Fig. 7.

Pyk2 promotes phosphorylation and activation of IKKβ and TBK1. a IB analysis of phosphorylated IKKβ or total IKKβ in lysates of Pyk2-silenced macrophages or Cd1d^–/– macrophages stimulated with LPS. b IB analysis of phosphorylated IKKβ or total IKKβ in lysates of macrophages pretreated with iGb3 and then stimulated with LPS. c IB analysis of phosphorylated IKKβ and phosphorylated MBP with or without pre-incubating IKKβ with Pyk2 recombinant protein. d Luciferase assay in lysates of HEK293 cells transfected with NF-κB-luc reporter plasmids, plus IKKβ plasmid alone or together with wild type Pyk2 plasmid or Pyk2 kinase inactive mutant (Y402F). e IB analysis of phosphorylated TBK1 or total TBK1 in lysates of LPS-stimulated Pyk2-silenced macrophages or Cd1d^–/– macrophages after IP with TBK1 Ab. f IB analysis of phosphorylated TBK1 or total TBK1 in lysates from Pyk2-silenced macrophages, Cd1d^–/– macrophages or wild type macrophages treated with iGb3, followed by stimulation with LPS. g IB analysis of phosphorylated TBK1 and phosphorylated MBP with or without preincubation TBK1 with Pyk2 protein. h Luciferase assay in lysates of HEK293 cells transfected with IFN-β-luc reporter plasmids, plus TBK1 plasmid alone or together with Pyk2 plasmid or Pyk2 kinase inactive mutant (Y402F). Data are representative of three independent experiments with similar results (a–c, e–g) or are from three independent experiments (d, h; mean ± SEM). **P < 0.01 (ANOVA)