴露于β-肾上腺素能激动剂会导致腺苷酸环化酶对异丙肾上腺素的反应性脱敏，并伴有“下调”，即细胞表面β-肾上腺素能受体数量减少。当通过细胞的渗透裂解制备青蛙红细胞质膜时，脱敏过程中从细胞表面丢失的受体可以在“轻膜组分”中回收，该组分是通过将细胞胞质溶胶在158,000×g下离心1小时获得的。这些受体被隔离在含有腺苷酸环化酶和鸟嘌呤核苷酸调节蛋白的质膜组分之外。然而，如果用更温和的冻融程序破坏脱敏的青蛙红细胞，可以证明隔离的β-肾上腺素能受体与质膜存在物理关联。通常，以这种方式制备的质膜尽管异丙肾上腺素刺激的腺苷酸环化酶活性减弱，但并未显示出明显的下调。在这些条件下，来自对照和脱敏制剂的β-肾上腺素能受体在蔗糖密度梯度上与质膜标记物腺苷酸环化酶在完全相同的位置共同迁移。相反，当对渗透裂解的脱敏细胞的膜进行蔗糖梯度分级分离时，下调的受体被隔离在一个几乎不进入梯度且与腺苷酸环化酶活性在物理上分离的轻膜组分中。这些数据与一种新的受体下调机制一致，该机制似乎涉及将β-肾上腺素能受体从细胞表面隔离到一个与质膜保持物理关联的膜区室中。
