Human TP63 was first isolated from human total genomic DNA by screening with an 800 bp PCR fragment obtained from probing with primers designed to anneal to regions in contiguous exons of p53 and p73, including the intervening introns (Yang et al., 1998). The resulting clones recovered from SK-N-MC neuroepithelioma total genomic DNA encoded proteins with two different N termini (TA and ΔN) and five different C termini (α, β, γ, δ, and ε; figure 1) (Yang et al., 1998; Mangiulli et al., 2009). Short splicing isoforms lacking exon 4 have been observed in invasive breast carcinomas (de Biase et al., 2010) that are not presented in figure 1.

The TP63 gene spans 267 kb, contains 16 exons, encodes 680 amino acids for the longest isoform, and has 12 isoforms formed by alternative promoter usage and alternative splicing (Augustin et al., 1998; Muzny et al., 2006; Parsons et al., 2009; Vanbokhoven et al., 2011). The six variants with TA and ΔN at the N-terminal and α, β, γ at the C-terminal are the most known and studied isoforms.

Human TP63 encodes the p63 protein, of which the longest isoform contains 680 amino acids, has molecular weights ranging from 44 to 77 kDa, depending on specific alternative promoter usage and alternative splicing (Augustin et al., 1998; Yang et al., 1998). TP63 comprises up to five types of domains, depending on the isoform, and may contain transactivation (TA), DNA Binding (DBD), oligomerization (OD), C-terminal sterile alpha motif (SAM), and/or C-terminal transcription inhibitory (ID) domains (Moll and Slade, 2004). The N-terminus may consist of a TA domain or a truncated version (ΔN) lacking the acidic TA domain that is derived from an alternative promoter and initiation codon in intron 3 (Yang et al., 1998). The 3 end of both TAp63 and ΔNp63 may be alternatively spliced to yield isoforms α, β, γ, and δ, while the ε-isoform is formed from transcriptional termination in exon 10 (Mangiulli et al., 2009). TAp63α and ΔNp63α contain a protein-protein interaction SAM domain and a trans-inhibitory domain, resulting from the masking of N-terminal TA domain residues. The δ- and ε-isoform proteins are formed by transcriptional termination in the second TA domain and after the OD, respectively (Mangiulli et al., 2009) (figure 1).

Structure
TP63 binds DNA as either a homo- or a heterotetramer, with isoform composition of the tetramer possibly determining transactivation activity. p63 may also form mixed dimers or tetramers with p73 at relatively higher affinity than with p53, suggesting functional cross talk to regulate transcriptional activity (Davison et al., 1999; Natan and Joerger, 2012). p63 appears to form a dimer of dimers, with monomers consisting of a β-strand followed by two helices (H1 and H2) that adopt a z-shaped double-hairpin conformation with little intramolecular contact between structural elements (Natan and Joerger, 2012). Monomers dimerize via intermolecular antiparallel β-sheet interactions and antiparallel packing of the H1 helices, with important hydrophobic contacts made by key leucine, valine, tyrosine, methionine, and isoleucine residues (Natan and Joerger, 2012). Tetramers are formed by hydrophobic H1-H1 interactions and H2-mediated contact where the H2 helices from the primary dimer clasps the adjacent dimer, packing the tetramer in an orthogonal fashion via H1 helices arrangement (Natan and Joerger, 2012). Analysis of the DBD of TP63 shows higher similarity to that of p73 than p53, and appears to bind a 10-bp DNA sequence containing a "CATG" motif with A/T-rich flanking regions (Chen et al., 2011).

TP63 expression is found in fetal and adult tissues, including the skin, cervix, vaginal epithelium, urothelium, prostate, heart, testis, kidney, thymus, brain, and spleen (Yang et al., 1998). Immunohistochemistry of p63 often shows strong nuclear-localized staining in basal epithelial cells. Additionally, TA and ΔN isoforms appear to be differentially expressed in particular tissue types. TAp63 variants are prevalent in the heart, testis, kidney, thymus, brain, and cerebellum. ΔNp63 transcripts are detected heavily in epithelial cells, kidney, spleen, and thymus, but not in the heart, liver, testis, or brain (Yang et al., 1998; Dötsch et al., 2010).

As a transcription factor, p63 is normally present in the nucleus (Yang et al., 1998).

