The Short and Long Forms of the α Subunit of the Stimulatory Guanine‐Nucleotide‐Binding Protein are Unequally Redistributed During (–)‐Isoproterenol‐Mediated Desensitization of Intact S49 Lymphoma Cells

Abstract

We report here that desensitization of the β‐adrenergic receptor‐triggered transmembrane signalling in S49 wild‐type lymphoma cells, induced by (–)‐isoproterenol (1 μM), results in unequal intracellular redistribution of the splicing variants of the α subunit of the stimulatory guanine‐nucleotide‐binding regulatory (Gsα protein (Gsα‐short and Gsα‐long) and alters the functional characteristics of the membrane‐associated signal transduction complex. We found that two cellular pools of membranes, light‐density membranes and plasma membranes prepared by sucrose‐density‐gradient centrifugation of cell homogenates differed in their content of Gsα splicing subforms and, moreover, that prolonged activation of the β‐adrenergic pathway induced intermembrane redistribution of the splicing variants of Gsα. Short (10 min) as well as prolonged (1 h) (–)‐isoproterenol treatment of the cells shifted Gsα‐short from light‐density membranes to plasma membranes and increased the total amount of light‐density membrane‐bound Gsα‐long; in parallel, the maximal (–)‐isoproterenol‐stimulated or AlF−4‐stimulated adenylyl cyclase activities measured in the plasma membrane pools prepared from treated cells decreased. The functional characteristics of the membrane‐bound Gsα pools were examined by a cyc−‐reconstitutive adenylyl cyclase assay where extracts of the plasma membrane and light‐density‐membrane pools, respectively, were mixed with plasma membranes derived from the mutant S49 cell line, cyc−, lacking Gsα. The maximal cyc−‐reconstitutive activities of the extracts prepared from light‐density membranes of short‐term as well as long‐term desensitized cells increased compared to control cells. These findings may indicate differences in the functioning of the splicing variants of Gsα.