Characterization of a BHK(TK-) cell clone resistant to postattachment entry by herpes simplex virus types 1 and 2

BHK(TK-) cells selected for resistance to polyethylene glycol-mediated fusion give rise to clones that are resistant to herpes simplex virus (HSV) infection. We have characterized one such clone, designated 95-19, and found that it is resistant to entry of HSV type 1 (HSV-1), HSV-2, and the related alphaherpesvirus pseudorabies virus (PRV). Single-step growth experiments show no detectable replication of multiple strains of HSV-1 and HSV-2 on 95-19 cells. Three lines of evidence suggest that these cells are resistant to postattachment entry. (i) Measurements of binding of radiolabeled virus show that heparin-sensitive binding of HSV-1 and HSV-2 to 95-19 cells is identical to binding to BHK(TK-) cells, suggesting that the block to replication occurs after attachment to heparan sulfate proteoglycan. (ii) 95-19 cells exposed to HSV-1 or HSV-2 at high multiplicity show no detectable immediate-early (IE) mRNA expression. (iii) Exposure of attached virus and cells to polyethylene glycol results in partial recovery of both IE gene expression and virus yield in single-step growth. The degrees of recovery of single-step yield and IE gene expression are similar, suggesting that the only block to single-step replication is at the point of virus entry and that these cells are deficient in some cellular factor required for efficient postattachment entry of free virus. 95-19 cells are also highly resistant to entry by cell-to-cell spread, suggesting that the same cellular factor participates in both types of entry.