Fig 1.

Function of ICP47 in mouse cells in vitro and in vivo. (A) Schematic showing nucleotide modifications (in red) in HSV-1 ICP47del and ICP47rev. Initiating codons (green) and stop (*) are marked, as are HindIII and XbaI enzyme sites (underlined) used to identify the recombinants. (B) Replication of recombinants assessed in Vero cells in comparison to parent virus. Data are shown as mean ± SEM from three replicates (error bars are obscured by data points). (C) Surface MHC-I (H-2K^b) levels were quantified on 293KbC2 (left) and MC57G (right) cells after infection with HSV-1 recombinants or parent. Data are H-2K^b level relative to mock-infected samples from four independent experiments. (D) Surface MHC-I (H-2K^k and H-2D^k) levels were quantified on L929 cells after infection with HSV-1 recombinants or parent. Data are H-2K^k or H-2D^k level relative to mock-infected samples from two independent experiments. (E) 293KbC2 (left) and MC57G (right) cells were infected with HSV-1 recombinants or parent, then with SIINFEKL-expressing M-miniOVA₂₅₇, or M-miniFlu (as negative control). Data are the fold change in the levels of H-2K^b-SIINFEKL relative to the negative control, and the dashed line represents the limit of detection. (F and G) Mice were infected on the flank with HSV-1 parent or recombinants by tattoo and viral loads (F) in the skin and DRG measured after 4 and 6 days, and CD8⁺ T cell responses (G) in spleen were measured. (F) Data are virus titers in an individual mouse derived from two independent experiments, and the dotted line signifies the limit of detection. (G) The percentage of CD8⁺ cells expressing IFNγ⁺ after stimulation with the HSV-1 peptides as marked is shown. Data are pooled from four independent experiments, each with three mice per group. The ends of boxes signify the interquartile range, the line shows the median, and the whiskers indicate maximum and minimum values. All panels unless specified: bars show mean ± SEM and statistical significance was determined using one-way ANOVA with Tukey’s multiple-comparison test (*P < 0.05; **P < 0.01; ***P < 0.001; and unmarked, not significant).