Pertussis toxin treatment in vivo is associated with a decline in G-protein β-subunits

The effects of pertussis toxin on the steady-state levels of G-protein α- and β-subunits were investigated both in vitro and in vivo. The steady-state level G(oα), a major substrate for pertussis toxin-catalyzed ADP-ribosylation, was unaltered by pertussis toxin treatment for periods up to 100 h for 3T3-L1 cells in culture or up to 3 days in vivo. In 3T3-L1 cells pertussis toxin treatment did not alter levels of G(sα)-subunits; in S49 cells the level of G(sα)-subunits declined moderately following by pertussis toxin treatment. The steady-state levels of G(β)-subunits, in contrast, were found to decline to less than 50% of the normal cellular complement following pertussis toxin treatment in vitro and in vivo. Inhibitory control of adenylate cyclase, pertussis toxin-catalyzed ADP-ribosylation of G(iα) and G(oα), and the GTP-dependent shift in agonist-specific binding to β-adrenergic receptors were attenuated or abolished within 5 h of pertussis toxin treatment, representing 'early' effects of the toxin. Stimulatory regulation of adenylate cyclase, in contrast, displayed a progressive enhancement that was first observed 4 h after pertussis toxin treatment, increasing thereafter up until 100 h, the last time point measured. This progressive enhancement of the stimulatory pathway of adenylate cyclase was not manifest at the level of stimulatory receptors, since the K(d) and B(max) for one such receptor, the β-adrenergic receptor, were shown to be unaltered in toxin-treated cells. Furthermore, the potentiation of stimulation of adenylate cyclase was observed in cells stimulated by the β-adrenergic agonist isoproterenol and PGE₁ alike. The progressive enhancement of the stimulatory pathway correlated best with the decline in G(β)-subunit levels that occurs following pertussis intoxication. The changes in both of these parameters occur 'late' (12-48 h), as compared to the early events that occur within 5 h. Pertussis toxin action appears to be composed of two, temporally distinct, groups of effects. Pertussis toxin-catalyzed ADP-ribosylation of G(α)-subunits, attenuation of the inhibitory regulation of adenylate cyclase, and attenuation of the ability of GTP to induce an agonist-specific shift in receptor affinity are members of the early group of effects. The second group of late effects includes the decline in G(β)-subunit levels and the progressive enhancement of the stimulatory pathway of adenylate cyclase. This enhanced stimulatory control at these later times cannot be explained by the attenuation of the inhibitory pathway occurring early, but rather appears as G(β)-subunit levels decline.