AML1 physically interacts with IKK complex and inhibits its kinase activity. (A-B) Interaction between AML1 and IKK complex. HEK293T cells were transfected with plasmids encoding for HA-AML1, FLAG-IKKα (A) and FLAG-IKKβ (B), as indicated, and extracts were immunoprecipitated with the antibody against FLAG. Western blots of the input lysate or immunoprecipitates were analyzed using the indicated antibodies. (C) Endogenous interaction between AML1 and the IKK complex in Jurkat cells. Cell extract from the Jurkat cells was immunoprecipitated with the antibody against AML1. Western blots of the input lysate or immunoprecipitates were analyzed using the indicated antibodies. (D) Interaction between AML1 and the IKK complex in cytoplasmic or nuclear fraction in HEK293T cells. (E) Endogenous interaction between AML1 and the IKK complex in cytoplasmic or nuclear fraction in Jurkat cells. (F) Endogenous interaction between AML1 and the IKK complex in U937 cells. Cell extract from the U937 cells was immunoprecipitated with the antibody against AML1. Western blots of the total lysate or immunoprecipitates were analyzed using the indicated antibodies. (G-H) In vitro kinase assays of IKKα (G) or IKKβ (H) in HEK293T cells transduced with AML1 or mock. Kinase activities were detected by autoradiography and quantified with ImageJ Version 1.41o software.³¹  Error bars show mean ± SEM. (I) Western blotting of IκBα degradation in HEK293T cells transduced with AML1 or mock. Protein levels of IκBα were quantified with ImageJ Version 1.41o software.³¹  Error bars show mean ± SEM.