Analysis by cell hybridization of mechanisms that regulate beta-adrenergic responses in reticulocytes and in differentiating erythroid cells.

In intact reticulocytes, but not in fragmented membranes, the loss of adenylate cyclase activity during cell maturation followed a biphasic time course. A rapid phase (t1/2 approximately 2 h) during which the initial activity was reduced by 40-50% was followed by a slow phase with t1/2 close to 3 days. The fast decay seemed to occur on the adenylate cyclase level since (-)isoprenaline- or forskolin-stimulated activities behaved similarly and bacterial toxin-monitored Gs and Gi proteins remained stable. The mechanism of the initial decrease in hormonal responsiveness was further analysed in hybrid cells prepared by fusing reticulocytes with Friend erythroleukemia (MEL) cells. The hybrids contained reticulocyte-derived beta-adrenoceptors and MEL cell-derived adenylate cyclase and G proteins. Fusion of reticulocytes to native MEL cells caused adenylate cyclase activity to drop by 30% at 2 h and 45% at 18 h after fusion. By contrast, hybrids prepared after dimethylsulfoxide-induced differentiation of MEL cells showed stable or increasing rates of receptor-coupled cAMP formation between 2 and 18 h after fusion, concomitant with the enhanced activity of the Gs protein in these cells. A cyclase-stimulating factor present in the cytosol of MEL cells and of reticulocytes appeared not to be involved in short-term regulation of hormonal responsiveness. We conclude that the strength of beta-adrenergic responses in erythroid progenitor cells is primarily regulated by modulating G protein-mediated receptor cyclase coupling while reticulocytes, during early maturation, seem to rely on direct inactivation of adenylate cyclase, probably via a cytosolic proteolytic pathway.