Fig. 6.

VP24 of all ebolaviruses except RESTV show robust, noncanonical inhibition of upstream IFN induction under multiple conditions. Reporter assays as described in Fig. 2 were conducted to assess potential VP24 antagonism of upstream IFN induction, using the same induction reporters. (A) Cells cotransfected with IFNβ reporter vector were induced 1dpt by tenfold increasing amounts of SeV during the same experiment as that conducted for VP35 in Fig. 2A. (B) Cells cotransfected with an increasing concentration of indicated filovirus VP24 vectors ranging from 20 to 240 ng/well along with the IFNβ reporter across two plates were induced 1dpt by a high MOI of SeV (~22.2) during the same experiment as that conducted with the ISG54 reporter in Fig 5E. (C and D) Cells were induced 1dpt by a high MOI of SeV and harvested 8hpi or 24hpi to assess IRF3 (C) and NF-κB (D) reporter activities, respectively. (E and F) Cells were cotransfected with IFNβ reporter vector, induced 1dpt by human RIG-I CARD vector (E) or poly(I:C) (F) transfection and harvested 24hpi. (G) Activities of FLAG-tagged filovirus VP24 proteins in cells cotransfected with the IFNβ reporter and induced by a high amount of SeV 1dpt were harvested in an experiment analogous to that described in Fig. 5F. FC=fold change of positive induction relative to uninduced negative control. RLU = relative light units. *=p < 0.05; **=p < 0.005; ***=p < 0.0005; ****=p < 0.0001; asterisks above horizontal lines indicate direct comparison of activity values between two antagonists.