MONOCLONAL-ANTIBODIES TO DISTINCT SITES ON HERPES-SIMPLEX VIRUS (HSV)GLYCOPROTEIN-D BLOCK HSV BINDING TO HVEM

HVEM (for herpesvirus entry mediator) is a member of the tumor necrosi s factor receptor superfamily and mediates entry of many strains of he rpes simplex virus (HSV) into normally nonpermissive Chinese hamster o vary (CHO) cells. We used sucrose density centrifugation to demonstrat e that purified HSV-1 KOS virions bind directly to a soluble, truncate d form of HVEM (HVEMt) in the absence of any other cell-associated com ponents. Therefore, HVEM mediates HSV entry by serving as a receptor f or the virus. We previously showed that soluble, truncated forms of HS V glycoprotein D (gDt) bind to HVEMt in vitro. Here we show that antib odies specific for gD, but not the other entry glycoproteins gB, gC, o r the gH/gL complex, completely block HSV binding to HVEM. Thus, virio n go is the principal mediator of HSV binding to HVEM. To map sites on virion go which are necessary for its interaction with HVEM, we prein cubated virions with gD-specific monoclonal antibodies (MAbs). MAbs th at recognize antigenic sites Ib and VII of go were the only MAbs which blocked the HSV-HVEM interaction, MAbs from these two groups failed t o coprecipitate HVEMt in the presence of soluble got, whereas the othe r anti-go MAbs coprecipitated HVEMt and got. Previous mapping data ind icated that site VII includes amino acids 11 to 19 and site Ib include s 222 to 252. The current experiments indicate that these sites contai n residues important for HSV binding to HVEM. Group Ib and VII MAbs al so blocked HSV entry into HVEM-expressing CHO cells. These results sug gest that the mechanism of neutralization by these MAbs is via interfe rence with the interaction between go in the virus and HVEM on the cel l. Group Ia and II MAbs failed to block HSV binding to HVEM yet still neutralized HVEM-mediated entry, suggesting that these MAbs block entr y at a step other than HVEM binding.