Adenylyl cyclase inhibition and altered G protein subunit expression and ADP-ribosylation patterns in tissues and cells from G_i2α-/- mice

The inhibition of α_i2-/- mouse cardiac isoproterenol-stimulated adenylyl cyclase (AC; EC 4.6.1.1) activity by carbachol and that of α_i2-/- adipocyte AC by phenylisopropyladenosine (PIA), prostaglandin K:, and nicotinic acid were partially, but not completely, inhibited. While the inhibition of cardiac AC was affected in all α_i2-/-animals tested, only 50% of the α_i2-/- animals showed an impaired inhibition of adipocyte AC, indicative of a partial penetrance of this phenotype. In agreement with previous results, the data show that G_i2 mediates hormonal inhibition of AC and that G_i3 and/or G_i1, is capable of doing the same but with a lower efficacy. Disruption of the α_i2 gene affected about equally the actions of all the receptors studied, indicating that none of them exhibits a striking specificity for one type of G_i over another and that receptors are likely to be selective rather than specific in their interaction with functionally homologous G proteins (e.g., G_i1, G_i2, G_i3). Western analysis of G protein subunit levels in simian virus 40-transformed primary embryonic fibroblasts from α_i2+/+ and α_i2-/- animals showed that α_i2 accounts for about 50% of the immunopositive G protein α subunits and that loss of the α_i2 is accompanied by a parallel reduction in Gβ₃₅ and Gβ₃₆ subunits and by a 30-50% increase in α_i3. This suggests that Gβ_γ levels may be regulated passively through differential rates of turnover in their free vs. trimeric states. The existence of compensatory increase(s) in α_i, subunit expression raises the possibility that the lack of effect of a missing α_i2 on AC inhibition in adipocytes of some α_i2-/- animals may be the reflection of a more pronounced compensatory expression of α_i3 and/or α_i1.