Fig 6.

Generation of a host cell line stably expressing levels of target receptor SSTR comparable to the levels of ecotropic receptor on NIH 3T3 cells. 293 cells were transfected with a cDNA-encoding human SSTR5 with an amino-terminal HA epitope tag (SSTR-HA), selected for G418 resistance, and then further selected for surface expression of the target receptor by three rounds of FACS experiments using anti-HA antibody, yielding a population of cells designated SSTR-HA/293. Left panel, aliquots of this cell line were incubated at 37°C with Sst-RBS pseudovirions or mock incubated (No virus), and then both samples were incubated with goat anti-SU antiserum followed by a mouse anti-goat FITC-conjugated secondary antibody plus propidium iodide and analyzed by flow cytometry. Values shown represent FITC intensities determined with live (propidium iodide-negative) cells. Right panel, aliquots of murine NIH 3T3 cells were incubated with MoMLV Env-pseudotyped virus or mock infected (No Virus), and virus binding was performed as described for the left panel. The histograms shown are representative of the results of three independent binding experiments.