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During the 1960s, the field of cancer research lacked clear direction. Several facts appeared to be well-established and correct, but the relationships among these observations were not apparent. Fifty years of research had demonstrated that viruses with both DNA and RNA genomes could cause cancer in animals. Over the next 45 years six new viruses were to be discovered that were able to initiate cancers in humans (Epstein-Barr Virus, Human T-Cell Leukemia Virus, Hepatitis B and C Viruses, Kaposi Sarcoma Virus and the Papilloma Viruses) (McKinnel et al., 1998). It was equally clear from the perspective of the 1960s that certain chemicals, when applied to animals, were able to initiate cancers (Yamagawa et al., 1918). Chemical carcinogenesis was a field both separate and distinct (both in the experiments one did and the experimentalists who did them) from viral carcinogenesis and very few scientists thought to find a common ground between concepts generated in each field. Thirdly, the study of mouse genetics demonstrated that some cancers were clearly inherited and these observations confirmed many prior publications that suggested a role for cancer causing genes in humans and other animals (DeOme, 1965). Finally epidemiologists, studying a variety of important variables that predispose humans to developing cancers, had made the very striking observation that the rates of cancer incidence increase exponentially with age and begin to rise dramatically by the fifth and sixth decade of life (Miller, 1991). While these four observations were all accepted facts the relationship between these concepts was not clear and researchers who studied viruses hardly ever discussed chemicals and those who thought about genes and viruses didn’t know what to make of aging as an important variable. Literally researchers from each of these fields, virology, chemical carcinogenesis, genetics and epidemiology never got together to discuss these issues.

p53 is one of the most frequently mutated genes in human cancers and, as a result, is also one of the most well-studied genes in the history of cancer research. Although many functions have been ascribed to p53 over the years, one of the first activities to be characterized was the ability to bind DNA sequence-specifically through its central domain (reviewed in Vogelstein & Kinzler, 1992). This domain, also frequently referred to as “the core” due to its protease resistance (Bargonetti et al., 1993; Pavletich et al., 1993), contains the most evolutionarily conserved sequences of the protein, both between p53 proteins from different species and between the different p53 family members, p63 and p73 (reviewed in Yang et al., 2002). This region is also the most frequently mutated domain of p53 in the major forms of human cancer (Hainaut & Hollstein, 2000; Olivier et al., 2002). Consequently much research has focused on understanding this crucial ability as well as its regulation. Indeed, the regulation of p53 DNA binding has generated much debate recently, specifically with regard to the role of the C-terminus.

The short history of p53 contains an overwhelming number of facts and hypotheses, presenting the challenge of integrating diverse and sometimes mutually exclusive ideas into a coherent picture. It is important to make a distinction between p53 tumor suppressor activity, the mechanism of which remains speculative, and p53 responses to DNA damage, which are well characterized. Because critical steps in tumorigenesis involve genomic fixation of DNA damage-induced mutations, it seems reasonable to assume that DNA damage signaling to p53 would activate p53 tumor suppressor activity. However, this has not been demonstrated, and p53 tumor suppressor activity may not require the acute p53 response to DNA damage (Komarov et al., 1999). Nonetheless, the genotoxic chemicals and ionizing radiation that are clinically used to treat human cancer indisputably activate wild type p53.

Happy 25th Anniversary p53 ! Since this is such a special occasion, I (GW) thought of explaining how fate brought p53 and me together. It was a snowy day in Utah when Arnie Levine came to the University of Utah in 1976 to present a lecture on genetic approaches to differentiation of teratocarcinoma cells. My graduate work with Mario Capecchi (starting at Harvard and continuing at the University of Utah) led me to appreciate the potential power of genetics in cancer research. I therefore arranged to visit Arnie’s lab to learn more about his research program. We discussed many topics, but not about how the large transforming protein (T antigen) of SV40 (SV40TAg) interacted with a putative ~54kDα cellular protein (Linzer and Levine, 1979).

In response to abnormal proliferative signals and many forms of cellular stress including DNA damage and ribonucleotide depletion, p53 induces cells to undergo a transient arrest in G1 that is believed to allow time for repair of damaged DNA before the initiation of S phase. Failure to arrest in G1 can lead to chromosome aberrations and genomic instability. Activated p53 can also eliminate cells from the proliferative population through mechanisms that involve prolonged arrest in G1 (as seen during telomereinitiated replicative senescence and stress/DNA damage-induced premature senescence) and apoptosis (Levine, 1997; Oren, 2003; Vogelstein et al., 2000). The elimination of damaged, stressed or abnormally proliferating cells by p53 is considered to be the principal means by which p53 mediates tumour suppression (Symonds et al., 1994; Schmitt et al., 2002). Inappropriate or prolonged activation of p53 in normal tissues, however, can lead to tissue damage and has been associated with multiple sclerosis (Wosik et al., 2003), neurodegenerative disorders and exacerbation of ischemic damage from stroke or cardiac arrest (Mattson et al., 2001; Komarova and Gudkov, 2001). Accordingly, the regulation of p53 function is important for the maintenance of tissue homeostasis.

