FIG 3.

HSV-1 US3 inhibits TNF-α-induced NF-κB cytokine expression. (A) HEK293T cells were cotransfected with pCMV-Flag control vector or US3-Flag expression plasmid along with pIL-8-Luc and RL-TK. Twenty-four hours posttransfection, cells were treated with TNF-α for 6 h and luciferase activity was measured. The expression of US3 was analyzed by WB using anti-Flag and anti-β-actin monoclonal antibodies. (B) HEK293T cells were transfected with pCMV-Flag control vector or US3-Flag. Twenty-four hours posttransfection, cells were mock treated or stimulated with TNF-α for 6 h before RT-PCR was performed using GAPDH and IL-8-primers. (C) HEK293T cells were mock infected or infected with WT HSV-1, US3-Del-HSV-1, K220M-HSV-1, D305A-HSV-1, and US3-repaired-HSV-1. Sixteen hours postinfection, cells were mock treated or treated with TNF-α for 6 h, and then the total RNA was analyzed by RT-PCR. The data represent means plus standard deviations for three replicates. Statistical analysis was performed using Student's t test. **, P < 0.01.