Figure 2

Matrin 3 suppression enhances ZAP-mediated restriction of retroviral infection. a Enhanced ZAP-mediated HIV-1 restriction in the absence of Matrin 3 expression. 293TrexhZAP2 cells (Ctrl) or cells silenced for Matrin 3 expression (Matrin 3 KD) were infected with HIV-luc or HR’-CMV-luc then treated with (ZAP) or without doxycycline treatment at 200 ng/ml. Cells were then harvested 2 days postinfection and lysates were subjected to luciferase assays to measure Nef-luc reporter activity. Data presented are the mean RLU/mg values of at least three experiments done in triplicate ±SD. Fold restriction was calculated from dividing Ctrl siRNA values by the results of each treatment indicated. b MoMuLV restriction also improved by Matrin 3 silencing. Data presented are the mean RLU/mg values of three independent experiments done in triplicate. P values ≤0.05 are shown. c Knockdown of Matrin 3 by siRNA treatment. Lysates from a representative experiment were subjected to immunoblot to illustrate Matrin 3 knockdown. Lysates were analyzed for at least three independent experiments. 293TrexhZAP2 were treated with control siRNA (siCtrl) or siRNA to Matrin 3 (siMatrin 3) for two consecutive days followed by infection and doxycycline treatment as stated above. Induction of myc-ZAP2 overexpression was detected with mouse α-myc antibody. GAPDH is the protein loading the control. d Cells transiently expressing mycDDK-tagged Matrin 3 (MDM3) or vector alone (MD) were infected with HIV-luc at two different dilutions, then treated with (ZAP) or without doxycycline (Dox) treatment at 200 ng/ml. Cells were then harvested as stated above. Data presented are the mean RLU/mg values of two experiments done in triplicate. Fold restriction was calculated from dividing MD values by the results of each treatment indicated for each dilution.