Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P43146 (tumour suppressor)
 5,935 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Epstein-Barr virus (EBV) efficiently converts resting human B cells into actively cycling, immortal, lymphoblastoid cell lines (LCLs). Here we show that LCLs expressing the full complement of latent viral genes are very sensitive to DNA-damaging agents such as cisplatin. The response includes a rapid accumulation of the tumour suppressor protein p53 and induction of the cellular genes mdm2 and WAF1/p21. Although the levels of Bcl2 protein and Bax mRNA appear unaltered by the activation of p53, within 24 h the majority of cells undergo apoptosis. Over-expression of wild-type p53 in an LCL also resulted in apoptosis; this was preceded by the dephosphorylation of the retinoblastoma gene product, pRb. Primary resting B cells showed no response to cisplatin and even after drug treatment, p53 remained undetectable. However, after infection with EBV, p53 gene expression was induced to a similar level to that found in mitogen-activated B cells. When the physiologically activated primary B cells were exposed to cisplatin, although p53 accumulated as in LCLs, the outcome was growth-arrest rather than gross cell death. We conclude that, in contrast to the transformation of fibroblasts by adenovirus, SV40 or HPV, when B cells become activated and immortalized by EBV they are sensitized to the p53-mediated damage response. When the resulting LCLs are treated with genotoxic agents such as cisplatin, they are unable to arrest like normal cells because they are driven to proliferate by EBV and consequently undergo apoptosis.
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PMID:Epstein-Barr virus efficiently immortalizes human B cells without neutralizing the function of p53. 772 16

The tumour suppressor protein p53 normally functions as a tetramer in a defined conformational state. Mutations within p53 which contribute to cancer development frequently induce a conformational shift in the protein which correlates with loss of wild type growth suppressor functions. Both the cell encoded mdm2 protein and the human papillomavirus oncoprotein E6 can regulate p53 function and we have examined the interaction of these proteins with p53. The E6/p53 association is sensitive to conformational alterations in the p53 protein, although oligomerisation is not necessary for this interaction to occur. Analysis of C-terminal p53 truncations has indicated that the region between residues 327 and 347 may play a role in E6 binding. Since monomeric forms of p53 retain transcriptional and transformation suppressor activities, our results indicate that E6 targets p53 proteins which retain these wild type functions. Conversely, the interaction of p53 with mdm2 is not dependent on the conformation of the p53 protein but is significantly impaired by loss of quaternary structure. It is possible that mdm2 plays a role in mediating activities of p53 which, unlike transcriptional activation, depend on oligomerisation.
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PMID:Oligomerisation of full length p53 contributes to the interaction with mdm2 but not HPV E6. 775 47

The oncogene mdm2 and its human homologue hdm2 bind to the tumour suppressor protein p53 and inactivate its function as a transcription factor. This has been implied as a possible mechanism for cancer development in several tumours including human sarcomas. The mdm2-p53 interaction is therefore a much persued target for the development of anti-cancer drugs. In order to find novel high affinity ligands for hdm2 which would interfere with its binding to p53 we screened phage display peptide libraries for mdm2 binding phage. We found a series of 12 and 15mer peptides which interact strongly with hdm2. The peptide sequences show striking homology with the previously established mdm2 binding site on p53, confirming that the peptide defined 18TFSDLW23 region is crucial for the interaction but that contact between the two molecules extends to position L26 on p53. Free synthetic peptides derived from the phage selected sequences proved to be up to 100 times stronger inhibitors of the p53-mdm2 interaction than the p53 derived wt-peptide in several ELISA-assays. This illustrates the potency of phage display libraries in the search for new peptide based lead structures designed to mimic or inhibit therapeutically important protein-protein interactions.
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PMID:Identification of novel mdm2 binding peptides by phage display. 895 Sep 81

Wild-type p53 (wtp53) is a tumour suppressor gene involved in cell cycle regulation. The mdm2 protein can complex with the p53 protein and influence its function as a regulator of cell growth. To detect and quantify wtp53 and mdm2 mRNA expression, we established the competitive reverse transcription/polymerase chain reaction for these genes and for the housekeeping gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The target RNA differed from the competitor cRNA by having 183 bp, 205 bp and 173 bp deletions for p53, mdm2 and GAPDH, respectively. Target RNA and known concentrations of competitor cRNA were co-reverse transcribed and co-amplified with the same primers. Target cDNA and the corresponding competitor cDNA were amplified at the same efficiency.
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PMID:Competitive reverse transcription/polymerase chain reaction for the quantification of p53 and mdm2 mRNA expression. 902 80

