FIG 3.
vPK epigenetically regulates to enhance transcription of BZLF1. (A and B) CLIX-FZ BL cells were transfected with empty vector (EV) or pFLAG-vPK (A) or with scrambled siRNA or si-BGLF4 (B), treated with doxycycline after 24 h (A) or 8 h (B), and harvested at different times after treatment for immunoblotting with the indicated antibodies. Numbers indicate the relative amounts of FLAG-ZEBRA and vPK after normalizing to β-actin. (C and D) HH514-16 BL cells were transfected with empty vector (EV) or pFLAG-vPK and treated with NaB after 24 h (C) or transfected with scrambled siRNA or si-BGLF4 and treated with NaB after 8 h (D). Cells were harvested at indicated times after treatment and examined by qRT-PCR using primers directed against BZLF1 transcript and using the ΔΔCT method. (E and F) CLIX-G4 eBL cells were left untreated or were treated with NaB or NaB plus doxycycline (to induce expression of vPK) and harvested after 24 h for chromatin immunoprecipitation with control antibody (IgG) or antibody against RNA polymerase II (E), histone 3 (F), H3K9me3 (F), or H3Ac (F). Relative occupancy of BZLF1 promoter (Zp) by RNA polymerase II (E) and modified histone 3 (F) was determined via qPCR using primers targeting the BZLF1 promoter. All experiments were performed between 2 and 3 times. Error bars in panels C to F represent 3 technical replicates from 2 experiments.