Fig. 2: XYLT2 and EXT2 as essential genes for HSV-1 binding and infection. | Communications Biology

Fig. 2: XYLT2 and EXT2 as essential genes for HSV-1 binding and infection.

From: Genome-wide CRISPR screen for HSV-1 host factors reveals PAPSS1 contributes to heparan sulfate synthesis

Fig. 2

a Western blotting analysis confirming the absence or complementation of XYLT2, the EXT2 expression in KO, and rescued clones of HAP1 cells. b, c HepS expression analysis by FACS. After treatment with or without heparinase for 1 h at 37 °C, the cells were stained using an anti-HepS antibody. The graph summarized the mean fluorescent intensity (MFI) of the HepS expression analysis of three independent experiments (c). Results are presented as means ± SEM. Viability of the nontargeting control clone (d) and the XYLT2- and EXT2-KO clones (e) of HAP1 cells after HSV-1 infection. The cells were either pretreated with 5 μM surfen hydrate or DMSO and infected with HSV-1 at a MOI of 9. The cell viabilities were measured via the MTS assay at 48 hpi. The results are presented as means ± SEM of three independent experiments. f Progeny virus production of HSV-1 in the control, XYLT2-KO, and EXT2-KO clones of HAP1 cells. Following surfen hydrate or DMSO treatment, the cells were infected with HSV-1 at a MOI of 1. The progeny viruses were collected at 48 hpi and titrated via the plaque assay. The results are presented as means ± SEM of three independent experiments. g Binding of HSV-1 in the control, XYLT2-KO, and EXT2-KO clones of HAP1 cells. After surfen hydrate or DMSO treatment, the cells were adsorbed with HSV-1 at an MOI of 50 for 1 h and 4 °C. The nontargeting control cells that were mock-infected with PBS were used as a negative control. After removing the residual viruses, the viral DNA was extracted and quantified using real-time PCR. The results are presented as means ± SEM of three independent experiments and shown as the relative DNA level of the control cells. h Viability of the nontargeting control clone, XYLT2-KO, and EXT2-KO human retinal pigment epithelial-1 cell clones after HSV-1 infection. The cells were infected with HSV-1 at an MOI of 9. Then, the viabilities were measured via the MTS assay at 48 hpi. The results are presented as means ± SEM of three independent experiments. Asterisks, p < 0.05; Double asterisks, p < 0.01; n.s. not significant; N.D. not detected.

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