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. Author manuscript; available in PMC: 2012 May 18.
Published in final edited form as: Mol Cell. 2011 Nov 18;44(4):559–571. doi: 10.1016/j.molcel.2011.09.015

Figure 1. A20 inhibits TNFα-induced IKK activation without antagonizing RIP1 ubiquitination.

Figure 1

(A) HEK293 cells stably overexpressing A20 and the parental (WT) HEK293 cells were stimulated with TNFα for the indicated time, then IKK complex was immunoprecipitated with a NEMO antibody for kinase assay using GST-IκBα N-terminus and γ-32P-ATP as substrates. Immunoprecipitates and cell lysates were immunoblotted with the indicated antibodies. N.S.: non-specific. (B) TNF receptor complexes were pulled down using glutathione Sepharose following stimulation of cells with GST-TNFα, and the proteins in the complexes were detected by immunoblotting with the indicated antibodies. (C) Similar to (B), except that a NEMO antibody was used for immunoprecipitation. (D) Parental HEK293 cells (WT) or HEK293 cells stably expressing A20 WT or A20 C103A with N-terminal TAP tag and C-terminal Flag tag were treated with TNFα for the indicated timepoints. Cell lysates were used for immunoblotting with the indicated antibodies.