Identification of transcriptional targets of Wnt/β-catenin signaling in dermal papilla cells of human scalp hair follicles: EP2 is a novel transcriptional target of Wnt3a

BackgroundRecent studies showed that Wnt signaling through the β-catenin pathway (canonical Wnt signaling) act on mouse dermal papilla cells (DPCs) enabling hair follicles to keep growing.ObjectiveTo investigate whether human DPCs respond to canonical Wnt signaling and, if so, to identify target genes of Wnt/β-catenin pathway.MethodsCultured human DPCs were transiently transfected with the β-catenin responsive TCF reporter plasmid (pTopflash) and corresponding negative control reporter (pFopflash) to assess the activity of β-catenin signaling by Wnt3a (one of the canonical Wnts). Immunofluorescence staining was also performed to localize β-catenin in the presence or absence of Wnt3a. Microarray was carried out using Affymetrix gene chips. RT-PCR analysis and immunoblot were employed to verify microarray data. Cyclic AMP (cAMP) levels were measured using EIA assay after Wnt3a and PGE2 treatment in DPCs.ResultsWnt3a significantly stimulated the transcriptional activity of pTopflash but not pFopflash. In line with this, we identified a number of genes that are regulated by Wnt3a. Some of the differently expressed genes including EP2 were confirmed by RT-PCR analysis. Immunoblot further confirmed that EP2 protein is indeed increased by Wnt3a. DPCs pretreated with Wnt3a showed higher responsiveness to PGE2 as measured by cAMP levels.ConclusionsElucidation of the role of Wnt3a-regulated genes identified in this study including EP2 would help our understanding of hair-induction and maintenance of anagen phase.