TP63 acts as a motif-specific transcriptional activator or repressor, depending on the presence of TA or ID domains in the specific p63 isoform (Yang et al., 1998). It plays a critical role in the maintenance of progenitor-cell populations that encourage epithelial development and morphogenesis (Romano et al., 2009). ΔNp63, once thought to serve as a dominant negative regulator due to its lack of a full TA domain, has recently been implicated in transcriptionally activating and repressing target genes such as Keratin 5 and Keratin 14 to dictate early epithelial development and determine keratinocyte cell fate and lineage choices (Yang et al., 1998; Romano et al., 2009). Less is known about TAp63 function. Mice with germline deletion of TAp63 display blisters, decreased hair morphogenesis, and potential maintenance of adult stem cells, though full characterization of TAp63 function is lacking (Su et al., 2009).
While the interactomes of TP53 and TP73 have been systematically analyzed and cataloged, until recently, no such information had been compiled for TP63 (Tozluoglu et al., 2008; Collavin et al., 2010). Recent protein chip analysis has elucidated 144 proteins that specifically interact with TP63 and are implicated in cell growth/death/survival, chromatin remodeling and gene regulation, RNA processing, protein trafficking and degradation, and other and epithelial differentiation (Huang et al., 2012). A representative protein and its function from each of the aforementioned categories are described below, while a complete list may be found in the supplemental data of Huang et al. (2012).
Cell growth/survival: Signal Transducer and Activator of Transcription 3 (STAT3) - STAT3 is a latent cytoplasmic transcription factor activated in response to various interleukins and growth factors that binds the promoter regions of IL-6-induced plasma protein and intermediate-early genes (Akira et al., 1994; McLoughlin et al., 2005; Huang et al., 2012).
Chromatin remodeling: SWI/SNF Regulator of Chromatin 1 (SMARCC1) - SMARCC1 is a component of the larger SWI/SNF complex responsible for chromatin remodeling necessary for transcriptional activation of certain genes (Wang et al., 1996; Ring et al., 1998; Huang et al., 2012).
Gene regulation/expression: Eukaryotic Translation Initiation Factor 4A2 (EIF4A2) - EIF4A2 plays an important role in the binding of mRNA to the 43S pre-initiation complex during the beginning of protein synthesis (Sudo et al., 1995; Huang et al., 2012).
RNA processing: Splicing Factor 3b, Subunit 4 (SF3B4) - SF3B4 interacts with other spliceosome proteins to help cross-link a 29-nucleotide region in the pre-mRNA near the branch-point sequence in the A complex (Champion-Arnaud and Reed, 1994; Huang et al., 2012).
Protein trafficking: Trafficking Protein Particle Complex 2-Like (TRAPPC2L) - TRAPPC2L is part of the TRAPP multi-subunit tethering complex involved in intracellular vesicle trafficking (Scrivens et al., 2011; Huang et al., 2012).
Protein degradation: Ubiquitin Conjugation Factor E4 B Isoform 1 (UFD2A) - UFD2A has been shown to promote monoubiquitination of p53 and, in combination with MDM2, to promote p53 polyubiquitination (Wu and Leng, 2011; Wu et al., 2011; Huang et al., 2012).
Epithelial differentiation: Keratin 1 (KRT1) - KRT1 is a marker for terminal differentiation in the mammalian epidermis (Lessin et al., 1988; Huang et al., 2012).
Through protein-protein interactions with other transcription and cofactors, TP63 also contributes to the transcriptional regulation of genes involved in cellular differentiation, proliferation/survival, growth suppression, apoptosis, adhesion, inflammation, and metabolism (Perez and Pietenpol, 2007; Viganò and Mantovani, 2007; Yang et al., 2011). Recent work by our laboratory has identified protein-protein interactions between ΔNp63α, TAp73, and c-REL that function to regulate key genes involved in growth arrest and apoptosis of mutant p53 head and neck squamous cell carcinoma (HNSCC) (Lu et al., 2011). Functionally important genes representing each of the aforementioned categories are described below, while more comprehensive reviews may be found in Perez and Pietenpol (2007) and Viganò and Mantovani (2007).
Cellular differentiation: Jagged 1 (JAG1) - JAG1 is the ligand of the Notch receptor. Its binding causes a proteolytic cleavage cascade, leading to translocation of the intracellular component of the Notch receptor and subsequent activation of transcription factors with key roles in cell differentiation and morphogenesis (Gray et al., 1999; Sasaki et al., 2002; Guarnaccia et al., 2004). Overexpression of p63γ has been shown to dramatically upregulate JAG1 protein levels in colorectal cancer, osteogenic sarcoma, lung cancer, hepatocellular carcinoma, and glioma (Sasaki et al., 2002).
