EXPRESSION OF FUNCTIONAL GROWTH-HORMONE RECEPTOR IN A MOUSE L-CELL-LINE INFECTED WITH RECOMBINANT VACCINIA VIRUS

The growth hormone receptor is a member of a large family of receptors including the receptors for prolactin and interleukins. Upon binding to one molecule of growth hormone two growth hormone receptor polypept ides dimerize. We have expressed the rabbit growth hormone receptor DN A in transfected mouse L cells infected with polymerase T7-producing v accinia virus. The growth hormone receptor was synthesized as a 85-kDa protein and transported to the cell surface. Western blotting and met abolic labeling combined with immunoprecipitation using a rabbit antib ody probably directed against the cytosolic domain of the receptor sho wed that its expression was dependent on both transfection of the grow th hormone receptor DNA and vaccinia infection. Binding studies with I -125-labeled growth hormone demonstrated specific binding sites at the cell surface 20 h after transfection. Permeabilization with saponin s howed that the growth hormone receptor binding sites were almost exclu sively present at the cell surface with little intracellularly, Chemic al crosslinking of the I-125-growth hormone complex resulted in a 180- kDa complex which could specifically be immunoprecipitated with the an tiserum. Electron microscopic immunocytochemistry confirmed the presen ce of growth hormone receptor at the cell surface. Furthermore, specif ic growth hormone receptor antigen was also associated with intracellu lar membranes. These results thus show that this transient transfectio n system will be useful for cell biological studies of growth hormone receptor regulation. (C) 1994 Academic Press, Inc.