DNp63a, implicated as an oncogene, is upregulated by activated Akt, part of a well-known cell survival pathway. Inhibition of Akt activation by phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and the presence of putative p63-binding sites in the pten promoter led us to investigate whether DNp63a regulates PTEN expression. Knockdown of DNp63a led to increases in PTEN levels and loss of activated Akt, while overexpression of DNp63a decreased PTEN levels and elevated active Akt. The repression of PTEN by DNp63a occurs independently of p53 status, as loss of DNp63a increases PTEN expression in cell lines with and without functional p53. In addition, decreased levels of DNp63a resulted in an increase in nuclear PTEN. Conversely, in vivo nuclear PTEN was absent in the proliferative basal layer of the epidermis where DNp63a expression is highest. Additionally, we show that in keratinocytes a balance between DNp63a and PTEN regulates Akt activation and maintains normal proliferation rates. This balance is disrupted in non-melanoma skin cancers through increased DNp63a levels, and could enhance proliferation and subsequent neoplastic development. Our studies show that DNp63a negatively regulates PTEN, thereby providing a feedback loop between PTEN, Akt and DNp63a, which has an integral role in skin cancer development. Cell Death and Differentiation (2011) 18, 1924-1933 doi:10.1038/cdd.2011; published online 3 June 2011The p53 transcription factor family consists of the tumor suppressor p53 and the homologous p63 and p73. Unlike p53, p63 is essential for normal epidermal stratification and the proliferative potential of the epithelial stem cells. 1,2 p63 exists as various isoforms with contrasting functions. 2 The TA isoforms (TAp63a, TAp63b and TAp63g) have a full-length N-terminal transactivation domain, whereas the DN isoforms (DNp63a, DNp63b and DNp63g) have a short but distinct transactivation domain. All isoforms have a DNA-binding domain that shares high homology with p53 that allows p63 proteins to bind to p53 DNA-binding sites. 3,4 Similar to p53, the TA isoforms of p63 and p73 can promote apoptosis and growth arrest through the induction of antiproliferative genes. In contrast, the DN isoforms have been shown to induce pro-survival genes and inhibit anti-proliferative genes. [4][5][6] Several studies indicate that DNp63a, the predominant isoform in adult tissue, may function as an oncogene as it can exert a dominant-negative effect over p53 and the TAp63 and TAp73 isoforms. 2 Additionally, DNp63a is frequently overexpressed in a variety of squamous cell (SCC) and basal cell carcinomas (BCCs). 7,8 The survival factor Akt can increase DNp63a levels and in turn, DNp63a protects against UV-B-induced apoptosis via Akt activation. 9,10 However, the mechanism behind the positive feedback loop between DNp63a and Akt has not been described. Akt activation can be negated by phosphatase and tensin homolog deleted on chromosome 10 (PTEN). PTEN dephosphorylates phosphatidylinositol 3,4,5-trisphosphate, thereby...

p63 and p73, members of the p53 family, have been shown to be functionally distinct from p53. Vitamin D receptor (VDR) is a ligand (vitamin D 3 )-dependent transcription factor, which is shown to play a major role in calcium homeostasis and keratinocyte differentiation. Vitamin D and its analogues in combination with DNA-damaging agents are extensively used for cancer chemotherapy. In this report, we examined whether p53 affects p63-mediated induction of VDR and studied the effect of DNA damage on VDR induction in p53 null cell lines. Our results demonstrate that p53 itself does not induce VDR expression, nor does it affect p63-mediated VDR induction in the cell lines tested in this study. Furthermore, we observed p53-independent activation of VDR upon DNA damage and associated the induction of VDR to p73. We have demonstrated that ectopic expression of various p73 isoforms can induce VDR expression. Inhibition of p73 in cells treated with DNA-damaging agents exhibited decreased VDR expression. Finally, we show that upon DNA damage, induction of VDR sensitizes the cells to vitamin D treatment. In conclusion, our results indicate that VDR is regulated by p63 and p73 and that the induction of VDR expression upon DNA damage is p73-dependent. p53, the most frequently altered gene in human cancers, controls multiple signaling pathways by regulating the genes involved in cell cycle arrest, apoptosis, DNA repair, cellular senescence, and inhibition of angiogenesis (1). Stabilization of p53 protein occurs in response to various stress stimuli including DNA damage, viral infection, or oncogenic activation (1, 2). Although the functional significance of p53 and its interacting proteins have been studied extensively, it was not until 1997 that the functional homologues of p53, namely p63 and p73, were discovered. Despite being structurally similar to p53, p63 and p73 were shown to be functionally more diverse and distinct from p53 (3, 4). p63 knock-out mice were born with severe developmental defects, which included lack of skin and various epithelial tissues, whereas p73 knock-out mice were born with neuronal defects as well as changes in sexual behavior (5, 6).Functional diversities associated with p63 and p73 are partly due to their ability to generate multiple transcripts. Differential promoter usage by both p63 and p73 results in the generation of either transactivation domain containing isoforms (TAp63 and TAp73) or NH 2 -terminally truncated isoforms (⌬Np63 and ⌬Np73) (7). Additionally, multiple carboxyl termini variants, ␣, ␤, and ␥ of p63 and ␣, ␤, ␥, ⑀, and ␦ of p73, are generated due to the differential splicing of COOH terminus. Due to their structural similarity with p53, TA isoforms of both p63 and p73 have been shown to activate various p53-responsive genes and promote cell cycle arrest and apoptosis (8, 9). On the contrary, elevated levels of ⌬Np63 and ⌬Np73 isoforms have been reported in several human cancers, and these isoforms have also been associated with the induction of various genes involved in prom...

TRAP is an 11 subunit RNA binding protein that regulates expression of genes involved in tryptophan biosynthesis and transport in Bacillus subtilis. TRAP is activated to bind RNA by binding up to 11 molecules of l-tryptophan in pockets formed by adjacent subunits. The precise mechanism by which tryptophan binding activates TRAP is not known. Thr30 is in the tryptophan binding pocket. A TRAP mutant in which Thr30 is substituted with Val (T30V) does not bind tryptophan but binds RNA constitutively, suggesting that Thr30 plays a key role in the activation mechanism. We have examined the effects of other substitutions of Thr30. TRAP proteins with small β-branched aliphatic side chains at residue 30 bind RNA constitutively, whereas those with a small polar side chain show tryptophan-dependent RNA binding. Several mutant proteins exhibited constitutive RNA binding that was enhanced by tryptophan. Although the tryptophan and RNA binding sites on TRAP are distinct and are separated by ∼7.5 Å, several substitutions of residues that interact with the bound RNA restored tryptophan binding to T30V TRAP. These observations support the hypothesis that conformational changes in TRAP relay information between the tryptophan and RNA binding sites of the protein.