NEMO C417R mutation impairs RelA activation and abolishes c-Rel induction in MoDCs stimulated with CD40L but not with LPS. (A) Left panel shows DNA binding activity of NF-κB dimers in MoDCs stimulated with CD40L (2.5 μg/mL) or CD40L plus IFN-γ (2000 U/mL) for 4 hours. Anti–c-Rel antibody was added prior to incubation with the labeled NF-κB DNA probe to supershift c-Rel containing dimers. Right panel shows quantification of RelA and c-Rel activity represented as means ± SEM of fold change compared with unstimulated cells (set at 1.0). (B) NEMO, c-Rel, and RelA protein levels in MoDCs. Cell lysates from 2 healthy volunteers and 1 patient with EDI were subjected to immunoblotting. Blots were probed with anti–c-Rel, anti-RelA, or anti-NEMO antibody. (C) Left panel shows DNA-binding activity of NF-κB dimers in MoDcs stimulated with LPS (2.5 μg/mL) for 4 hours and processed as in panel A. Right panel shows quantification of RelA and c-Rel activity represented as means ± SEM of fold change compared with unstimulated cells (set at 1.0).