Resolution and identification of Gq/G11alpha and Gialpha/Goalpha proteins in brown adipose tissue: effect of cold acclimation

Abstract

Levels of guanine nucleotide-binding proteins G(q)alpha and G(11)alpha, which produce receptor regulation of phospholipase C, were measured immunologically in purified plasma membrane fractions of hamster brown adipose tissue (BAT). This was achieved by immunoblotting with antisera (CQ series) that identify these two polypeptides equally, following separation of the plasma membranes using SDS-PAGE in the presence of 6 M urea, i.e. conditions that can resolve G(q)alpha and G(11)alpha. The ratio of levels of G(q)alpha to G(11)alpha was 1:1. A similar approach was used for resolution and identification of G(o)1alpha and G(o)2alpha, the latter representing the prevailing form of G(o)alpha proteins in this tissue. Although clearly recognized in brain microsomes, which were used as positive controls, no detected levels of G(o)*alpha protein were noted. Using specific anti-peptide antibodies directed against the carboxy-terminal decapeptide of G(i)3alpha, this G protein was also found to be expressed in BAT tissue. Cold acclimation resulted in reduction of the plasma membrane levels of all these Galpha proteins.