Regulation of PCNA and Cyclin D1 Expression and Epithelial Morphogenesis by the ZO-1-Regulated Transcription Factor ZONAB/DbpA

Abstract

The tight junction protein ZO-1 inhibits G₁/S-phase transition by cytoplasmic sequestration of a complex formed by CDK4 and the transcription factor ZONAB. Canine ZONAB is the homologue of human DbpA, an E2F target gene that is overexpressed in different carcinomas. Since the ZONAB target genes that are involved in G₁/S-phase transition are unknown, we employed the mammary epithelial cell line MCF-10A and cDNA arrays to screen for such genes. We identified genes encoding cell cycle and replication proteins whose expression was altered due to increased ZONAB expression. For proliferative cell nuclear antigen and cyclin D1 genes, we show that increased mRNA levels resulted in increased protein levels and we identified ZONAB-responsive elements in their promoters by using different approaches, including chromatin immunoprecipitation assays. RNA interference and overexpression of ZONAB affected the proliferation of both MCF-10A and MDCK cells as well as the differentiation of MDCK cells into polarized cysts in three-dimensional cultures. These results indicate that ZONAB regulates the transcription of genes that are important for G₁/S-phase progression and links tight junctions to the transcriptional control of key cell cycle regulators and epithelial cell differentiation.

Supplemental material for this article may be found at http://mcb.asm.org/.

We thank Micheal B. Mathews for PCNA promoter plasmids and the members of our laboratories for discussions. We thank Parminder Mann for her work during the early stage of this project.

This research was supported by the Wellcome Trust (063661 and 066100), MRC, BBSRC and Fight for Sight. A.G. was supported by the European Union (Integrated Project EVI-GENORET: LSHG-CT-2005-512036).