Figure 4.

TG promptly induces an extended antiviral state targeting influenza virus post-translationally. (A) NPTr cells primed with 0.5 µM for 30 min were immediately infected or further cultured for 48 h prior to infection with USSR H1N1 virus at 0.5 MOI for 24 h. Spun sns were used to infect MDCK cells for 6 h in FFAs. Significance determined by paired t-test within each time point. (B) NPTr and (C) NHBE cells were incubated with TG (0.5 and 0.01 µM, respectively) for 30 min and immediately infected (TG pre-infection) or were first infected for 6 h followed by TG exposure for 30 min (TG post-infection). NPTr and NHBE cells were infected with USSR H1N1 virus at 0.5 and 1.0 MOI, respectively. TG pre-primed cells were infected for 2 h, PBS washed and cultured in fresh infection media overnight. Cells 6 h post-infection were treated with TG for 30 min, washed with PBS and cultured in fresh infection media overnight. Spun sns were used to infect MDCK cells for 6 h in FFAs. Significance determined by paired t-test within each time point. (D,F) NPTr primed with (0.5 or 1.0 µM) and (E,G) NHBE cells primed with TG (0.005 or 0.01 µM respectively) were subsequently infected with USSR H1N1 at 0.5 and 1.0 MOI respectively for 24 h. (D,E) Total RNA extracted to determine M gene expression for all samples, normalised to 18S rRNA. Significance determined by one-way ANOVA, relative to DMSO control. (F,G) Protein lysates (DMSO and highest selected TG concentration) harvested at 24 h post-infection were used to ascertain viral NP and M1 protein levels, normalised to beta-actin. Representative blots displayed. Significance determined by paired t-test. * p < 0.05, **** p < 0.0001. All assays were in triplicates and were performed three times.