RB and c-Myc Activate Expression of the E-Cadherin Gene in Epithelial Cells through Interaction with Transcription Factor AP-2

ABSTRACT

E-cadherin plays a pivotal role in the biogenesis of the first epithelium during development, and its down-regulation is associated with metastasis of carcinomas. We recently reported that inactivation of RB family proteins by simian virus 40 large T antigen (LT) in MDCK epithelial cells results in a mesenchymal conversion associated with invasiveness and a down-regulation of c-Myc. Reexpression of RB or c-Myc in such cells allows the reexpression of epithelial markers including E-cadherin. Here we show that both RB and c-Myc specifically activate transcription of the E-cadherin promoter in epithelial cells but not in NIH 3T3 mesenchymal cells. This transcriptional activity is mediated in both cases by the transcription factor AP-2. In vitro AP-2 and RB interaction involves the N-terminal domain of AP-2 and the oncoprotein binding domain and C-terminal domain of RB. In vivo physical interaction between RB and AP-2 was demonstrated in MDCK and HaCat cells. In LT-transformed MDCK cells, LT, RB, and AP-2 were all coimmunoprecipitated by each of the corresponding antibodies, and a mutation of the RB binding domain of the oncoprotein inhibited its binding to both RB and AP-2. Taken together, our results suggest that there is a tripartite complex between LT, RB, and AP-2 and that the physical and functional interactions between LT and AP-2 are mediated by RB. Moreover, they define RB and c-Myc as coactivators of AP-2 in epithelial cells and shed new light on the significance of the LT-RB complex, linking it to the dedifferentiation processes occurring during tumor progression. These data confirm the important role for RB and c-Myc in the maintenance of the epithelial phenotype and reveal a novel mechanism of gene activation by c-Myc.

ACKNOWLEDGMENTS

We thank T. Williams, A. Israël, J.-S. Seeler, and R. White for pPADH-AP2, AP-2 conaβ₂ CAT, GST-HP1, and some GST-RB constructs. We are very grateful to M. Buckingham, M. Yaniv, J. C. Reyes, and G. Butler Brown for critical reading of the manuscript.

This work was supported by grants from Association pour la Recherche sur le Cancer (6256), the Fondation pour la Recherche Médicale, and the GEFLUX. E.B. was supported by predoctoral fellowships from the Ministère de l’enseignement Supérieur et de la Recherche and from La Ligue contre le Cancer.