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. 2003 Nov;23(22):8282–8294. doi: 10.1128/MCB.23.22.8282-8294.2003

FIG. 5.

FIG. 5.

Enhanced KSHV lytic replication by RTA MUT2 mutant. (A) Level of KSHV lytic reactivation. After the stimulation with doxycycline (0.1 μg/ml), TRExBCBL1-pcDNA5, TRExBCBL1-RTA, and TRExBCBL1-MUT2 cells were fixed by paraformaldehyde and reacted with K8.1 rabbit sera followed by fluorescein isothiocyanate-conjugated anti-rabbit secondary antibody. The data were reproduced in three independent experiments. (B) Expression of KSHV vIL-6 and K8 protein induced by RTA and the RTA MUT2 mutant. After doxycycline treatment for 0, 1, 2, and 3 days, KSHV-infected TRExBCBL1-RTA (WT) and TRExBCBL1-RTA MUT2 (MUT2) cells were lysed, and 10 μg of total proteins was subjected to immunoblotting with anti-RTA, anti-vIL-6, and anti-K8 antibodies. (C) ChIP assay of RTA-dependent promoters. KSHV-infected TRExBCBL1-pcDNA5, TRExBCBL1-RTA, and TRExBCBL1-TRA MUT2 cells were collected after 24 h of 0.1-μg/ml doxycycline treatment. ChIP assays were performed with an anti-His (αHis) antibody (Ab). PCR products corresponding to each viral promoter were generated from an aliquot (2%) of total immunoprecipitated material (Input).