The p53 tumor suppressor is the central component of a system that helps maintain the genetic identity of cells in multicellular organisms. The major task for p53 is to ensure that an individual cell's behavior conforms to the functional needs of the entire organism. Loss of function of p53 results in the removal of numerous restrictions imposed on a cell, restrictions that normally abrogate or limit its activities. The cell is then no longer bound by the rules of the multicellular organism: it starts to compete with its neighbors, giving rise to a genetically unstable and rapidly evolving population of autonomous cells. Impairment of p53-dependent mechanisms is a necessary precondition for cancer; once the function of p53 is lost, the probability of cancer development increases dramatically. We study the mechanisms by which p53 enforces the altruistic behavior of individual cells in a multicellular organism. It is widely accepted that the major function of p53 is either to restrict growth or to kill abnormal or damaged, stress-exposed cells.
Recently, we established that, in addition to its restrictive functions, p53 helps maintain genetic stability by protecting the genome against excessive oxidation by reactive oxygen species (ROS). Several of the genes regulated by p53 encode antioxidant proteins, including two members of the sestrin family. We found that the protein products of the SESN1 and SESN2 genes participate in controlling the function of peroxiredoxins. The peroxiredoxins are required for decomposition of H2O2, which not only is a harmful by-product of respiration but also is an important signaling molecule. H2O2 is produced in response to activation of membrane receptors, which results in oxidation and functional modulation of several redox-sensitive components of signal transduction machinery. Thus, the task of peroxiredoxins is to ensure the delivery of an oxidant signal to its target and to remove excess H2O2, thereby protecting against oxidative damage. We found that p53-regulated sestrins control the regeneration of overoxidized peroxiredoxins, thus regulating their activity.
Our further studies established that the antioxidant activity of p53 contributes substantially to its tumor suppressor function. Even under physiological conditions in response to a mild oxidative burden, p53 regulates sestrin levels, which means that p53 protects the genome not only in emergencies but also in everyday life. Inhibition of p53 by various treatments (overexpression of MDM2, inhibition by RNAi, expression of transdominant p53 mutants, and so forth) results in an increased intracellular level of ROS, excessive oxidation of DNA, and an increased mutation rate, which are all reversed by incubation with antioxidants or by overexpression of sestrins. We found that dietary supplementation with antioxidants can efficiently prevent the frequent occurrence of lymphomas observed in p53-knockout mice and can also significantly improve karyotype stability.
We also found that changes in the expression of sestrins occur during carcinogenesis, particularly during malignant transformation of cells with the Ras oncogene. Overexpression of Ras results in downregulation of sestrins, which contributes to elevation of the intracellular ROS level. The latter is recognized as a signal that induces p53, which contributes to the development of Ras-induced senescence. A mutation or other defect in p53 function is required to overcome the proliferation block after oncogenic activation of Ras. We observed a similar downregulation of sestrins in human lung carcinomas. The effect was accompanied by an increase in ROS and a substantial acceleration of xenograft growth in nude mice. Our studies indicate that downregulation of p53-modulated sestrins in human cancer might be an important factor in tumor progression.
To understand the mechanisms underlying antioxidant and signal transduction functions of p53-modulated sestrins, we are studying the interactions and relationships of sestrins with other proteins as well as changes in the intracellular localization of sestrin-containing protein complexes following treatment with oxidants after generation of a signal. We obtained evidence suggesting that p53-modulated sestrins can efficiently protect the nuclear compartment from ROS produced during physiological signaling, protection that contributes substantially to the p53-mediated control of genetic stability. We found that large protein complexes containing sestrins move to the nuclei shortly after the release of H2O2 and that the translocated nuclear complexes function as efficient peroxidases, thus protecting the genome and the transcription machinery from excessive oxidation.
In human cancer cells, the activity of the p53 tumor suppressor is known to be impaired by multiple mechanisms. However, in general, human cancer cells retain and even acquire high sensitivity to exogenous p53, which may lead to massive death of cancer cells. Therefore, reactivation of damaged p53-dependent mechanisms in cancer has potential as an anticancer therapy. One of our programs aims at reactivating p53 in human cancer cells through the use of small molecules. We have developed several reporter human carcinoma cell lines that measure the transcriptional activity of p53 by expressing a reporter gene under the control of a p53-inducuble promoter. Thus, untreated carcinoma cell lines do not express the reporter, whereas p53 reactivation can be quantitatively measured in a colorimetric reaction. We used the cell lines as readout systems in high-throughput screening of chemical libraries and isolated several classes of small molecules that reactivate the transcriptional activity of p53. In particular, we identified small molecules that rescue p53 in cervical carcinoma, where p53 is suppressed by a human papilloma virus gene product. In addition, we identified a small-molecule compound that rescues the p53 tumor suppressor pathway in human carcinomas bearing a mutated p53. The latter compound activates the p53 family member p73, which is frequently suppressed in cancer cells by binding to the abundantly expressed mutant p53. The compound releases the activity of p73, which results in massive apoptosis of cancer cells. We found that the compound displays highly specific toxicity toward mutant p53-expressing carcinoma cells, with little or no effect on normal cells. Further optimization of similar compounds could result in the development of novel classes of anticancer therapeutics.
Last updated September 2008