A number of viral oncogenes target the tumour suppressor protein p53 and inactivate its function. This is an important step in tumourogenesis. The cellular oncogene hdm2 acts through a similar mechanism. It binds the N terminus of p53, thereby interfering with the ability of p53 transcriptionally to activate genes responsible for growth arrest or apoptosis after genotoxic insults. The disruption of the interaction of the two proteins therefore comprises a promising therapeutic target for treatment of the subset of human cancers in which this pathway is active. In this paper we attempt to characterize the p53-hdm2 interaction biochemically. We analyse the potential of a series of peptide inhibitors, derived from previously described mdm2 binding peptide display phage, to disrupt this interaction in ELISA assays. We conclude that F19, W23 and L26 of p53 are critical contact points for p53 binding to hdm2. Furthermore, we show the potential of the monoclonal antibody 3G5 to interfere with binding of p53 to hdm2 in ELISA assays. Consequently, we define the binding site of 3G5 on hdm2 using overlapping peptides derived from the N terminus of hdm2 and phage display libraries. The result indicates L66, Y67 and E69 on hdm2 as critical binding points for 3G5. In electrophoretic mobility shift assay we demonstrate the formation of hdm2-p53 complexes that can be disrupted in the presence of 3G5 or inhibitory peptides. Finally, we describe the effects of NEM and DTT on the interaction between the two molecules in ELISA assays. All our results are discussed in the light of the recently published crystal structure of the mdm2-p53 complex. A striking correspondence between our findings and the crystal structure is revealed.
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PMID:Molecular characterization of the hdm2-p53 interaction. 922 38

Loss of function of the tumour suppressor gene p53 is a key event in most human cancers. Although usually occurring through mutation, in some tumour types this appears to be achieved via an indirect mechanism involving inappropriate expression of a functional inhibitor, mdm2, which binds to the transactivation domain of p53. This interaction offers an ideal potential target for novel cancer therapies. However, therapeutic specificity may depend on the extent to which this p53-inhibitory action of mdm2 is also required by normal cells. Transgenic data have already established that mdm2 is needed to prevent embryonic lethality, but the situation in adult cells is still unclear. Here we show that micro-injection of normal human fibroblasts with an antibody directed against the p53-binding domain of mdm2 induces expression of p53-responsive genes, and furthermore results in p53-dependent growth arrest. We conclude that normal cell proliferation can be dependent on negative regulation of p53 by mdm2, a finding which raises an important note of caution for mdm2-directed cancer therapies.
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PMID:The proliferation of normal human fibroblasts is dependent upon negative regulation of p53 function by mdm2. 968 31

The p53 tumour suppressor protein is a transcriptional activator, which can induce cell cycle arrest and apoptosis. p53 Gene mutations occur in more than 50% of all human tumours. Reintroduction of wild-type p53 but also of oligomerisation-independent p53 variants into tumour cells by gene transfer methods has been considered. We have investigated the biological properties of two carboxy-terminal deletion mutants of p53, p53 delta 300 (comprising amino acids 1-300) and p53 delta 326 (amino acids 1-326), to evaluate their potential deployment in gene therapy. Transactivation was measured in transiently transfected HeLa and SKBR3 cells. Both monomeric variants showed reduced activities compared with wild-type p53. Individual promoters were differently affected. In contrast to wild-type p53, monomeric variants were not able to induce apoptosis. We also provided wild-type p53 and p53 delta 326 with tetracycline-regulated promoters and stably introduced these constructs into Saos2 and SKBR3 cells. Upon induction, wild-type p53 expressing cells, but not p53 delta 326 expressing cells underwent apoptosis. Consistently, only wild-type p53 expressing cells accumulated p21/waf1/cip1 mRNA and protein and showed increased bax, Gadd45 and mdm2 mRNA. Neither wild-type p53 nor p53 delta 326 repressed the transcription of the IGF-1R gene in these cell lines. We conclude that the transactivation potential of monomeric, carboxy-terminally truncated p53 is not sufficient to cause induction of the endogenous target genes which trigger apoptosis.
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PMID:Transcriptional regulation and induction of apoptosis: implications for the use of monomeric p53 variants in gene therapy. 1002 42

