Clinical implications of p53 mutations

The ultimate goal of basic cancer research is to provide a theoretical foundation for rational approaches to improve cancer therapy. Our extensive insight into the biology of the p53 tumour suppressor and the clinical behaviour of tumours harbouring p53 mutations indicates that information concerning p53 will be useful in diagnosis and prognosis, and may ultimately produce new therapeutic strategies. At the same time, efforts to understand the clinical implications of p53 mutations have revealed conceptual and technical limitations in translating basic biology to the clinic. The lessons learned from p53 may lay the groundwork for future efforts to synthesize cancer gene function, cancer genetics and cancer therapy.     

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.     

Covalent and noncovalent modifiers of the p53 protein

Despite the massive attention it has received, there is still much to learn about the p53 tumour suppressor protein. Given that it plays complex and multiple roles in cells, it is not surprising that p53 is subjected to an intricate array of regulatory processes. p53 receives signals from cells in multiple ways, leading to its stabilization and activation. The functions of the protein are altered by phosphorylation and other covalent modifications. However, a number of proteins can regulate p53 function dramatically by noncovalent means. p53 is thus subjected to numerous signalling and regulatory pathways which we have only begun to decipher.     

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.     

Isoform specific phosphorylation of p53 by protein kinase CK1

The ability of three isoforms of protein kinase CK1 (α, γ₁, and δ) to phosphorylate the N-terminal region of p53 has been assessed using either recombinant p53 or a synthetic peptide reproducing its 1–28 sequence. Both substrates are readily phosphoylated by CK1δ and CK1α, but not by the γ isoform. Affinity of full size p53 for CK1 is 3 orders of magnitude higher than that of its N-terminal peptide (K _m 0.82 μM vs 1.51 mM). The preferred target is S20, whose phosphorylation critically relies on E17, while S6 is unaffected despite displaying the same consensus (E-x-x-S). Our data support the concept that non-primed phosphorylation of p53 by CK1 is an isoform-specific reaction preferentially affecting S20 by a mechanism which is grounded both on a local consensus and on a remote docking site mapped to the K²²¹RQK²²⁴ loop according to modeling and mutational analysis.     

Progesterone inhibits human endothelial cell proliferation through a p53-dependent pathway

Previous studies have shown that progesterone inhibits endothelial cell proliferation through a nuclear receptor-mediated mechanism. Here, we further demonstrate that progesterone at physiologic levels (5 – 500 nM) dose- and time-dependently inhibited DNA synthesis of cultured human umbilical vein endothelial cells (HUVEC). The mRNA and protein levels of p21, p27, and p53 in HUVEC were increased by progesterone. The formation of CDK2-p21 and CDK2-p27 were increased and the CDK2 activity was decreased in the progesterone-treated HUVEC. The progesterone-inhibited [3H]thymidine incorporation was completely blocked when the expressions of p21 and p27 were knocked-down together. Transfection of HUVEC with dominant negative p53 cDNA prevented the progesterone-induced increases in p21 and p27 promoter activity and protein level, decreases in thymidine incorporation, and capillary-like tube formation. Matrigel angiogenesis assay in mice demonstrated the antiangiogenic effect of progesterone in vivo. These findings demonstrate for the first time that progesterone inhibited endothelial cell proliferation through a p53-dependent pathway. Received 28 July 2008; received after revision 25 September 2008; accepted 26 September 2008     

Developing pharmacological therapies for Alzheimer disease

Alzheimer disease (AD), while chronic and progressive with an average progression of 7 – 10 years, is both multifactorial and heterogeneous. Thus, AD offers a large window of opportunity and a large number of therapeutic targets to inhibit it. The selection of a therapeutic target, however, is one of the biggest challenges in developing a pharmacological treatment of this multifactorial disease. Inhibition of a pivotal downstream event is likely to benefit more patients than inhibition of an upstream event in AD pathogenesis. Neurofibrillary degeneration of abnormally hyperphosphorylated tau offers such a pivotal therapeutic target. Abnormal hyperphosphorylation of tau and not its aggregation into filaments appears to be the most deleterious step in neurofibrillary degeneration. Tau can be abnormally hyperphosphorylated by downregulation of protein phosphatase-2A activity or by upregulation of more than one tau kinase. Restoration of the phosphatase activity which is downregulated in AD brain or inhibition of GSK-3β and cdk5, which are required for AD-type abnormal hyperphosphorylation of tau, are among the most promising therapeutic strategies.     

