Protein kinase PAK4 is involved in keratinocyte differentiation and modulates transcription factor p63

The epidermis is a multilayered, stratified epithelium continuously regenerated by keratinocytes through a terminal differentiation process called cornification. Recently, it has been shown that in epithelial cells PAK4 interacts with Keratinocyte Growth Factor (KGF) Receptor, and is activated following KGF or UV exposure. PAK4 is a member of the PAK family of serine/threonine kinases, originally identified as an effector protein of the RhoGTPase CDC42. While PAK4 is expressed at low levels in most adult tissues, it is found overexpressed in tumor cell lines and primary tumors, and its locus amplification is recurrent in many cancers, including squamous cell carcinoma. Considering that PAK4 is involved in many cellular processes necessary for proper development and homeostasis of epidermis, like cell adhesion, motility and control of cell death, and that its expression is increased during keratinocyte differentiation, in this work we evaluated PAK4 role in signalling mechanisms that control differentiation and survival in human keratinocytes. Using the RNA interference technique, we first showed that PAK4 down-regulation sensitizes HaCaT cells to UV induced apoptosis, confirming its pro-survival role also in keratinocytes. Secondly, we could demonstrate that PAK4 is necessary for the differentiation process induced by confluence growth: PAK4 knock-down cells show a loss of Keratin1 induction and impaired down-regulation of transcription factor p63, a master regulator of epithelial tissue differentiation. We then evaluated activated PAK4 effects on p63 stability and function. Our data show that active PAK4 can both modulate protein levels of the α and β isoforms of DNp63, and inhibit its trancriptional activity. Our results show that PAK4 plays a relevant role in differentiation of human keratinocytes, and suggest a novel p63 regulatory network that could be implicated in epithelial tissues pathologies.