Development of the skin epidermis and appendages such as hair follicles involves coordinated processes of keratinocyte proliferation and differentiation. The transcription factor p63 plays a critical role in these steps as evident by a complete lack of these structures in p63 null mice. The p63 gene encodes for two proteins TAp63 and DeltaNp63, the latter being the more prevalent and dominant isoform expressed in keratinocytes. Although numerous p63 target genes have been identified, these studies have been limited to transformed human keratinocyte cell lines. Here, we have employed a genomic screening approach of chromatin immunoprecipitation (ChIP) coupled with an enrichment strategy to identify DeltaNp63 response elements in primary mouse keratinocytes. Analysis of p63-ChIP-derived DNA segments has revealed more than 100 potential target genes including novel as well as mouse counterparts of established human p63 targets. Among these is Runx1, a transcription factor important for hair follicle development. We demonstrate that DeltaNp63 binds to a p63-response element located within a well-conserved enhancer of the Runx1 gene. Furthermore, siRNA mediated reduction of DeltaNp63 in mouse keratinocytes reduces Runx1 expression. Consistent with this, endogenous Runx1 levels are lower in the skin of p63(+/-) animals as compared to wild type animals. Lastly, we demonstrate that DeltaNp63 and Runx1 are co-expressed in specific compartments of the hair follicle in a dynamic fashion. Taken together our data demonstrate that p63 directly regulates Runx1 gene expression through a novel enhancer element and suggests a role for these two transcription factors in dictating skin keratinocyte and appendage development.
2008 Wiley-Liss, Inc.