IκBα-mediated inhibition of v-Rel DNA binding requires direct interaction with the RXXRXRXXC Rel/κB DNA-binding motif

Rel family proteins bind to κB DNA sites, form heterodimers with one another, and modulate expression of genes linked to κB motifs. IκB factors associate with Rel proteins, inhibit Rel DNA binding in vitro, and displace DNA from DNA-bound Rel complexes. We have investigated the mechanism by which the p40/IκBα inhibitor interferes with Rel DNA-binding activity. Here, we report that p40 contacts the RXXRXRXXC DNA-binding motif conserved in all Rel family proteins, in addition to associating with the nuclear localizing sequence. Competition assays with a Rel-derived peptide comprising the DNA-binding region specifically alleviated p40-mediated inhibition of v-Rel DNA-binding activity, whereas a covalently modified Rel peptide was inactive. Combined, these results indicate that IκBα interaction with the RXXRXRXXC motif is required for inhibition of v-Rel DNA binding and suggest that nuclear IκB factors may be critical for regulating transcription by Rel family proteins.