A retroviral vector was used to express various amounts of the receptor (ecoR) for ecotropic host range murine retroviruses on naturally barren hamster, mink, and human cells. These cells and murine cells were then incubated for 2 h with dilutions of a helper-free ecotropic retrovirus that encodes human growth hormone, and the number of infected cells was later determined by growth hormone-specific immunofluorescence. For all cells under the conditions of these studies, virus adsorption was the limiting step of infection and the cellular capacities for infection were unsaturated either at cell surfaces or at intracellular sites. Thus, infections occurred at low multiplicities of infection per cell and were directly proportional to virus and cell concentrations, and only a small percentage (ca. 5%) of the infectious virions became adsorbed from the medium during the 2-h incubations. Although increasing the adsorption by raising virus or cell concentrations results in more infections in the cultures, increasing adsorption by raising the number of ecoR above a low threshold had no effect on infections. Thus, cells with a low number of ecoR were infected as efficiently as highly adsorbing cells that contained many times more ecoR. To reconcile these results, we conclude that only a small, set number of cell surface ecoR can be functional for infection and that all excess ecoR can only bind virus into an unsalvageable pool. Therefore, retroviral receptors on single cells are functionally diverse. Our results suggest that activity of ecoR in infection requires a limiting second cellular component. * Corresponding author. 25) and for gibbon ape leukemia virus (39) were isolated. Although these receptors appear to be unrelated, they both may be glycoproteins with multiple membrane-spanning regions (1, 39). The ecotropic receptor (ecoR) occurs on cells of mice and rats but not on cells of other species. Genetic analysis has suggested that ecoR may be essential for the viability of cultured murine fibroblasts (16). In this study, we have developed efficient methods to express ecoR cDNA in nonmurine cells. We used these cells to study the role of ecoR in virus binding and infection. Our results suggest that a cellular component in addition to ecoR is required for infection. MATERIALS AND METHODS Cells and viruses. Swiss mouse NIH 3T3 fibroblasts, mink lung CCL64 fibroblasts, Chinese hamster ovary cells (CHO), and human osteogenic sarcoma cells (Ostk-) were from American Type Culture Collection (Rockville, Md.). Ii-2 ecotropic packaging cells (34) and PA-12 amphotropic packaging cells (38) were from R. C. Mulligan (Massachusetts Institute of Technology, Cambridge) and A. D. Miller (Fred Hutchinson Cancer Center, Seattle, Wash.), respectively. CHO cells were grown in alpha minimum essential medium supplemented with 10% fetal bovine serum. Other cells were maintained in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum. Helper-free ecotropic host range viruses from ti-2 cells that encoded the neomycin...

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