Figure 4

Apoptotic phenotype of EBV-loss Kem-BL clones re-expressing Latency I-associated genes. EBV-loss Kem-BL clones (n1–n3) re-expressing EBNA1 (a and b), EBERs (c and d), miR-BARTs (e and f) or empty vector (controls), marked (+) and (−), respectively, compared with isogenic EBV-loss (EBV −ve) and parental EBV-positive cells (EBV +ve). (a) EBNA1 protein expression, blotted with human AMo serum with probing for actin used as a loading control. (b) Survival of ionomycin-treated EBNA1-expressing EBV-loss BL cells compared with controls. (c) EBER expression was detected by Northern blotting using a probe to the EcoRI J fragment of the EBV genome, using 5 S as a loading control. (d) Survival of ionomycin-treated, EBER-expressing EBV-loss BL cells compared with controls. (e) Expression of mature BART miRs by q-PCR, expressed from the Cluster 1 (top), Cluster 2 (middle) or miR-BART-5 only (bottom) constructs, relative to levels in Kem-BL. (f) Survival of ionomycin-treated miR-BART-expressing EBV-loss BL cells compared with controls. All F-UTG-transduced cells in (a,b,e and f) induced with dox for 24 h before experiments were carried out. In apoptosis assays (b,d and f), cell death was induced by treatment with ionomycin for 48 h. Data are representative of assays that were carried out in triplicate on three independent occasions