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. 2008 Mar 17;28(10):3538–3547. doi: 10.1128/MCB.02098-07

FIG. 5.

FIG. 5.

IRAK-1 lysines 134 and 180 are required for IL-1-induced NF-κB but not p38 activation. (A) NF-κB activation in 293/IL-1R/TLR4 cells transiently transfected with increasing amounts of IRAK-1 (WT), IRAK-1K134R (K134R), IRAK-1K180R (K180R), or IRAK-1K134/180R (K134/180R). Values were normalized to β-galactosidase activity and are expressed as the induction level compared to luciferase in cells transfected with empty vector (-). Expression of WT, K134R, K180R, and K134/180R was confirmed by immunoblotting (IB) with anti-Flag. A large nonspecific band just below IRAK-1 is denoted by ns. (B) IL-1- and LPS-induced NF-κB luciferase activity was measured in IRAK-1-deficient MEFs transiently transfected with an NF-κB-driven luciferase reporter in combination with vector (-), WT, K134R, K180R, or K134/180R. Each transfection was performed in triplicate, and error bars represent the standard errors of the means. This experiment is representative of five independent experiments. (C) IL-1-induced p38 activation was measured in IRAK-1-deficient MEFs transiently transfected with vector, IRAK-1, or IRAK-1K134/180R. The amount of phosphorylated (p-p38) and total p38 was determined by immunoblotting.