Beta(2)-adrenoceptors increase translocation of GLUT4 via GPCR kinase sites in the receptor C-terminal tail

Background and purpose: beta-Adrenoceptor (beta-AR) stimulation induces glucose uptake in several insulin-sensitive tissues, by poorly understood mechanisms. Experimental approach: We used a model system in CHO-K1 (Chinese Hamster ovary K1) cells expressing the human beta(2) -AR and glucose transporter 4 (GLUT4) to investigate the signalling mechanisms involved. Key results: In CHO-K1 cells there was no response to beta-AR agonists. The introduction of beta(2) -ARs and GLUT4 into these cells caused increased glucose uptake in response to beta-AR agonists. GLUT4 translocation occurred in response to insulin and beta(2) -AR stimulation, although the key insulin signalling intermediate Akt was not phosphorylated in response to beta(2) -AR stimulation. Truncation of the C-terminus of the beta(2) -AR at position 349 to remove known phosphorylation sites for G protein-coupled receptor kinases (GRKs) or at position 344 to remove an additional protein kinase A (PKA) site together with the GRK phosphorylation sites did not significantly affect cyclic AMP accumulation but decreased beta(2) -AR stimulated glucose uptake. Furthermore, inhibition of GRK by transfection of betaARKct inhibited beta(2) -AR-mediated glucose uptake and GLUT4 translocation, and over-expression of a kinase-dead GRK2 mutant (GRK2 K220R) also inhibited GLUT4 translocation. Introducing beta(2) -AR lacking phosphorylation sites for GRK or PKA demonstrated that the GRK sites, but not the PKA sites, were necessary for GLUT4 translocation. Conclusions and implications: Glucose uptake in response to activation of beta(2) -ARs involves translocation of GLUT4 in this model system. The mechanism is dependent on the C-terminus of the beta(2) -AR, requires GRK phosphorylation sites, and involves a signalling pathway distinct from that stimulated by insulin.