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A
293T cells were transfected with control or TRAF6 siRNA for 24 h, followed by transfection with an IFN‐β promoter reporter plasmid and pRL‐TK plasmid for 12 h, and analyzed for IFN‐β‐dependent luciferase activity (fold induction) after SeV (MOI = 0.1) infection for indicated time points.
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B
Luciferase activity in 293T cells transfected with the ISRE promoter reporter plasmid and pRL‐TK plasmid, together with expression plasmid encoding TRAF6 and increasing amount of NLRP11.
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C
293T cells were transfected with Flag‐NLRP11, and then infected by SeV (MOI = 1) for indicated time points. MG132 was used to treat the cells for 6 h before harvesting. Whole‐cell extracts were immunoprecipitated with anti‐Flag beads, followed by IB analysis with anti‐TRAF6 antibody.
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D, E
Immunoblot analysis of HA‐TRAF6 (D) or endogenous TRAF6 (E) in 293T cells transfected with Flag‐NLRP11 or Flag‐NLRP4 (D) or infected with Sendai virus (SeV) (MOI = 1) (E) for 24 h.
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F
WT or NLRP11 KO THP‐1 cells were pretreated with CHX for 2 h, and then infected with SeV (MOI = 1) for indicated time points, followed by immunoblot analysis.
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G
293T cells were transfected with Myc‐NLRP11 and HA‐TRAF6 for 24 h, and then treated with the indicated inhibitors for 6 h, followed by immunoblot analysis.
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H
293T cells were transfected with empty vector or Myc‐NLRP11 for 12 h, infected with SeV (MOI = 1) for 12 h, and then subjected to MG132 treatment for 6 h before harvesting. The cell lysates were subjected to immunoprecipitation with anti‐TRAF6 antibody followed by immunoblot analysis with indicated antibodies.
Data information: Data in (A and B) are expressed as means ± SEM of three independent experiments (**
< 0.001, versus cells transfected with control siRNA or EV with the same treatment, Student's
‐test).
