(A-B) Immunoblot for total levels of IκBα, phospho-IκBα (Ser32), p65, p50 and phospho-p65 (Ser536) in C2C12 cells with RhoE knockdown (A), or with RhoE overexpression (B). In TNFα treatment groups, cells were treated with 40 ng/ml of TNFα for 15 minutes. (C-D) Nuclear and cytosolic lysates extracted from above cells were immunoblotted for p65 and p50. (E) C2C12 cells were transfected with control siRNA or IκBα-specific siRNA together with empty vector or myc-RhoE expression construct. Total lysates were immunoblotted for IκBα and myc-RhoE. Nuclear and cytosolic lysates were immunoblotted for p65 and p50.