Fig. 1.

Inflammasome sensors regulate the EBV replication switch. (A) HH514-16 BL cells were exposed to control siRNA or inflammasome sensor-directed siRNAs, treated with the replicative/lytic cycle-inducing agent NaB 24 h later, and harvested after another 24 h to assay intracellular levels of the EBV replication switch protein ZEBRA and GAPDH by in-cell Western. (B) HH514-16 BL cells were treated with NaB (versus untreated control), harvested 24 h later, stained with anti-ZEBRA antibody, and subjected to flow cytometry. ZEBRA⁺ gate was set based on staining with isotype control antibody; numbers represent percent ZEBRA⁺ cells. (C) Cells treated and harvested as in A were immunoblotted for ZEBRA and β-actin. Negative (Neg) control, untreated cells exposed to control siRNA; positive (Pos) control, NaB-treated cells exposed to control siRNA in A and C. Data in A represent means of three independent experiments; error bars, SEM; *P ≤ 0.05. Sensors whose knockdown resulted in at least 25% reduction in ZEBRA levels compared to control (Pos) in A were tested in C. (D) ZEBRA levels (normalized to β-actin) in siRNA-knocked down cells relative to control siRNA but NaB-treated cells (Pos; shown in C). Inflammasome components whose knockdown resulted in at least 25% loss of ZEBRA in A and D (via in-cell Western and immunoblot) are indicated in blue.