Abnormalities of the p53 tumour suppressor gene are among the most frequent molecular events in human and animal neoplasia. Moreover, p53 is one of the most studied proteins in the whole of contemporary biology, with more than 12,500 papers so far written! In this review the choice has been deliberately made not to be fully comprehensive in the coverage of the huge p53 literature. Rather attention is focused on a small number of recent developments which are reviewed in the context of modern models of p53 function. Progress in the analysis of signalling to p53 including phosphorylation cascades, and interactions with proteins such as mdm2 and ARF are highlighted. The plethora of protein-protein interactions is discussed, as are the strategies for defining downstream targets of p53. Finally, the emerging biology of p53 homologues is considered. The need for bridging the gap between reductionist, biochemical and biophysical studies and biological and genetic analysis is emphasized. Only this will provide the needed framework for utilizing the information in clinical care.
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PMID:The p53 pathway. 1034 12

The transcriptional activity of the p53 tumour suppressor is inhibited by binding to MDM2. The in vivo significance of this interaction was established in mdm2 null mice. Embryonic lethality due to loss of mdm2 is completely rescued by deletion of p53, indicating that the lethality is due to inability to down-modulate p53 function. The production of mice null for both p53 and mdm2 led to an assessment of the role of MDM2 in tumour development. Tumour latency and spectrum in p53 null mice were monitored in the presence or absence of mdm2. Two unusual findings resulted: tumour latency in p53 null/mdm2 heterozygous mice was longer than in p53/mdm2 double-null mice; and the incidence of sarcomas was higher in p53 null/mdm2 heterozygous mice than in p53 null or p53/mdm2 double-null mice. These data raise the possibility that heterozygosity at the mdm2 locus in the absence of p53 affects the development of tumours of mesenchymal origin.
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PMID:Loss of one but not two mdm2 null alleles alters the tumour spectrum in p53 null mice. 1041 3

The ability to separate the isoforms of human tumour suppressor protein p53 expressed in insect cells using heparin-Sepharose correlates with differences in the isoelectric point of p53, demonstrating that p53 can be heterogeneously modified and providing support for the use of insect cells as a model system for identifying novel signalling pathways that target p53. One p53 isoform that was reduced in its binding to the monoclonal antibody DO-1 could be stimulated in its binding to DO-1 by prior incubation with protein phosphatases, suggesting the presence of a previously unidentified N-terminal phosphorylation site capable of masking the DO-1 epitope. A synthetic peptide from the N-terminal domain of p53 containing phosphate at Ser(20) inhibited DO-1 binding, thus identifying the phosphorylation site responsible for DO-1 epitope masking. Monoclonal antibodies overlapping the DO-1 epitope were developed that are specific for phospho-Thr(18) (adjacent to the DO-1 epitope) and phospho-Ser(20) (within the DO-1 epitope) to determine whether direct evidence could be obtained for novel phosphorylation sites in human p53. A monoclonal antibody highly specific for phospho-Ser(20) detected significant phosphorylation of human p53 expressed in insect cells, whereas the relative proportion of p53 modified at Thr(18) was substantially lower. The relevance of these two novel phosphorylation sites to p53 regulation in human cells was made evident by the extensive phosphorylation of human p53 at Thr(18) and Ser(20) in a panel of human breast cancers with a wild-type p53 status. Phospho-Ser(20) or phospho-Thr(18) containing p53 peptides are as effective as the phospho-Ser(15) peptide at reducing mdm2 (mouse double minute 2) protein binding, indicating that the functional effects of these phosphorylation events might be to regulate the binding of heterologous proteins to p53. These results provide evidence in vivo for two novel phosphorylation sites within p53 at Ser(20) and Thr(18) that can affect p53 protein-protein interactions and indicate that some human cancers might have amplified one or more Ser(20) and Thr(18) kinase signalling cascades to modulate p53 activity.
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PMID:Novel phosphorylation sites of human tumour suppressor protein p53 at Ser20 and Thr18 that disrupt the binding of mdm2 (mouse double minute 2) protein are modified in human cancers. 1043 10


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