Figure 7.

HDAC5 Restricts VACV and HSV-1 Infection

(A) Diminished replication of VACV and HSV-1 in an U2OS cell line inducibly expressing HDAC5-FLAG. Infections were performed at MOI = 0.001, for 2 days with VACV and 3 days with HSV-1, after induction with 100 ng/ml doxycycline overnight. Data are represented as mean ± SEM, p values were calculated using a two-tailed t test (n = 3). ^∗p < 0.05 and ^∗∗p < 0.01; ns: not significant.

(B) Immunoblot of HDAC5-FLAG expression in U2OS cell lines.

(C) Enhanced replication of VACV and HSV-1 in HeLa CRISPR/Cas9 HDAC5^−/− knockout clones compared to parental cell lines. Infections were performed as detailed in (A). Data are represented as mean ± SEM, p values were calculated using a two-tailed t test (n = 3). ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001.

(D) Immunoblot confirmed knockout of HDAC5. Sequencing of genomic DNA from clones H5KO1 and H5KO2 confirmed frameshift mutations in both alleles (Table S7A).

(E) Reintroduction of HDAC5 in HDAC5^−/− cells restored restriction of VACV and HSV-1 replication. H5KO1 cells were transduced with either empty vector (EV) or HDAC5-FLAG. Infections were performed as detailed in (A). Data are represented as mean ± SEM, p values were calculated using a two-tailed t test (n = 3). ^∗∗∗p < 0.001 and ^∗∗∗∗p < 0.0001.

(F) Immunoblot analysis of HDAC5-FLAG expression in H5KO1 cell lines.

(G) Enhanced replication of VACV and HSV-1 in HEK293T CRISPR/Cas9 HDAC5^−/− knockout clones compared to parental cell lines. Infections were performed as detailed in (A). Data are represented as mean ± SEM, p values were calculated using a two-tailed t test (n = 3). ^∗p < 0.05 and ^∗∗p < 0.01.

(H) Immunoblot confirming knockout of HDAC5. Sequencing of genomic DNA from clones H5KO3 and H5KO4 confirmed frameshift mutations in both alleles (Table S7A).