Figure 5.
In vitro (A) and in vivo (B,C) replication of 17VP16pEgr1/Sp1, its rescue, and wild-type parental strain 17syn+. (A). Multistep in vitro replication kinetics of parental strain 17syn+ and three independent isolates of mutant 17VP16pEgr1/Sp1. Shown is one of two biological replicates. These mutants contain a refined 4-bp mutation in the putative Egr-1/Sp1 site at position 105,134 to 105,137 in the HSV-1 VP16 promoter/5′UTR sequence (see Figure 1B). (B) In vivo replication in eyes. Groups of 4–5 mice were inoculated on scarified corneas with 2 × 105 pfu of each of the 17VP16pEgr1/Sp1 and rescued viruses as indicated. In addition, groups of mice were infected with 2 × 105 pfu 17VP16πRR or its rescue 17VP16πRR-R. On day 4 pi, eyes and TG were harvested and infectious viral titers quantified by standard plaque assay. Replication on the eye (B) is not influenced by this mutation in the Egr-1/Sp1 site in the 5′UTR. By contrast, this Egr-1/Sp1 mutation results in a significant reduction in viral titers in the TG. This effect was also observed for the triple TF 5′UTR mutant, 17VP16πRR which was reported previously (Sawtell and Thompson, 2016a,b) (C) Shown are results from two independent biological replicates. ***p ≤ 0.001.