Proliferation/survival: Epidermal Growth Factor Receptor (EGFR) - EGFR is the receptor for epidermal growth factor and is involved in modulating cellular functions such as cell proliferation, differentiation, and survival by activating various intracellular signaling cascades such as RAS and STAT that transcribe target genes important in cellular proliferation and survival (Carpenter, 1984; Jamnongjit et al., 2005; Testoni et al., 2006). Knockdown of ΔNp63α has been shown to reduce expression of EGFR in keratinocytes (Testoni et al., 2006).
Growth suppression: Cyclin-Dependent Kinase Inhibitor 1A (CDKN1A) - CDKN1A (p21) associates with cyclins A, D, and E to prevent the G1-S phase transition in mammals (el-Deiry et al., 1993; Westfall et al., 2003). Overexpression of ΔNp63α has been shown to reduce expression of p21 in HEK cells (Westfall et al., 2003).
Apoptosis: p53-Upregulated Modulator of Apoptosis (PUMA) - PUMA binds to BCL2 to induce rapid and profound apoptosis by cytochrome c release (Yu et al., 2001; Flores et al., 2002). Additionally, recent work has shown that a complex of ΔNp63α, with TAp73 or c-REL, can modify expression of key growth arrest and apoptotic genes such as p21WAF1, NOXA, and PUMA (Lu et al., 2011; Yang et al., 2011).
Adhesion: Integrin Alpha 6 (ITGA6) - ITGA6 is a cell surface adhesion molecule that may help regulate migration and layer formation, especially in epithelial cells. Mouse models deficient in ITGA6 display severe blistering of the skin and other epithelia and die shortly after birth (Georges-Labouesse et al., 1996; Carroll et al., 2006). Loss of endogenous p63 in mammary epithelial cells has been shown to induce detachment and cell death, presumably because p63 regulates expression of key adhesion molecules such as ITGA6, ITGB1, ITGB4 other ECM components, and cadherins-catenins (Carroll et al., 2006; Yang et al., 2011).
Inflammation: ΔNp63 overexpression has been observed in head and neck squamous cell carcinoma (HNSCC), which are associated with inflammation. TNF-α, a potent pro-inflammatory cytokine, promoted NF-κB, c-REL and RELA complexes with nuclear ΔNp63, to promote a broad-spectrum production of cytokines and chemokines (Lu et al., 2011; Yang et al., 2011). In addition, in squamous epithelia of ΔNp63α transgenic mice, severe inflammation, skin lesions and erythema were observed after ΔNp63 expression was induced. Microscopically, hyperplastic and hyperproliferative epidermis with diffuse infiltration of inflammatory cells in the dermis was observed (Yang et al., 2011). NF-κB family proteins, c-Rel and RelA, and numerous inflammatory cytokines and chemokines were significantly upregulated in ΔNp63 transgenic mice.
Metabolism: Fatty Acid Synthase (FASN) - FASN catalyzes the conversion of Acetyl-CoA and Malonyl-CoA into long-chain saturated fatty acids with the help of NADPH (Wakil, 1989; DErchia et al., 2006). ΔNp63α (and TAp73α) expression has also been shown to induce promoter and enhancer activity in human FASN gene by binding to p53 response elements (DErchia et al., 2006).
Mouse models TP63 knockout mice are born alive but have striking developmental defects including absent or truncated limbs, lack of stratified epithelia, and lack hair follicles, teeth, and mammary glands. Thus, TP63 is essential for epidermal-mesenchymal interactions during embryonic development, including regenerative limb proliferation, craniofacial and epithelial development, and differentiation of squamous epithelia (Yang et al., 1998; Mills et al., 1999; McKeon, 2004; Laurikkala et al., 2006). Recently, ΔNp63α transgenic mice have been developed using tissue-specific tetracycline inducible expression. Mice expressing ΔNp63α under a tissue-specific promoter (SPC) for lung epithelium exhibited Keratin 5 and 14 induction, trans-differentiation to an epidermal cell lineage, and squamous metaplasia. Overexpression of ΔNp63α under a K5 promoter in wild-type epidermis results in severe defects in hair follicle development and cycling, leading to severe hair loss and a depleted hair follicle stem-cell niche (Romano et al., 2009; Romano et al., 2010). In addition, ΔNp63α overexpression induced marked skin inflammation, and skin hyperplasia (Yang et al., 2011). A more detailed summary of the phenotypes of different p63 knockout and transgenic mouse models has recently been published (Vanbokhoven et al., 2011).

In addition to humans, the TP63 gene is conserved in chimpanzees, dogs, cows, mice, rats, chicken, and zebrafish, and evolutionary precursors have been detected in Cnidarians.