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. Author manuscript; available in PMC: 2018 Jun 12.
Published in final edited form as: Cell Host Microbe. 2017 Apr 12;21(4):507–517.e5. doi: 10.1016/j.chom.2017.03.007

Figure 3. AFF4 localizes to early HSV transcriptional foci and is required for efficient IE expression.

Figure 3

(A) MRC5 cells were mock infected or infected with HSV-1 for 2 h. Cells were co-stained with antibodies to the SEC scaffold component AFF4 and the HSV-1 lytic activator ICP4. (B–C) MRC5 cells were transfected with siControl or siRNAs to AFF4 or BRD4. Cells were infected with HSV-1 for 2 h and mRNA levels of the siRNA target (AFF4, BRD4), control cellular gene (GAPDH), and viral IE genes (ICP4, ICP27, ICP22) were determined. Data are levels in siAFF4 or siBRD4 cells relative to control siRNA cells. (Means +/− s.e.m., n > = 6). (D) JQ1+, IBET-762, and HMBA bind to the bromodomains of BRD4 and inhibit its binding to chromatin. These compounds also induce release of 7SK snRNP sequestered P-TEFb. (E–F) MRC5 cells treated with JQ1+ 1 uM, IBET-762 1 uM, or HMBA 5 mM and infected with HSV-1 for 2 h. The mRNA levels of control cellular gene GAPDH and viral IE genes (ICP4, ICP27, ICP22) are relative to those in vehicle treated cells. (Means +/− s.e.m., n > = 6). (G–H) mRNA levels of HSV IE genes and controls (GAPDH, HPRT) in cells treated with the indicated concentrations of CDK9 inhibitors. (Means +/− s.e.m., n = 3). (I–J) MRC5 cells treated with vehicle, JQ1+, or dBET1 and infected with HSV-1 for 2 h. (I) Western blot of ICP4, BRD4, and control GAPDH protein levels. (J) mRNA levels of viral IE and GAPDH are relative to those in vehicle treated cells. (Means +/− s.e.m., n = 3). See also Figures S1–S6.