Thesis presented June 24, 2010
Abstract: The outer layer of human skin, the epidermis, is a self-renewing, stratified squamous epithelial tissue. In adult tissue, the maintenance of epidermal homeostasis depends on an exquisite regulation of the balance between keratinocyte proliferation and differentiation. However, the specific molecular mechanisms governing each of these processes are not completely understood. p63 (TP63), a member of tumor suppressor p53 family, acts as a key regulator in controlling proliferation and differentiation balance in human keratinocytes. Although our knowledge of p63 was tremendously extended in the past years, little is known on the molecular mechanisms downstream of p63 that regulate the epidermal homeostasis. The identification of functional “effectors” acting downstream of p63 is an open question. By analyzing the transcriptional changes and phenotypic consequences of the loss of either p63 or MYC in human mature keratinocytes, I have characterized the networks acting downstream of these two genes to control epidermis homeostasis. I have shown that p63 is required to maintain growth and to commit to differentiation by two distinct mechanisms. Knockdown of p63 led to downregulation of MYC
via the Wnt/ß-catenin and Notch signaling pathways and in turn reduced keratinocyte proliferation. In addition, p63 controls the keratinocyte proliferation by directly repressing the transcription regulator of bHLH family, ID2. I demonstrated that a p63-controlled keratinocyte cell fate (KCF) network is essential to induce the onset of keratinocyte differentiation. This network contains several secreted proteins involved in cell migration/adhesion, including fibronectin 1 (FN1), interleukin 1 beta (IL1B), cysteine-rich protein 61 (CYR61), and jagged-1 (JAG1), that act downstream of p63 as key effectors to trigger differentiation. My findings establish for the first time a connection between p63 and MYC and show that the balance between MYC-controlled cell-cycle progression network and p63-controlled cell migration-related network dictate skin cell fate. Finally, I have characterized a set of miRNAs controlled by p63, and specified their roles in the onset of keratinocyte terminal differentiation. They target MAPKs, which have a strong effect on differentiation. Actually, inhibition of MAPKs was able to rescue the defects of differentiation caused by the loss of p63. My PhD work enabled the emergence of a general view on the “differentiation effectors”, both protein coding and non-coding small RNA genes, acting downstream of p63 to promote the commitment of human keratinocytes to terminal differentiation.
Keywords: P63, Keratinocytes, Cells, Proliferation, Differentiation
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