Differential display analysis of gene expression in mammals: a p53 story

Differential display is used worldwide as a method to identify changes in gene expression and to discover novel genes that are involved in important biological pathways. The principle of differential display is the systematic amplification of the 3' termini of messenger RNAs by using anchored oligo-dT primers in combination with upstream arbitrary primers. The separation of the polymerase chain reaction products by gel electrophoresis and their direct comparison allows the identification of differentially regulated genes. Recently, fluorescent differential display was established as the first nonradioactive differential display system with equivalent sensitivity to originally 33P isotopic labeling method. Because of its simplicity, sensitivity, reproducibility and automation, which increase the throughput and accuracy, differential display has become one of the most widely used gene-screening methods in biomedical research involving mammals. This chapter provides a glimpse of the application of differential display in search of target genes of the p53 tumor suppressor gene.     

The role of p53 in tumour suppression: lessons from mouse models

The use of mouse models has greatly contributed to our understanding of the role of p53 in tumour suppression. Mice homozygous for a deletion in the p53 gene develop tumours at high frequency, providing essential evidence for the importance of p53 as a tumour suppressor. Additionally, crossing these knockout mice or transgenic expression p53 dominant negative alleles with other tumour-prone mouse strains has allowed the effect of p53 loss on tumour development to be examined further. In a variety of mouse models, absence of p53 facilitates tumorigenesis, thus providing a means to study how the lack of p53 enhances tumour development and to define genetic pathways of p53 action. Depending on the particular model system, loss of p53 either results in deregulated cell-cylce entry or aberrant apoptosis (programmed cell death), confirming results found in cell culture systems and providing insight into in vitro function of p53. Finally, as p53 null mice rapidly develop tumours, they are useful for evaluating agents for either chemopreventative or therapeutic activities.     

Engineering microbes for targeted strikes against human pathogens

Lack of pathogen specificity in antimicrobial therapy causes non-discriminant microbial cell killing that disrupts the microflora present. As a result, potentially helpful microbial cells are killed along with the pathogen, altering the biodiversity and dynamic interactions within the population. Moreover, the unwarranted exposure of antibiotics to microbes increases the likelihood of developing resistance and perpetuates the emergence of multidrug resistance. Synthetic biology offers an alternative solution where specificity can be conferred to reduce the non-specific, non-targeted activity of currently available antibiotics, and instead provides targeted therapy against specific pathogens and minimising collateral damage to the host’s inherent microbiota. With a greater understanding of the microbiome and the available genetic engineering tools for microbial cells, it is possible to devise antimicrobial strategies for novel antimicrobial therapy that are able to precisely and selectively remove infectious pathogens. Herein, we review the strategies developed by unlocking some of the natural mechanisms used by the microbes and how these may be utilised in targeted antimicrobial therapy, with the promise of reducing the current global bane of multidrug antimicrobial resistance.     

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     

亚洲人群p53Arg72Pro基因多态性与胃癌易感性的Meta分析

目的探讨p53基因多态性与亚洲人群胃癌易感性的关系。方法检索PubMed、EMbase、The Cochrane Library和CBM、中国学术文献总库、维普、万方数据库,收集有关p53基因多态性与亚洲人群胃癌易感性有关的文献,最后检索日期为2014年5月,并追溯纳入研究的参考文献。由两位研究者按照纳入与排除标准独立筛选文献、提取资料和评价质量后,采用Stata13.0统计软件进行统计分析。以病例组及对照组p53基因型分布的比值比(OR值)为效应指标,计算合并OR值及95%可信区间,用漏斗图和Egger图评估。结果在亚洲人群中,病例组和对照组相比,携带杂合基因型(Arg/Pro)和突变纯合基因型(Pro/Pro)的个体发生胃癌的风险和野生纯合型(Arg/Arg)相比差异有统计学意义(OR=1.159,95%CI:1.025~1.331,P=0.019)(P0.05)。结论与野生纯合型相比,携带杂合基因型和突变纯合基因型增加了患胃癌的风险。