Divalent cation-induced desensitization of glucagon-stimulable adenylyl cyclase in rat liver plasma membrane. GTP-dependent stimulation by glucagon

Treatment of rat liver plasma membrane with Mg ion in the presence of 1.0 mM ATP leads to a concentration-dependent loss of glucagon stimulability of adenylyl cyclase without loss of guanine nucleotide- or fluoride- stimulated activity; apparent K(a) for Mg ion: 22 mM. Lowering of ATP to 0.1 mM or substituting with adenyl-5'yl imidodiphosphate abolished this effect. Addition of glucagon during treatment lowered the apparent K(a) for Mg ion to 4.0 mM. Desensitization stimulated by glucagon was dependent upon co-addition of GTP, that by Mg ion alone was not. Mn ion could substitute for Mg ion in the desensitization process with apparent K(a) values of 0.4 and 0.1 mM in the absence and presence of glucagon, respectively. Desensitization in the presence of glucagon, was time-dependent, also required ATP (apparent K(a) of 0.4 mM), and exhibited an apparent K(a) for glucagon of 1 nM, which is indistinguishable from the apparent K(a) with which glucagon stimulates adenylyl cyclase. Full desensitization obtained in the presence of glucagon led to the reappearance of the characterisitic lag in the time course of activation of liver adenylyl cyclase by guanine-5'-yl imidodiphosphate. This lag is otherwise seen only in the absence of glucagon. Loss of glucagon stimulability was resistant to washing and persisted upon lowering of ATP in the assay to 0.1 mM. The apparent K(a) for glucagon remained unchanged upon partial (40%) desensitization. After desensitization, there was no significant change in NaF-stimulated activity. Although close to 100% of glucagon-stimulated activity was observed upon desensitization with 0.1 μM glucagon and 5 mM MgCl₂, this was accompanied by only a 25% decrease in the number of ¹²⁵I-glucagon binding sites. Upon desensitization with 100 mM MgCl₂, no loss of glucagon binding sites was observable. Thus, it appears that liver plasma membranes contain a divalent cation-dependent process that leads to uncoupling of glucagon receptors from adenylyl cyclase in an ATP-dependent manner and that glucagon receptors themselves are capable, upon occupation, of stimulating this uncoupling process in a GTP-dependent manner. Furthermore, the 40- to 50-fold difference between the apparent K(a) values for Mg ion and those for Mn ion in promoting desensitization suggests that this effect of divalent cations is mediated by a site siye allosteric to that (or those) with which ATP interacts in this system.