Role of Chromatin and Transcriptional Co-regulators in Mediating P63-genome Interactions in Keratinocytes

Background: The Transcription Factor (TF) p63 is a master regulator of epidermal development and differentiation as evident from the remarkable skin phenotype of p63 mouse knockouts. Furthermore, ectopic expression of p63 alone is sufficient to convert simple epithelium into stratified epithelial tissues in vivo and p63 is required for efficient transdifferentiation of fibroblasts into keratinocytes. However, little is known about the molecular mechanisms of p63 function, in particular how it selects its target sites in the genome. p63, which acts both as an activator and repressor of transcription, recognizes a canonical binding motif that occurs over 1 million times in the human genome. But, in human keratinocytes less than 12,000 of these sites are bound in vivo suggesting that underlying chromatin architecture and cooperating TFs mediate p63-genome interactions.

Results: We find that the chromatin architecture at p63-bound targets possess distinctive features and can be used to categorize p63 targets into proximal promoters (1%), enhancers (59%) and repressed or inactive (40%) regulatory elements. Our analysis shows that the chromatin modifications H3K4me1, H3K27me3, along with overall chromatin accessibility status can accurately predict bonafide p63-bound sites without a priori DNA sequence information. Interestingly, however there exists a qualitative correlation between the p63 binding motif and accessibility and H3K4me1 levels. Furthermore, we use a comprehensive in silico approach that leverages ENCODE data to identify several known TFs such as AP1, AP2 and novel TFs (RFX5 for e.g.) that can potentially cooperate with p63 to modulate its myriad biological functions in keratinocytes.

Conclusions: Our analysis shows that p63 bound genomic locations in keratinocytes are accessible, marked by active histone modifications, and co-targeted by other developmentally important transcriptional regulators. Collectively, our results suggest that p63 might actively remodel and/or influence chromatin dynamics at its target sites and in the process dictate its own DNA binding and possibly that of adjacent TFs.