Introduction: p53 – the first twenty years

The p53 protein was discovered 20 years ago, as a cellular protein tightly bound to the large T oncoprotein of the SV40 DNA tumour virus. Since then, research on p53 has developed in many exciting and sometimes unexpected directions. p53 is now known to be the product of a major tumour suppressor gene that is the most common target for genetic alterations in human cancer. The nonmutated wild-type p53 protein (wtp53) is often found within cells in a latent state and is activated in response to various intracellular and extracellular signals. Activation involves an increase in overall p53 protein levels, as well as qualitative changes in the protein. Upon activation, wtp53 can induce a variety of cellular responses, most notable among which are cell cycle arrest and apoptosis. To a great extent, these effects are mediated by the ability of p53 to activate specific target genes. In addition, the p53 protein itself possesses biochemical functions which may facilitate DNA repair as well as apoptosis. The role of p53 in normal development and particularly in carcinogenesis has been elucidated in depth through the use of mouse model systems. The insights provided by p53 research over the years are now beginning to be utilized towards better diagnosis, prognosis and treatment of cancer.     

p53 in embryonic development: maintaining a fine balance

In addition to its role as a tumour suppressor and cell-cycle checkpoint control protein, p53 has been implicated as an important protein in embryonic development. Despite the viability of most p53 null mice, evidence has accumulated that p53 may regulate differentiation and the response of embryonic cells to diverse environmental stresses. Moreover, it appears that maintenance of a fine balance of p53 protein levels within embryonic cells is important for optimal development. Inappropriate overexpression or underexpression of p53 can lead to embryonic lethality or increased risk of malformations. The p53 protein may utilize multiple functional activities in its regulation of developmental processes.     

Regulation of the cell cycle following DNA damage in normal and Ataxia telangiectasia cells

A proportion of the population is exposed to acute doses of ionizing radiation through medical treatment or occupational accidents, with little knowledge of the immedate effects. At the cellular level, ionizing radiation leads to the activation of a genetic program which enables the cell to increase its chances of survival and to minimize detrimental manifestations of radiation damage. Cytotoxic stress due to ionizing radiation causes genetic instability, alterations in the cell cycle, apoptosis, or necrosis. Alterations in the G1, S and G2 phases of the cell cycle coincide with improved survival and genome stability. The main cellular factors which are activated by DNA damage and interfere with the cell cycle controls are: p53, delaying the transition through the G1-S boundary; p21^WAF1/CIPI, preventing the entrance into S-phase; proliferating cell nuclear antigen (PCNA) and replication protein A (RPA), blocking DNA replication; and the p53 variant protein p53as together with the retinoblastoma protein (Rb), with less defined functions during the G2 phase of the cell cycle. By comparing a variety of radioresistant cell lines derived from radiosensitive ataxia talangiectasia cells with the parental cells, some essential mechanisms that allow cells to gain radioresistance have been identified. The results so far emphasise the importance of an adequate delay in the transition from G2 to M and the inhibition of DNA replication in the regulation of the cell cycle after exposure to ionizing radiation.     

Cisplatin-induced genes as potential markers for thyroid cancer

Despite the uncontested role of p53 in cycle arrest/cell death after cisplatin treatment, to date the question whether wild-type p53 confers a resistant or sensitive status on the cell is still a matter of debate. Isogenic and isophenotypic human thyroid papillary carcinoma cell line variants for p53 differently expressed cycle genes after cisplatin treatment. Seven genes (CDC6-related protein, CCNC, GAS1, TFDP2, MAPK10/JNK3, WEE1, RPA1) selected after expression on an Atlas human cell cycle array were analyzed by quantitative real-time PCR. While cisplatin treatment increased their expression in p53 wild-type cells it decreased it in cells with inactivated p53 and had no or less effect on cells with mutated p53. These results show that in a well-defined system, different alterations of p53 can lead to a different regulation of genes and hence to either resistance or sensitivity to cisplatin. Moreover for the first time, MAPK10/JNK3 was identified in human thyroid cells and tissue. Four of the genes (CDC6-related protein, CCNC, GAS1 and TFDP2) were decreased in human papillary carcinoma tissues. Relevance of these genes (especially a decrease in GAS1 in thyroid papillary carcinoma) in various malignant pathologies has already been shown. These genes may be explored as new markers in advanced thyroid cancer such as metastatic and anaplastic forms displaying p53 alterations.Received 26 July 2004; received after revision 14 September 2004; accepted 26 October 2004     

Calcium-mediated activation of PI3K and p53 leads to apoptosis in thyroid carcinoma cells

The molecular mechanism responsible for cadmium-induced cell death in thyroid cancer cells (FRO) is unknown. We demonstrated that apoptosis of FRO cells induced by cadmium was concentration and time dependent. Cadmium caused the rapid elevation of intracellular calcium and induced phosphorylation of Akt, p53, JNK, ERK and p38. Inhibition of PI3K/Akt attenuated the cadmium-induced apoptosis, but the inhibition of JNK inhibitor, ERK or p38 aggravated it, indicating that activation of PI3K/Akt was a pro-apoptosis signal in response to cadmium treatment, whereas the activation of stress-activated protein kinase JNK, ERK and p38 functioned as survival signals to counteract the cadmium-induced apoptosis. Buffering of the calcium response attenuated mitochondrial impairment, recovered the cadmium-activated Akt, p53, JNK, ERK and p38, and subsequently blocked the apoptosis. These results suggested that apoptosis induced by cadmium in FRO cells was initiated by the rapid elevation of intracellular calcium, followed by calcium-mediated activation of PI3K/Akt and mitochondrial impairment. Received 28 February 2007; received after revision 2 April 2007; accepted 23 April 2007     

