PEST-DEPENDENT CYTOPLASMIC RETENTION OF V-REL BY I-KAPPA-B-ALPHA - EVIDENCE THAT I-KAPPA-B-ALPHA REGULATES CELLULAR-LOCALIZATION OF C-REL AND V-REL BY DISTINCT MECHANISMS

Association of c-Rel with the inhibitor of kappa B-alpha (I kappa B-al pha) protein regulates both cellular localization and DNA binding. The ability of v-Rel, the oncogenic viral counterpart of avian c-Rel, to evade regulation by p10, the avian I kappa B-alpha protein, contribute s to v-Rel-mediated oncogenesis. The yeast two-hybrid system was utili zed to dissect Rel:I kappa B-alpha interactions in vivo. We find that distinct domains in c-Rel and v-Rel are required for association with p40. Furthermore, while the ankyrin repeat domain of p40 is sufficient for association with c-Rel, both the ankyrin repeat domain and the PE ST domain are required for association with v-Rel. Two amino acid diff erences between c-Rel and v-Rel that are principally responsible for P EST-dependent association of v-Rel with p40 were identified. These sam e amino acids were principally responsible for PEST-dependent cytoplas mic retention of v-Rel by p40. The presence of mutations in c-Rel that were sufficient to confer PEST-dependent association of the mutant c- Rel protein with p40 did not increase the weak oncogenicity of c-Rel, However, the introduction of these two c-Rel-derived amino acids into v-Rel markedly reduced the oncogenicity of v-Rel. Deletion of the NLS of either c-Rel or v-Rel did not abolish association with p40, but did confer PEST-dependent association of c-Rel with p40. Surprisingly, de letion of the nuclear localization signal in v-Rel did not affect onco genicity by v-Rel, Analysis of several mutant c-Rel and v-Rel proteins demonstrated that association of Rel proteins with p40 is necessary b ut not sufficient for cytoplasmic retention. These results are not con sistent with the hypothesis that p40 regulates cellular localization o f v-Rel and c-Rel by the same mechanism. Rather, these results support the hypothesis that p40 regulates cellular localization of v-Rel and c-Rel by distinct mechanisms.