FIG 1.

Cells expressing high levels of STAT3 protein are refractory to spontaneous and induced KSHV lytic activation. BCBL-1 cells were treated with TPA or VPA or left untreated (U). Cells were harvested at 1, 24, and 48 h (A and B) and at 48 h (C) posttreatment. (A) RNA was isolated and subjected to qRT-PCR to determine the relative levels of lytic ORFK8.1 transcripts. Fold changes were calculated by the ΔΔC_T method, normalized to three housekeeping genes, GAPDH, HPRTI, and B2M (β2 microglobulin). The data are presented as means and standard errors of the mean (SEM) and are representative of the results of two experiments with 3 technical replicates. (B) Cell lysates were immunoblotted using antibodies to K8.1 and β-actin. (C) Cells were immunostained for K8.1 and STAT3 and subjected to flow cytometry. The numbers within the dot plots in the left column indicate the percent K8.1^hi cells under untreated or TPA- or VPA-treated conditions after comparison with similarly treated cells stained with isotype control antibody; the dashed oval gates indicate K8.1^lo cells. The numbers within the dot plots in the three middle columns indicate the percent K8.1^hi cells in subpopulations expressing different levels of STAT3, i.e., STAT3^hi (high), STAT3^int (intermediate), and STAT3^lo (low). Numbers in the dot plots on the right indicate the percent K8.1^lo cells (from the oval gates in the left-hand dot plots) expressing low, intermediate, and high levels of STAT3.