The dual role model for p53 in maintaining genomic integrity

The tumour suppressor p53 is a potent mediator of cellular responses against genotoxic insults. In this review we describe the multiple functions of p53 in response to DNA damage, with an emphasis on p53's role in DNA repair. We summarize data demonstrating that p53 actively participates in various processes of DNA repair and DNA recombination via its ability to interact with components of the repair and recombination machinery, and by its various biochemical activities. An important aspect in evaluating p53 functions is provided by the finding that the core domain of p53 harbours two mutually exclusive biochemical activities, sequence-specific DNA binding required for its transactivation function, and 3'-5' exonuclease activity, possibly involved in aspects of DNA repair. Based on the finding that modifications of p53 which lead to activation of its sequence-specific DNA-binding activity result in inactivation of its 3'-5' exonuclease activity, we propose that p53 exerts its functions as a 'guardian of the genome' at various levels: in its noninduced state, p53 should not be regarded as a 'dead' protein but, for example, via its exonuclease activity might be actively involved in prevention and repair of endogenous DNA damage. Upon induction through exogenous DNA damage, p53 will exert its well-documented functions as a superior response element in various types of cellular stress. This dual role model for p53 in maintaining genomic integrity significantly enhances p53's possibilities as a guardian of the genome.     

Kinetics of BRCA1 regulation in response to UVC radiation

To investigate changes in BRCA1 following DNA damage, we exposed MCF-7 cells to increasing doses of ultraviolet C. We observed an increase in BRCA1 protein levels above 78 J/m². This increase was observed as early as 5 min after irradiation. BRCA1 levels were then observed to decrease after 2 h, consistent with the previously published data. By pretreating with cycloheximide prior to irradiation, we observed a decrease in the protein half-life, from 3.5 h to 53 min, suggesting that a decrease in protein half-life may cause the lower levels of BRCA1 after irradiation. We also observed an increase in BRCA1 mRNA within 15 min of irradiation, followed by a decrease after 4 h. These data suggest that newly translated protein may contribute to increases in BRCA1 protein levels. The very rapid changes in BRCA1 support its role as a sensor of DNA damage, as opposed to being a repair gene. Received 6 April 2000; received after revision 23 May 2000; accepted 23 May 2000     

Cyclin A1 is a p53-induced gene that mediates apoptosis, G2/ M arrest, and mitotic catastrophe in renal, ovarian, and lung carcinoma cells

We were the first to identify cyclin A1 as a p53-induced gene by cDNA expression profiling of p53-sensitive and -resistant tumor cells [Maxwell S. A. and Davis G. E. (2000) Proc. Natl. Acad. Sci. USA 97, 13009–13014]. We show here that cyclin A1 can induce G2 cell cycle arrest, polyploidy, apoptosis, and mitotic catastrophe in H1299 non-small cell lung, TOV-21G ovarian, or 786-0 renal carcinoma cells. More cdk1 protein and kinase activities were observed in cyclin A1-induced cells than in GFP control-induced cells. Thus, cyclin A1 might mediate apoptosis and mitotic catastrophe through an unscheduled or inappropriate activation of cdk1. Two primary renal cell carcinomas expressing mutated p53 exhibited reduced or absent expression of cyclin A1 relative to the corresponding normal tissue. Moreover, renal carcinoma-derived mutant p53s were deficient in inducing cyclin A1 expression in p53-null cells. Cyclin A1 but not cyclin A2 was upregulated in etoposide-treated tumor cells undergoing p53-dependent apoptosis and mitotic catastrophe. Forced upregulation of cyclin A2 did not induce apoptosis. The data implicate cyclin A1 as a downstream player in p53-dependent apoptosis and G2 arrest. Received 1 November 2005; received after revision 17 February 2006; accepted 13 April 2006     

Nucleolar control of p53: a cellular Achilles’ heel and a target for cancer therapy

Nucleoli perform a crucial cell function, ribosome biogenesis, and of critical relevance to the subject of this review, they are also extremely sensitive to cellular stresses, which can cause loss of function and/or associated structural disruption. In recent years, we have learned that cells take advantage of this stress sensitivity of nucleoli, using them as stress sensors. One major protein regulated by this role of nucleoli is the tumor suppressor p53, which is activated in response to diverse cellular injuries in order to exert its onco-protective effects. Here we discuss a model of nucleolar regulation of p53, which proposes that key steps in the promotion of p53 degradation by the ubiquitin ligase MDM2 occur in nucleoli, thus providing an explanation for the observed link between nucleolar disruption and p53 stability. We review current evidence for this compartmentalization in p53 homeostasis and highlight current limitations of the model. Interestingly, a number of current chemotherapeutic agents capable of inducing a p53 response are likely to do so by targeting nucleolar functions and these compounds may serve to inform further improved therapeutic targeting of nucleoli.     

Cyclin D3 and p53 mediate sulforaphane-induced cell cycle delay and apoptosis in non-transformed human T lymphocytes

Despite experimental evidence that sulforaphane can exert chemopreventive effects, whether these effects are specific for neoplastic cells is not known. Following our previous demonstration that sulforaphane induces cell cycle arrest and apoptosis in human T lymphoblastoid Jurkat leukemia cells and increases p53 and bax protein expression, we tested sulforaphane on non-transformed phytohemagglutinin-stimulated human lymphocytes. Here, we demonstrate that sulforaphane arrested cell cycle progression in G, phase, through a decrease in the protein expression of cyclin D3. Moreover, sulforaphane induced apoptosis (and also necrosis), mediated by an increase in the expression of p53. These findings suggest that sulforaphane is a growth modulator for T cells. Our in vitro evidence that sulforaphane is active and even cytotoxic in normal as well as transformed lymphocytes raises important questions regarding its suitability for cancer chemoprevention.