Mutation of Host Cell Determinants which Discriminate between Lytic and Persistent Mouse Hepatitis Virus Infection Results in a Fusion-resistant Phenotype

Abstract

SUMMARYThe expression of mouse hepatitis virus (MHV) E2-specific mRNA, the E 2 polypeptide and its associated cell fusing activity was monitored in various cell types inoculated with a recombinant vaccinia virus, designated vMS containing the Ez gene. The results suggest that host cell permissiveness to MHV infection correlates with cellular susceptibility to membrane fusion mediated by the MHV E2 glycoprotein. In addition, we utilized a genetic approach to the analysis of host cell functions involved in determining permissiveness to MHV. By using the chemical mutagen ethyl methanesulphonate, mouse fibroblast cell mutants were generated and selected for their resistance to cell killing by MHV. When challenged with MHV, all five mutants examined gave rise to persistent infections, in contrast to wild-type L-2 cells which were rapidly killed by the virus. The results provide genetic evidence in support of a previous correlation proposed between MHV permissiveness and two host determinants, namely susceptibility to MHV infection and to MHV-mediated cell fusion. Fusion resistance was specific to fusion mediated by the MHV E2 glycoprotein as shown in contact fusion assays between uninfected cells and cells infected either with MHV or with an E2-expressing recombinant vaccinia virus. In contrast, mutant cells were not resistant to fusion after treatment with polyethylene glycol. The observed high rate of generation of these mutants suggests that the conversion of a fully MHVsusceptible cell to a semi-resistant one is a fairly common event, possibly involving a single mutation. In this case, resistance to MHV infection and to E2-mediated membrane fusion may depend on a common host function. This result provides prospects for the precise genetic and biochemical characterization of the steps involved in host cell permissiveness to MHV infection. INTRODUCTIONMouse hepatitis virus (MHV) produces infections ranging from persistent to acute, depending on the host cell type as well as other factors. Despite recent advances in the molecular biology of coronaviruses such as MHV, the mechanisms which determine the outcome of infection remain largely unclear. MHV gives rise to certain cytopathic functions, such as cell fusion and inhibition of host cell protein synthesis, which, if differentially expressed in a cell-specific manner, could determine the severity of MHV infection. In fact, cell differences in susceptibility to MHV-induced fusion have been implicated as host cell factors which, at least in part,

Molecular cloning and expression of a spike protein of neurovirulent murine coronavirus JHMV variant c1-2

“…Interestingly, syncytium formation was also observed in RK 13 cells, which are not susceptible to MHV infection. Previously, it was thought that MHV S protein induced syncytium formation only in mouse cells (Vennema et al, 1990); there is a correlation between permissiveness to MHV and susceptibility to fusion by the S protein (Daya et al, 1989).…”

Proteolytic cleavage of the murine coronavirus surface glycoprotein is not required for fusion activity

“…Sturman and coworkers (Frana et al, 1985) have found that there are significant differences in the susceptibility of murine cell lines to MHV-mediated fusion from without. Similarly Anderson and coworkers (Mizzen et al, 1983;Daya et al, 1989) conclude that host cell permissiveness to MHV infection correlates with cellular susceptibility to membrane fusion. Also, Kooi et al (1991), Cervin & Anderson (1991) and Roos et al (1990) have shown that the susceptibility of cells to MHV-induced fusion can be altered by modification of the membrane lipid composition.…”

The Role of the MHV Receptor and Related Glycoproteins in Murine Hepatitis Virus Infection of Murine Cell Lines

“…, 1985 ) . The lipid composition of the cell membrane (Daya, et al, 1988;RODS, et aI" 1990), an unknown membrane component (Daya, et al , 1989) may also affect coronav irus-induced cell-cell fusion . The structure of MHV spike glycoprotein has important effects on MHV -induced cell fusion .…”