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MAVS activates TBK1 and IKKε through TRAFs in NEMO dependent and independent manner

Fig 5

TRAFs’ coiled-coil domain is important for both interaction and activation of TBK1/IKKε.

(A) Left: A diagram illustrating TRAF truncations. dR deletions: lacking amino acids 34–73 for TRAF2, 68–77 for TRAF3 and 70–109 for TRAF6; dC deletions: lacking amino acids 299–348 for TRAF2, 267–338 for TRAF3 and 288–348 for TRAF6; d1 deletions: lacking amino acids 1–233 for TRAF2, 1–249 for TRAF3 and 1–259 for TRAF6; d2 deletions: lacking amino acids 1–348 for TRAF2, 1–338 for TRAF3 and 1–347 for TRAF6. Right: 293T cells were transfected with Flag-tagged TRAFs or their truncations and HA-tagged TBK1 for 24 h. Cell lysates were immunoprecipitated with the anti-Flag antibody. The precipitates and whole cell lysates (WCL) were analyzed by Western blot with the indicated antibodies. (B) and (C) TRAFs/NEMO-deficient cells reconstituted with TRAF2, 3, or 6 or their truncations were infected with SeV for the indicated times. Cells were analyzed by Western blot to detect the phosphorylation and expression of the indicated proteins (B), or RT-PCR analysis of the indicated gene induction (C). (D) TRAFs/NEMO-deficient cells were transfected with Flag-tagged TRAFs or their truncations and HA-tagged MAVS for 24 h. Cells were analyzed by Western blot to detect the phosphorylation and expression of the indicated proteins. (E) TRAFs–deficient cells reconstituted with TRAF2 or its truncations were infected with SeV for the indicated times. Cells were analyzed by Western blot to detect the phosphorylation and expression of the indicated proteins.

Fig 5

doi: https://doi.org/10.1371/journal.ppat.1006720.g005