The p53 gene, first described in 1979, was the first tumor suppressor gene to be identified (Lane and Crawford, 1979; Linzer and Levine, 1979). It was originally identified as an oncogene- a cell cycle accelerator, but subsequent studies ten years after its discovery confirmed it to be a tumor suppressor gene that is highly mutated in a wide variety of tumors (Baker et al., 1990; Finlay et al., 1989). In about half of the tumors, p53 is inactivated directly as a result of mutations in the p53 gene. In many others, it is inactivated indirectly through binding to viral proteins, or as a result of alterations in the genes whose products interact with p53 or transmit information to or from p53. The tumor suppressor protein p53 acts as a major node in a complex signaling pathway that evolved to sense a broad range of cellular stresses such as DNA damage, oncogene activation, nucleotide depletion, and hypoxia (Figure 1). In the absence of cellular stress, the p53 protein is expressed at low steady-state levels and exerts little, if any, effect on cell fate.

The basis for p53’s striking apoptotic and tumor suppressive potency lies in its pleiotropism that includes transcription-dependent and –independent functions. p53 kills cells predominantly via the mitochondrial death pathway rather than the death receptor pathway (Schuler et al. 2001). p53 can mediate apoptosis by transcriptional activation of pro-apoptotic genes like the BH3- only proteins Noxa and Puma, Bax, p53 AIP1, Apaf-1, DRAL and PERP, and by transcriptional repression of Bcl2 and IAPs. For Noxa, Puma and PIDD, downregulation decreases - but does not abolish – the extent of death after stress. Of note, induction of these target gene products show variable kinetics, with some being delayed in their response (over 24 h), e.g. Bax and p53AIP1 (Attardi et al. 2000; Nakano et al. 2001). Analysis of p53-regulated global gene expression shows that the type, strength and kinetics of the target gene profiles depends on p53 levels, stress type and cell type (Zhao et al. 2000). This indicates that only individual genes will be chosen from the complex spectrum of potentially inducible genes to mediate a specific p53 response in a given physiological situation.

The early discoveries elucidating p53 function were based on cell culture experiments. Most of our fundamental knowledge of the role of p53 in cell signaling, stress response, cell cycle control, and apoptosis are a result of these in vitro studies (Giaccia and Kastan, 1998; Ko and Prives, 1996; Levine, 1997; Vogelstein et al., 2000). However, a greater depth of understanding was facilitated by the advent first of transgenic mouse methodologies and then by embryonic stem (ES) cell-based genetic manipulations. The sequencing of the mouse genome (www.ensembl.org and www.myscience.appliedbiosystems.com) has greatly simplified and accelerated the generation of null alleles. Methods have been developed to generate single nucleotide substitutions in the germline of mice, and importantly, to generate somatic mutations in genes to study somatic inactivation as occurs in most human cancers. The availability of whole genome analysis at the RNA expression level (arrays) and at the genomic level (array CGH) provides another level of analysis that is sure to provide insights into the molecular changes that lead to the initiation, progression, and maintenance of the tumor phenotype.

The discoveries of p63 and p73 as genes related to the vaunted tumor suppressor p53 launched questions that remain largely unanswered, and fuel controversy and debate. Do these homologs behave like p53? Do they also act in tumor suppression? What were their origins – spin-offs of an ancestral p53 gene, or, in fact, predecessors of this famed ‘guardian of the genome’? In vivo studies clearly reveal distinct physiological roles for p53, p63, and p73. But do these belie cooperative or antagonistic interactions within the p53 gene family?

Approximately half of human tumors bear p53 mutations (Hollestein et al., 1997). The most prevalent type consists of missense mutations that are frequently accompanied by loss of the remaining wild-type p53 (wt-p53) allele (Hainaut et al., 1997; Levine, 1997). The major site of the p53 mutations is the highly conserved DNA binding core domain (Hussain et al., 1998; Prives et al., 1999). Thus, mutant p53 (mt-p53) proteins are unable to specifically bind DNA and to activate specific wt-p53 target genes. Unlike wt-p53, whose half-life is short, mutant p53 proteins are quite stable and abundantly present in cancer cells. One certain outcome of p53 mutations is the loss of wild type activities such as growth arrest, apoptosis, and differentiation (Michalovitz et al., 1990; Yonish-Rouach et al., 1991; Soddu et al., 1996; Almog et al., 1997). However, at variance with other tumor suppressor genes, cells with p53 mutations maintain expression of the fulllength protein. This may suggest that, at least certain mutant forms of p53 can gain additional functions through which actively contribute to cancer progression (Prives et al., 1999; Sigal et al., 2000; Strano et al., 2001; Bullock et al., 2001). Such evidence is provided by several in vitro and in vivo studies (Haley et al., 1990; Dittmer et al., 1993; Gualberto et al., 1998; Frazier et al., 1998; Li et al., 1998; Blandino et al., 1999; Aas et al., 1996; Irwin et al., 2003; Strano et al., 2003)