Figure 1.

NF-κB activation by flagellin is Toll dependent. NCIH292 cells were co-transfected with an NF-κB TA-luciferase reporter (BD Biosciences) and either empty vector (EV, solid black bar) or DNM (hatched bar) constructs for TLR5 (left panel), TLR2 (middle panel), or MyD88 (right panel). Thirty-six hours after transfection, cells were stimulated with flagellin (Flgn, 10 μg/ml), an agonist antibody directed against the flagellin receptor, asialoGM1 (α-ASGM1, 1:100 dilution), or lipoteichoic acid (LTA, 50 μg/ml) for 6 h before cell lysis. Both flagellin and α-ASGM1 induced NF-κB luciferase activity. The induction by both agents was abolished by DNMs directed against TLR5 and MyD88. To indicate specificity, flagellin-induced NF-κB activation was unaffected by the TLR2 DNM and NF-κB activation by LTA was unaffected by the MyD88 DNM (22).