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. Author manuscript; available in PMC: 2014 Feb 13.
Published in final edited form as: Cell Host Microbe. 2013 Feb 13;13(2):204–214. doi: 10.1016/j.chom.2012.12.005

Figure 3. The Major Lytic Switch Protein, KSHV ORF50/RTA, is Activated Following TLK2 Knockdown.

Figure 3

A) The KSHV ORF50 promoter is activated by TLK2 knockdown. HEK-293 cells were transfected with GAPDH siRNA or a pool of TLK1 or TLK2 siRNAs (50nM) for 24 hours followed by transfection with an ORF50 luciferase reporter construct. Lysate was harvested 48 hours after ORF50-luciferase transfection and a luciferase assay was performed. Values are normalized to protein levels determined by Bradford assay. B) Western blots were performed for TLK1, TLK2, and tubulin to confirm knockdown. C) Increase in ORF50 mRNA. Levels of ORF50 viral transcript were measured by qPCR as described in 2C. D) Induction of other viral lytic genes. KSHV-293 cells were reverse transfected with GAPDH siRNA or a pool of TLK1 or TLK2 siRNAs (50nM) for 96 hours. Total RNA was harvested and reverse transcriptase PCR was performed on the RNA (+/− reverse transcriptase (RT)) and PCR products were run out on agarose gels. M, Marker lane. Error bars represent standard deviation from the mean. Data were analyzed using a two-tailed type II Student’s t test for significance.