Figure 3. The IL-1-stimulated activation of IRAK1 does not require the catalytic activity of IRAK4.

(A) IL-1R cells were incubated for 1 h without (−) or with (+) 3 µM IRAK4-IN-1 to inhibit cellular IRAK4 and stimulated with IL-1β for the times indicated. IRAK1 was immunoprecipitated from 1 mg of cell extract protein with anti-IRAK1 or control IgG and assayed with GST-Pellino1 and Mg[γ³²P-ATP] as substrates in the presence (+) of 1 µM IRAK4-IN-1. Further details are given in Figure 2A. (B) Same as in A, except that the cell culture medium was incubated with 10 µM JNK-IN-7 to inactivate cellular IRAK1 activity, and IRAK4 was immunoprecipitated from the cell extracts instead of IRAK1, and co-immunoprecipitating IRAK4 was assayed by including JNK-IN-7 in the assay to inhibit IRAK1.