Figure 3.

Effect of MG 132 on R. rickettsii induced NF-κB activation. Gel shift analysis using a ³²P-labeled oligonucleotide probe was performed on nuclear extracts prepared from control ECs (C) and ECs infected for 3 hr with R. rickettsii (RR) in the absence and presence of 25 μM (25) or 50 μM (50) MG 132. Analysis of complex formation was performed on 4% nonreducing polyacrylamide gels followed by autoradiography. An R. rickettsii-inducible gel-shifted complex representing interaction of dimeric NF-κB molecules with the labeled probe is indicated in the gel (NF-κB) as is the presence of a nonspecific noninducible gel-shifted complex (NS). Specificity of the NF-κB gel-shifted complexes was demonstrated by inclusion of excess unlabeled oligonucleotide probe (+cold).