Hypoxia induces accumulation of p53 protein, but activation of a G1-phase checkpoint by low-oxygen conditions is independent of p53 status.

It has been convincingly demonstrated that genotoxic stresses cause the accumulation of the tumor suppressor gene p53. One important consequence of increased p53 protein levels in response to DNA damage is the activation of a G1-phase cell cycle checkpoint. It has also been shown that G1-phase cell cycle checkpoints are activated in response to other stresses, such as lack of oxygen. Here we show that hypoxia and heat, agents that induce cellular stress primarily by inhibiting oxygen-dependent metabolism and denaturing proteins, respectively, also cause an increase in p53 protein levels. The p53 protein induced by heat is localized in the cytoplasm and forms a complex with the heat shock protein hsc70. The increase in nuclear p53 protein levels and DNA-binding activity and the induction of reporter gene constructs containing p53 binding sites following hypoxia occur in cells that are wild type for p53 but not in cells that possess mutant p53. However, unlike ionizing radiation, the accumulation of cells in G1 phase by hypoxia is not strictly dependent on wild-type p53 function. In addition, cells expressing the human papillomavirus E6 gene, which show increased degradation of p53 by ubiquitination and fail to accumulate p53 in response to DNA-damaging agents, do increase their p53 levels following heat and hypoxia. These results suggest that hypoxia is an example of a "nongenotoxic" stress which induces p53 activity by a different pathway than DNA-damaging agents.     

Depletion of the novel p53-target gene carnitine palmitoyltransferase 1C delays tumor growth in the neurofibromatosis type I tumor model

Despite the prominent pro-apoptotic role of p53, this protein has also been shown to promote cell survival in response to metabolic stress. However, the specific mechanism by which p53 protects cells from metabolic stress-induced death is unknown. Earlier we reported that carnitine palmitoyltransferase 1C (CPT1C), a brain-specific member of a family of mitochondria-associated enzymes that have a central role in fatty acid metabolism promotes cell survival and tumor growth. Unlike other members of the CPT family, the subcellular localization of CPT1C and its cellular function remains elusive. Here, we report that CPT1C is a novel p53-target gene with a bona fide p53-responsive element within the first intron. CPT1C is upregulated in vitro and in vivo in a p53-dependent manner. Interestingly, expression of CPT1C is induced by metabolic stress factors such as hypoxia and glucose deprivation in a p53 and AMP activated kinase-dependent manner. Furthermore, in a murine tumor model, depletion of Cpt1c leads to delayed tumor development and a striking increase in survival. Taken together, our results indicate that p53 protects cells from metabolic stress via induction of CPT1C and that CPT1C may have a crucial role in carcinogenesis. CPT1C may therefore represent an exciting new therapeutic target for the treatment of hypoxic and otherwise treatment-resistant tumors.     

p53: new roles in metabolism

Virtually all cancers show metabolic changes that result in upregulation of glycolysis and glucose consumption. Although discovered in the 1920s, how this glycolytic switch happens, and whether it is a cause or a consequence of the malignant process, has remained a matter of debate. The p53 tumor suppressor gene, discovered some 30 years ago, has a key role in preventing cancer development. Recent discoveries revealing new functions for p53 in the regulation of glucose metabolism and oxidative stress have brought together these two venerable fields of cancer biology. These activities of p53 appear to be key in tumor suppression, and shed some light on the pathways that underlie the metabolic changes in cancer cells.     

Activation of the p53 tumor suppressor protein

The p53 tumor suppressor gene plays an important role in preventing cancer development, by arresting or killing potential tumor cells. Mutations within the p53 gene, leading to the loss of p53 activity, are found in about half of all human cancers, while many of the tumors that retain wild type p53 carry mutations in the pathways that allow full activation of p53. In either case, the result is a defect in the ability to induce a p53 response in cells undergoing oncogenic stress. Significant advances have been made recently in our understanding of the molecular pathways through which p53 activity is regulated, bringing with them fresh possibilities for the design of cancer therapies based on reactivation of the p53 response.     

2-Deoxyglucose combined with wild-type p53 overexpression enhances cytotoxicity in human prostate cancer cells via oxidative stress

Overexpression of the tumor suppressor gene, wild-type p53 (wtp53), using adenoviral vectors (Adp53) has been suggested to kill cancer cells by hydroperoxide-mediated oxidative stress [1,2] and nutrient distress induced by the glucose analog, 2-deoxyglucose (2DG), has been suggested to enhance tumor cell killing by agents that induce oxidative stress via disrupting hydroperoxide metabolism [3,4]. In the current study clonogenic cell killing of PC-3 and DU-145 human prostate cancer cells (lacking functional p53) mediated by 4 h exposure to 50 plaque forming units (pfus)/cell of Adp53 (that caused the enforced overexpression of wtp53) was significantly enhanced by treatment with 2DG. Accumulation of glutathione disulfide was found to be significantly greater in both cell lines treated with 2DG+Adp53 and both cell lines treated with 2DG+Adp53 showed a approximately 2-fold increases in dihydroethidine (DHE) and 5-(and-6)-carboxy-2',7'-dichlorodihydrofluorescein diacetate (CDCFH(2)) oxidation, indicative of increased steady-state levels of O(2)(.-) and hydroperoxides, respectively. Finally, overexpression of catalase or glutathione peroxidase using adenoviral vectors partially, but significantly, protected DU-145 cells from the toxicity induced by 2DG+Adp53 treatment. These results show that treatment of human prostate cancer cells with the combination of 2DG (a nutrient stress) and overexpression of the tumor suppressor gene, wtp53, enhances clonogenic cell killing by a mechanism that involves oxidative stress as well as allowing for the speculation that inhibitors of glucose and hydroperoxide metabolism can be used in combination with Adp53 gene therapy to enhance therapeutic responses.     

