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. 2013 Dec 10;24(3):359–371. doi: 10.1038/cr.2013.162

Figure 3.

Figure 3

RACK1 interacted with IKKα and IKKβ. (A) Overexpressed RACK1 interacted with the overexpressed IKK complex. Myc-RACK1-293T stable cell lines were transfected with the indicated plasmids. Twenty-four hours later, the cells were harvested, and immunoprecipitated with the anti-HA antibody and analyzed by immunoblotting with anti-Flag antibody. (B) GST-RACK1 pulled down endogenous IKKα and IKKβ, but not IKKγ. 293T cell extracts were incubated with GST alone or GST-RACK1 and then immunoprecipitated with Sepharose beads. Immunoblots of the immunoprecipitates were analyzed with immunoprecipitated IKK antibodies. (C) RACK1 physically interacted with IKKα and IKKβ. 293T or RAW264.7 cell extracts were immunoprecipitated with mouse IgM or anti-RACK1 antibody and then subjected to immunoblot analysis of the endogenous IKK proteins. (D) RACK1 interacted with IKK in a TNF-triggered manner. 293T cells were treated with TNF (10 ng/ml) for the indicated time, and the cells were then harvested for immunoprecipitation with mouse IgM or anti-RACK1 antibody. The interacting IKK proteins were analyzed by immunoblotting.