FIG 3.

Deletion of aa 207 to 210 does not affect BZLF1 biochemical properties. (A) GFP-tagged BZLF1 and BZLF1Δ207-210 were transfected into 293T cells. At 24 h after transfection, 20 μg/ml cycloheximide was added and the reaction mixtures were incubated for the different times as indicated; then, the cells were collected, and cell extracts were subjected to Western blotting with anti-BZLF1 antibody. The results of examination of the intensity of the BZLF1 bands were quantified, imaged, and analyzed using Li-COR Odyssey and are shown as the means of data from duplicate experiments. (B) GFP-fused BZLF1 and BZLF1Δ207-210 were transfected in 293T cells. After 36 h, the cell lysates were left untreated or incubated with 5 mM cross-linker disuccinimidyl suberate (DSS) at room temperature for 10 min, followed by Western blotting with an anti-BZLF1 antibody. (C) Different amounts of GFP-fused BZLF1 and BZLF1Δ207-210 were transfected in 293T cells for 48 h; then, the nuclear protein fractions were extracted, quantified, and validated with anti-GFP and anti-PCNA antibodies. The same amounts of nuclear proteins were incubated with Cys5.5-labeled BMLF1 promoter probes for 20 min; then, the mixtures were subjected to EMSA, and the images were visualized with Li-COR Odyssey.