The E6AP ubiquitin ligase catalyzes the ubiquitylation and proteasomal degradation of several cellular proteins. Recently, we have shown that E6AP can stabilize β- catenin and that this activity is enhanced by the human papillomavirus (HPV) E6 oncoprotein. In the present study we show that E6AP interacts with β- catenin and ubiquitylates it in a nonproteolytic manner, through atypical ubiquitin linkage that involves K63, K29 and K11, but not K48. The Ubiquitylation of β- catenin by E6AP is specific, it requires the ubiquitin ligase activity of E6AP and is independent of the phosphorylation of β- catenin by GSK3β and activity of the β-catenin “destruction complex”. E6 is not involved in the E6AP mediated ubiquitylation of β- catenin. Interestingly, the E6AP-induced stabilization and activation of β- catenin/TCF transcription, are absolutely dependent on the activity GSK3β, the susceptibility of β- catenin to GSK3β phosphorylation and the ubiquitin ligase activity of E6AP. Collectively, our studies uncover a role for E6AP in regulation of β- catenin ubiquitylation, stability and transcriptional activity, raising the possibility that the nonproteolytic ubiquitylation of β- catenin by E6AP may be a necessary but not sufficient for stabilizing β- catenin and stimulating its transcriptional activity.
