👤 Nadine Jarousse

Kaposi's sarcoma-associated herpesvirus (KSHV) and its murine homolog, murine gammaherpesvirus 68 (MHV68), are lymphotropic viruses that establish latent infection in their host. Surprisingly, while B cells are the main viral reservoir in vivo, B-cell lines are poorly permissive to infection by either MHV68 or KSHV. Here, we report that most B-cell lines express very little to no cell surface heparan sulfate (HS), a glycosaminoglycan that is essential for infection by these viruses. We found that Ext1, a key enzyme in the biosynthesis of HS, was expressed at a low level in these cells. Transfection of B-cell lines with Ext1 restored high HS expression at the cell surface. Overexpression of Ext1 in murine A20 and M12 B-cell lines increased MHV68 surface binding and enhanced the efficiency of infection. Finally, although it was not sufficient to allow efficient infection, the expression of HS on BJAB cells promoted KSHV binding at the cell surface. Thus, our results indicate that MHV68 and KSHV cycles are blocked in B-cell lines at the binding step due to a lack of surface HS. Show less

Murine gammaherpesvirus 68 (MHV68) is used as a model to study gammaherpesvirus pathogenesis both in tissue culture systems and in vivo. We used a gene-trapping approach to get insight into cellular factors involved in MHV68 infection. By generating a library of gene-trapped CHO cells, we were able to isolate several clones that exhibited various degrees of resistance to MHV68-induced cytopathic effect. Clones that showed the highest degree of resistance were affected at the early stage of the viral cycle, with the vast majority of these clones being deficient for heparan sulfate (HS) expression at the cell surface. Heparan sulfate expression could be restored in all the HS-deficient clones by expression of EXT1, an enzyme that is essential for the biosynthesis of HS. Consistent with the role of HS in viral entry, HS-deficient CHO cells did not support viral internalization. Cell surface heparan sulfate proteoglycans (HSPG) are mostly composed of HS chains attached to two families of core proteins, the transmembrane syndecans and the GPI-anchored glypicans. Treatment of CHO cells with phosphatidylinositol-specific phospholipase C (PI-PLC) did not significantly affect the level of HS expression, indicating that the glypicans are not a major source of HSPG in CHO cells. By contrast, treatment of CHO cells with PMA, a drug known to accelerate syndecan shedding, resulted in a decrease in both HS expression and susceptibility to MHV68; these effects were abolished by TIMP-3, a specific inhibitor of syndecan shedding. All together, our results confirm the essential role of HS in MHV68 infection and identify the syndecans as a major source of HSPG used by the virus as coreceptors to infect CHO cells. Show less