PITX2 and β-Catenin Interactions Regulate Lef-1 Isoform Expression

Abstract

Lef-1 and PITX2 function in the Wnt signaling pathway by recruiting and interacting with β-catenin to activate target genes. Chromatin immunoprecipitation (ChIP) assays identified the Lef-1 promoter as a PITX2 downstream target. Transgenic mice expressing LacZ driven by the 2.5-kb LEF-1 promoter demonstrated expression in the tooth epithelium correlated with endogenous Lef-1 FL epithelial expression. PITX2 isoforms regulate the LEF-1 promoter, and β-catenin synergistically enhanced activation of the LEF-1 promoter in combination with PITX2 and Lef-1 isoforms. PITX2 enhances endogenous expression of the full-length β-catenin-dependent Lef-1 isoform (Lef-1 FL) while decreasing expression of the N-terminally truncated β-catenin-independent isoform. Our research revealed a novel interaction between PITX2, Lef-1, and β-catenin in which the Lef-1 β-catenin binding domain is dispensable for its interaction with PITX2. PITX2 interacts with two sites within the Lef-1 protein. Furthermore, β-catenin interacts with the PITX2 homeodomain and Lef-1 interacts with the PITX2 C-terminal tail. Lef-1 and β-catenin interact simultaneously and independently with PITX2 through two different sites to regulate PITX2 transcriptional activity. These data support a role for PITX2 in cell proliferation, migration, and cell division through differential Lef-1 isoform expression and interactions with Lef-1 and β-catenin.

We thank Tord A. Hjalt (University of Lund, Lund, Sweden) for reagents, Jim Martin (IBT) and Jianbo Wang (IBT) for technical advice, and members of the Amendt laboratory for helpful discussions.

Support for this research was provided by grant NIH R37 DK047967 to J.F.E. and DE 13941 from the National Institute of Dental and Craniofacial Research to B.A.A.