Primate Gammaretroviruses Require an Ancillary Factor Not Required for Murine Gammaretroviruses To Infect BHK Cells

Abstract

BHK cells remain resistant to xenotropic murine retrovirus-related virus (XMRV) or gibbon ape leukemia virus (GALV) infection, even when their respective receptors, Xpr1 or PiT1, are expressed. We set out to determine the stage at which viral infection is blocked and whether this block is mediated by a dominantnegative factor or the absence of a requisite ancillary factor. BHK cells bind neither XMRV nor GALV envelope proteins. BHK cells expressing the appropriate receptors bind XMRV or GALV envelope proteins. BHK cells can be infected by NZB-XMV(New Zealand Black mouse xenotropic murine virus)-enveloped vectors, expressing an envelope derived from a xenotropic retrovirus that, like XMRV, employs Xpr1 as a receptor, and also by vectors bearing the envelope of 10A1 murine leukemia virus (MLV), a murine retrovirus that can use PiT1 as a receptor. The retroviral vectors used in these analyses differ solely in their viral envelope proteins, suggesting that the block to XMRV and GALV infection is mediated at the level of envelope-receptor interactions. N-linked glycosylation of the receptors was not found to mediate resistance of receptor-expressing BHK cells to GALV or XMRV, as shown by tunicamycin treatment and mutation of the specific glycosylation site of the PiT1 receptor. Hybrid cells produced by fusing BHKXpr1 or BHKPiT1 to XMRV-or GALV-resistant cells, respectively, can mediate efficient XMRV or GALV infection. These findings indicate that BHK cells lack a factor that is required for infection by primate xenotropic viruses. This factor is not required for viruses that use the same receptors but were directly isolated from mice.

Feline Leukemia Virus Infection Requires a Post-Receptor Binding Envelope-Dependent Cellular Component

“…5A), would strongly suggest that NIH 3T3 cells lack an additional cellular component required for FeLV entry rather than favor a role for actin and clathrin in the FeLV resistance. Our findings show strong similarity to previous reports of an ancillary factor for GALV and XMRV (47) and to the requirement of a second cell component for HIV-1 infection (4). However, we cannot exclude the possibility that MDTF and TE671 cells express a cellular factor that counteracts an inhibitor expressed by NIH 3T3 cells or that actin and clathrin are involved in resistance.…”

“…A recent report (47) has suggested that BHK cells lack an ancillary factor required for infection by GALV and by XMRV. To test whether BHK cells also lack a cellular component/ancillary factor required for FeLV infection, we tested the FeLV susceptibility of receptor-expressing BHK cells.…”

“…A limiting accessory factor has also been suggested for ecotropic murine leukemia virus infection (45). Recently, the requirement of an envelope-dependent ancillary factor was implicated for entry by gibbon ape leukemia virus (GALV) and the xenotropic murine retrovirus-related virus (XMRV) (47).…”

“…LacZ-encoding GALV pseudotype virus was also provided by Maribeth Eiden and used in infection studies. This virus was generated by transfection of HEK293T cells with MLV Gag-Pol, LacZ, and GALV envelope expression constructs as described previously (47). LacZ-encoding FeLV-A containing FeLV core (Gag-Pol) proteins was produced by first transducing feline FEA cells with ␤-galactosidase-encoding FeLV-C to introduce the lacZ gene and then transfecting the transduced FEA cells with FeLV-A molecular clone (kindly provided by Brian Willett) using PolyFect transfection reagent (Qiagen, Toronto, ON, Canada).…”

Escape variants of the XPR1 gammaretrovirus receptor are rare due to reliance on a splice donor site and a short hypervariable loop

“…With very few exceptions, mammalian XPR1 proteins have X-MLV receptor function (Xu and Eiden, 2011). This scarcity of escape mutations results in part from the fact that the critical ECL3 residue, K500, is evolutionarily conserved, and by the fact that functional X-MLV receptors can carry multiple substitutions in ECL4 (Yan et al, 2010).…”