Cellular and species resistance to murine amphotropic, gibbon ape, and feline subgroup C leukemia viruses is strongly influenced by receptor expression levels and by receptor masking mechanisms

Abstract

Chinese hamster ovary (CHO) cells are resistant to infections by gibbon ape leukemia virus (GALV) and amphotropic murine leukemia virus (A-MLV) unless they are pretreated with tunicamycin, an inhibitor of N-linked glycosylation. These viruses use the related sodium-phosphate symporters Pit1 and Pit2, respectively, as receptors in nonhamster cells, and evidence has suggested that the corresponding transporters of CHO cells may be masked by tunicamycin-sensitive secreted inhibitors. Although the E36 line of Chinese hamster cells was reported to secrete the putative Pit2 inhibitor and to be sensitive to the inhibitory CHO factors, E36 cells are highly susceptible to both GALV and A-MLV in the absence of tunicamycin. Moreover, expression of E36 Pit2 in CHO cells conferred tunicamycin-independent susceptibilities to both viruses. Based on the latter results, it was suggested that E36 Pit2 must functionally differ from the endogenous Pit2 of CHO cells. To test these ideas, we analyzed the receptor properties of CHO Pit1 and Pit2 in CHO cells. Surprisingly, and counterintuitively, transfection of a CHO Pit2 expression vector into CHO cells conferred strong susceptibility to both GALV and A-MLV, and similar overexpression of CHO Pit1 conferred susceptibility to GALV. Thus, CHO Pit2 is a promiscuous functional receptor for both viruses, and CHO Pit1 is a functional receptor for GALV. Similarly, we found that the natural resistance of Mus dunni tail fibroblasts to subgroup C feline leukemia viruses (FeLV-C) was eliminated simply by overexpression of the endogenous FeLV-C receptor homologue. These results demonstrate a novel and simple method to unmask latent retroviral receptor activities that occur in some cells. Specifically, resistances to retroviruses that are caused by subthreshold levels of receptor expression or by stoichiometrically limited masking or interference mechanisms can be efficiently overcome simply by overexpressing the endogenous receptors in the same cells.

FIG. 1

Phosphate uptake of CHO cells expressing Pit1 and Pit2 proteins. CHO cells expressing the mouse ecotropic MLV receptor (CERD9 cells) (31) were transduced with ecotropic virus carrying genes that encode for either rat Pit2 (CE/rPit2) (17), CHO Pit2 (CE/cPit2), human-CHO Pit1 (CE/hcPit1) and human Pit1 (CE/hPit1). CHO/EAR cells are CHO cells expressing E36 Pit2 and were generated by Wilson et al. (35). Phosphate transport was measured using the procedure outlined by Olah et al. (21). The phosphate uptake values are averages of four different replicates in the same experiment. The standard deviations (error bars) are shown.

FIG. 2

Comparison of the amino acid sequences of hFLVCR and mdFLVCR. Dots, identical amino acids; dashes, spaces introduced for alignment. ∗, N-linked glycosylation site for hFLVCR. Potential membrane-spanning segments are indicated by a line over the sequence, and the presumptive ECLs are indicated.