The alpha 2A-adrenergic receptor is targeted directly to the basolateral membrane domain of Madin-Darby canine kidney cells independent of coupling to pertussis toxin-sensitive GTP-binding proteins.

The alpha 2-adrenergic receptor (alpha 2-AR) is a member of the seven transmembrane-spanning G-protein-coupled receptor superfamily. In the kidney, the alpha 2-AR is most abundant in the epithelial cells of the proximal tubule where it is important in enhancing Na+ reabsorption via the modulation of Na+/H+ exchange. Radioligand binding and physiological studies suggest that the alpha 2-AR residues primarily on the basolateral surface of these proximal tubule cells in vivo. To investigate the mechanisms underlying alpha 2-AR polarization in epithelial cells, we permanently expressed wild-type and an epitope-tagged version of the alpha 2A-AR in Madin-Darby canine kidney (MDCK) cells. Using a steady-state surface biotinylation assay, we observe that 80-90% of the alpha 2A-AR in MDCK cell clones is located on the basolateral membrane domain. Immunolocalization studies confirm the biotinylation results and demonstrate that the alpha 2A-AR is actually confined primarily to the lateral domain of the basolateral surface. Metabolic labeling experiments suggest that basolateral polarization of the alpha 2A-AR is achieved by direct targeting of the receptor to the basolateral domain. Targeting of the alpha 2A-AR to the basolateral surface is not perturbed by pertussis toxin-treatment of MDCK cells, suggesting that coupling of the alpha 2A-AR to GTP-binding proteins is not important for receptor polarization.