Fig. 5: DUX4 expression is critical for HSV-1 and HSV-2 replication.

A DUX4 ko HAP1 cells were generated with CRISPR/Cas9 and DUX4 specific sgRNAs. HSV-1 protein expression in HAP1 wt cells and HAP1 cells with DUX4 knockout. Cells were infected with HSV-1-GFP (MOI of 2) for 20 h and analyzed by western blot. HSP90 was used as loading control. Representative experiment out of n = 3. B mRNA expression (RNA-seq.) of all host genes of HAP1-DUX4-ko cells plotted against HAP1 wt cells, both infected with HSV-1-GFP (MOI of 1) for 8 h. Depicted in red are known DUX4 target genes. C Heatmap of mRNA-expression of selected viral transcripts from HSV-1 (MOI of 1) infected wt and DUX4 ko HAP1 cells 8 h post infection. Marked are viral genes with immediate-early, early and late expression kinetics. D GFP-expression of HAP1 wt and HAP1 DUX4-ko cells infected with HSV-1-GFP (MOI of 0.1) at day 1–4 post infection, measured by flow cytometry (upper panel). In parallel, viral titers were assessed by plaque assays on Vero cells (lower panel). One representative experiment out of n = 3. Values shown are technical replicates. E GFP-expression of HAP1 wt and HAP1 DUX4-ko cells infected with HSV-2-GFP (MOI of 0.05) at day 1–4 post infection, measured by flow cytometry (upper panel). In parallel, viral titers were assessed by plaque assays on Vero cells (lower panel). One representative experiment out of n = 3. Values shown are technical replicates. F Western blot of 293T WT cells and 293T DUX4-ko cells infected with HSV-1 for 18 h. Comparison of HSV-1 protein expression in wt cells and cells with complete DUX4-ko. Actin was used as loading control. Representative experiment out of n = 4. G qRT-PCR analysis of the replication of HSV-1 GFP (F-strain) and HSV-1 KOS at 0,6 and 12 h after infection of HAP1 wt and HAP1 DUX4-ko cells (MOI of 1). Representative experiment out of n = 3. Values shown are technical replicates. H qRT-PCR analysis of the DUX4 target genes TRIM43 as well as the viral genes ICP0 and ICP27 in cells treated with 2,6 nM, 9,8 nM A485 or DMSO and infected with HSV-1 (MOI of 3). Values shown are the biological replicates (n = 3) and presented as mean fold induction +/- SD (normalized to HPRT RNA) relative to uninfected control cells. Source Data are provided as a Source Data file.