Skip to main content
. 2010 Feb 11;5(2):e9172. doi: 10.1371/journal.pone.0009172

Figure 4. TRAF5 is required for IRF3/7 and NF-κB activation in response to RLR signaling.

Figure 4

A HEK293T cells were initially transfected with siRNAs targeting GFP, TRAF3, or TRAF5. 48 hrs later, cells were transfected with pLuc-IFNβ and 6 hrs later, poly I:C was transfected. Luciferase activity was measured 18 hours following poly I:C transfection. In the inset figure, HA-TRAF3 or HA-TRAF5 was cotransfected with siRNAs targeting GFP, TRAF3, or TRAF5 and anti-HA immunoblots were performed on prepared whole cell lysates. B HEK293T cells were initially transfected with siRNAs targeting GFP, TRAF3, or TRAF5. 48 hrs later, cells were transfected with HA-IRF3, lysates were prepared, and anti-HA immunoprecipitates were probed for phosphorylated IRF3 (Ser 396) or total HA-IRF3. Cells were transfected with poly I:C 8 hrs prior to cell lysis. C HEK293T cells were transfected with siRNAs targeting GFP, TRAF3, or TRAF5. Cell lysates probed for phospho-STAT1 (Y701) or total STAT1. Poly I:C was transfected 8 hrs prior to cell lysis. D, E Luciferase reporter assays were performed in HEK293T cells transfected with siRNA targeting GFP, MAVS, or TRAF5. 48 hrs following siRNA transfection, cells were transfected with pLuc-IFN-β, pLuc-PRD(III-I)3 (IRF3/7) or pLuc-PRD(II)2 (NF-κB). SeV infection was performed 4 hrs following transfection and 20 hrs prior to luciferase assay.