Differential Impact of Tumor Suppressor Pathways on DNA Damage Response and Therapy-Induced Transformation in a Mouse Primary Cell Model

The RB and p53 tumor suppressors are mediators of DNA damage response, and compound inactivation of RB and p53 is a common occurrence in human cancers. Surprisingly, their cooperation in DNA damage signaling in relation to tumorigenesis and therapeutic response remains enigmatic. In the context of individuals with heritable retinoblastoma, there is a predilection for secondary tumor development, which has been associated with the use of radiation-therapy to treat the primary tumor. Furthermore, while germline mutations of the p53 gene are critical drivers for cancer predisposition syndromes, it is postulated that extrinsic stresses play a major role in promoting varying tumor spectrums and disease severities. In light of these studies, we examined the tumor suppressor functions of these proteins when challenged by exposure to therapeutic stress. To examine the cooperation of RB and p53 in tumorigenesis, and in response to therapy-induced DNA damage, a combination of genetic deletion and dominant negative strategies was employed. Results indicate that loss/inactivation of RB and p53 is not sufficient for cellular transformation. However, these proteins played distinct roles in response to therapy-induced DNA damage and subsequent tumorigenesis. Specifically, RB status was critical for cellular response to damage and senescence, irrespective of p53 function. Loss of RB resulted in a dramatic evolution of gene expression as a result of alterations in epigenetic programming. Critically, the observed changes in gene expression have been specifically associated with tumorigenesis, and RB-deficient, recurred cells displayed oncogenic characteristics, as well as increased resistance to subsequent challenge with discrete therapeutic agents. Taken together, these findings indicate that tumor suppressor functions of RB and p53 are particularly manifest when challenged by cellular stress. In the face of such challenge, RB is a critical suppressor of tumorigenesis beyond p53, and RB-deficiency could promote significant cellular evolution, ultimately contributing to a more aggressive disease.     

Regulation of p53: The next generation

The p53 protein is one of the most important tumor suppressor proteins. The most prevailing property of this tumor suppressor protein is its activation in response to DNA damage which counteracts the propagation of genetic alterations to daughter cells under conditions that provoke mutagenesis. In response to DNA damage and some other kinds of cellular stress the turnover of p53 is reduced or completely switched-off, which leads to a strong increase in the amount of the p53 protein and subsequently to the implementation of cell cycle arrest and apoptosis. Although post-translational modifications of p53 certainly contribute to the activation of p53 under physiologic conditions, an increase in the amount of the protein e.g. after overexpression, is sufficient for p53's deadly activities. This makes this tumor suppressor protein an interesting target for cancer therapy. This article summarizes the most important principles for the regulation of p53, with a particular focus on recent findings. Furthermore, open questions and possible future directions shall be discussed.     

The p53‐inducible long noncoding RNA TRINGS protects cancer cells from necrosis under glucose starvation

The tumor suppressor p53 is activated in response to cellular stress to prevent malignant transformation. However, several recent studies have shown that p53 can play protective roles in tumor cell survival under adversity. Whether p53‐regulated long noncoding RNAs are involved in this process remains to be fully understood. Here, we show that under glucose starvation condition, p53 directly upregulates a novel lncRNA named TRINGS (Tp53‐regulated inhibitor of necrosis under glucose starvation) in human tumor cells. TRINGS binds to STRAP and inhibits STRAP–GSK3β–NF‐κB necrotic signaling to protect tumor cells from cell death. Interestingly, TRINGS appears to respond to glucose starvation specifically, as it is not activated by serum, serine, or glutamine deprivation. Collectively, our findings reveal that p53‐induced lncRNA TRINGS controls the necrotic pathway and contributes to the survival of cancer cells harboring wild‐type p53 under glucose stress.     

Increased DNA damage sensitivity and apoptosis in cells lacking the Snf5/Ini1 subunit of the SWI/SNF chromatin remodeling complex

The gene encoding the SNF5/Ini1 core subunit of the SWI/SNF chromatin remodeling complex is a tumor suppressor in humans and mice, with an essential role in early embryonic development. To investigate further the function of this gene, we have generated a Cre/lox-conditional mouse line. We demonstrate that Snf5 deletion in primary fibroblasts impairs cell proliferation and survival without the expected derepression of most retinoblastoma protein-controlled, E2F-responsive genes. Furthermore, Snf5-deficient cells are hypersensitive to genotoxic stress, display increased aberrant mitotic features, and accumulate phosphorylated p53, leading to elevated expression of a specific subset of p53 target genes, suggesting a role for Snf5 in the DNA damage response. p53 inactivation does not rescue the proliferation defect caused by Snf5 deficiency but reduces apoptosis and strongly accelerates tumor formation in Snf5-heterozygous mice.