FIG 6.

Riplet DN suppresses RIG-I signaling but augments ZAP-mediated restriction of HIV-1 reporter virus. (A) Function of Riplet DN in the RIG-I pathway measured by a surrogate reporter of IFN-β induction upon poly(I·C) stimulation. HEK293 cells were untreated (Cntl) or transiently transfected with the indicated DNAs along with a construct expressing luciferase reporter under IFN-β promoter control. Twenty-four hours later, cells were stimulated with poly(I·C) (left) or were unstimulated (right), and reporter expression was measured 6 h later. Data points represent the mean ± SD values of three independent experiments. (B) Riplet DN overexpression augments ZAP-mediated restriction of retroviral infection. 293TrexhZAP cells transfected with empty vector (EV) or with DNAs expressing the indicated constructs were infected with HIV-luc reporter virus. Cells were treated with doxycycline to induce ZAP expression or DMSO as control at 6 h postinfection. Firefly luciferase reporter activity was measured at 24 h postinfection and normalized to total protein content measured by a Bradford assay for each sample. Data points presented are the mean RLU/mg ± SD values of four independent experiments done in triplicate. (C) Fold ZAP-mediated inhibition of HIV-1 reporter expression derived from data in panel B. Fold virus inhibition by ZAP was calculated as the ratio of luciferase expression levels in DMSO-treated cells to those in doxycycline-treated cells. Data points represent the mean ± SD values of four independent experiments.