Kaiso regulates Znf131-mediated transcriptional activation

Kaiso is a dual-specificity POZ-ZF transcription factor that regulates gene expression by binding to sequence-specific Kaiso binding sites (KBS) or methyl-CpG dinucleotide pairs. Kaiso was first identified as a binding partner for the epithelial cell adhesion regulator p120ctn. The p120ctn/Kaiso interaction is reminiscent of the β-catenin/TCF interaction and several studies have suggested that Kaiso is a negative regulator of the Wnt/β-catenin TCF signaling pathway. To gain further insight into Kaiso's function, we performed a yeast two-hybrid screen using the Kaiso POZ domain as bait. This screen identified the POZ-ZF protein, Znf131, as a Kaiso-specific binding partner. GST pull-down assays confirmed that the interaction is mediated via the POZ domain of each protein, and co-immunoprecipitation experiments further supported an in vivo Kaiso-Znf131 interaction. Using a Cyclic Amplification and Selection of Targets (CAST) approach, we identified the 12-base pair DNA palindrome sequence GTCGCR-(X)n-YGCGAC as a potential Znf131 binding element (ZBE). In vitro studies using electrophoretic mobility shift assay (EMSA) demonstrated that Znf131 binds the ZBE via its zinc finger domain. Znf131 DNA-binding specificity was confirmed using competition assays and ZBE mutational analyses. An artificial promoter–reporter construct containing four tandem copies of the ZBE was constructed and used to assess Znf131 transcriptional properties. We observed dose-dependent transcriptional activation of this artificial promoter–reporter by Znf131 in both epithelial and fibroblast cells, suggesting that Znf131 is a transcriptional activator. Kaiso overexpression significantly decreased the Znf131-mediated transcriptional activation, and interestingly, co-expression of the Kaiso-specific interaction partner p120ctn relieved Kaiso's inhibition of Znf131-mediated transcriptional activation. These findings indicate that Znf131 is a transcriptional activator, a less common function of POZ-ZF proteins, that is negatively regulated by its heterodimerization partner Kaiso.