Fig 1.
Inhibition of R8102 infection by nystantin (A), filipin (B), dynasore (C), or the dominant-negative DynK44A-GFP (D) in CHO cells expressing the following receptors: nectin1 alone (Nec), nectin1 plus αVβ3-integrin (Nec-αVβ3), nectin-GPI (NecGPI), and nectin-EGFR (NecEGFR). Cells were pretreated with inhibitors at the amounts indicated in the abscissas and infected with R8102 (3 PFU/ml) for 90 min at 37°C in the same medium. Inoculum was removed, and cells were overlaid with Dulbecco's minimal essential medium (DMEM) containing or not the appropriate inhibitor for a further 8 h. For filipin, cells were preincubated with the compound at 37°C for 30 min and infected for 30 min (30 PFU/ml) in the same medium. For nystatin, cells were preincubated with nystatin for 16 h, the inhibitor was removed, and cells were washed and infected for 60 min at 37°C (3 PFU/ml) in the absence of nystatin. With both inhibitors, infected cells were overlaid without inhibitor. For dynasore, cells were preincubated for 60 min at 37°C. (D) Cells were transfected with DynK44A-GFP or wt-DynGFP (16) encoding dynamin2 fused to green fluorescent protein (GFP) in the wt or dominant-negative (DN) version (K44A substitution) 18 h prior to infection. In all assays, each point represents averages ± standard deviations (vertical bars) of the results of experiments performed in triplicate. HSV R8102 recombinant expresses lacZ under the control of the α27 promoter (6). Infection was quantified as the β-Gal level; results are expressed as percentages compared to untreated cells.