G-Protein mRNA levels during adipocyte differentiation

G-protein-mediated transmembrane signaling in 3T3-L1 cells is modulated by differentiation. The regulation of G-protein expression in differentiating 3T3-L1 cells was probed at the level of mRNA by DNA-excess solution hybridization. Pertussis toxin-catalyzed ADP-ribosylation of G-protein α-subunits increased as fibroblasts differentiate to adipocytes. Steady-state levels of mRNA for G_iα2 and G_oα, in contrast, declined sharply. Immunoblotting with antipeptide antibodies specific for G_iα2, too, revealed a decline in the steady-state expression of this pertussis toxin substrate. ADP-ribosylation of G_sα by cholera toxin was less in the adipocyte than fibroblast. Analysis by immunoblotting revealed only a modest decline in G_sα. Analysis of mRNA levels also demonstrated a decline for G_sα. mRNA levels for the G_β-subunits rose initially (25%) on day 1, declined from day 1 to day 3, and remained 25% lower in adipocytes than in fibroblasts. In 3T3-L1 adipocytes the molar amounts of subunit mRNAs were: 60.6 (G_sα); 2.1 (G_iα2); and 1.5 (G_oα) amol/μg total cellular RNA. In rat fat cells these mRNA levels were 19.4 (G_sα); 7.0 (G_iα2); and 2.3 (G_oα). These data demonstrate that for G_iα2 and G_oα alike mRNA and protein expression decrease, not increase, in differentiation. A substrate for pertussis toxin other than G_iα2 and G_oα appears to be responsible for the increase in toxin-catalyzed labeling that accompanies differentiation of 3T3-L1 cells.