G proteins consist of three subunits: alpha, beta and gamma. Four beta subunits have been cloned: beta 1 and beta 4 (36 kDa), and beta 2 and beta 3 (35 kDa). We studied endogenous beta subunits in mouse NIH 3T3 fibroblasts stably expressing high levels of G protein alpha subunits after transfection with cDNAs encoding alpha i1, alpha i2, alpha i3 and alpha q. Immunoblots showed that NIH 3T3 cells express beta 36 and beta 35 subunits; in these cells, beta 35 subunits are four times more abundant than beta 36 subunits. We could detect beta 1 and beta 2 mRNA, but neither beta 3 nor beta 4 mRNA. We found that a stable increase in expression of wild-type alpha i1, alpha i2, alpha i3 or alpha q subunits is always accompanied by an increase in beta 1 and beta 2 mRNA and protein levels. There was no evidence of selectivity for an increase in beta 1 rather than beta 2 subunits depending on the type of alpha subunit overexpressed. However, constitutive activation or inactivation of alpha subunits induced specific changes in beta subunits. Expression of constitutively inactivated alpha i2 subunits was accompanied by an increase in mRNA and protein levels of both beta subunits. In contrast, cells expressing constitutively activated alpha i2 subunits did not show any change in the amount of beta proteins expressed in membranes, despite a significant increase in beta 1 and beta 2 mRNA. We conclude that stable changes in the levels of expression or degree of activation of G alpha subunits affect the level of expression, and possibly the turn-over, of beta subunits, without selectivity among beta 1 and beta 2 subunits.