Precise epitope mapping of the murine transformation-associated protein, p53.

Wade-Evans, A.M.; Jenkins, Jennifer

doi:10.1002/j.1460-2075.1985.tb03686.x

Cited by 131 publications

(83 citation statements)

References 28 publications

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“…Conformation-specific monoclonal antibodies can be used to analyze the structure of p53, and a number of antibodies recognizing a specific region of the protein associated with functional properties of p53 have been raised. By binding to the C terminus (amino acids 372-381), the antibody PAb421 can allosterically activate latent p53 for DNA binding (Wade-Evans and Jenkins, 1985;Hupp et al, 1992Hupp et al, , 1994. DO-1 binds to the highly antigenic N terminus (amino acids 20 -25) of p53 (Stephen et al, 1995).…”

Section: Resultsmentioning

confidence: 99%

Allosteric Regulation of the Thermostability and DNA Binding Activity of Human p53 by Specific Interacting Proteins

Hansen¹,

Hupp

Lane

1996

Journal of Biological Chemistry

101

118

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Conformational stability is a prerequisite for the physiological activity of the tumor suppressor protein p53. p53 protein can be allosterically activated for DNA binding by phosphorylation or through noncovalent interaction with proteins such as DnaK, the Escherichia coli homologue of the heat shock protein Hsp70. We present in vitro evidence for a rapid temperature-dependent change in the conformation and tetrameric nature of wild-type p53 upon incubation at 37°C, which correlates with a permanent loss in DNA binding activity. We show that p53 is allosterically regulated for stabilization of the wild-type conformation and DNA binding activity at 37°C by binding of two classes of ligands to regulatory sites on the N and C terminus of the molecule through which an intrinsic instability of p53 is neutralized. Deletion of the domain conferring instability at the C terminus is sufficient to confer enhanced stability to the total protein. DnaK binding to the C terminus can profoundly protect p53 at 37°C from a temperature-dependent loss of the DNA binding activity but does not renature or activate denatured p53. In contrast, another activator of the DNA binding activity of latent p53, the monoclonal antibody PAb421, which also interacts with the C terminus of the protein, is not able to protect p53 from thermal denaturation. Two monoclonal antibodies to the N terminus of p53, PAb1801 and DO-1, do not activate the latent DNA binding function of p53 but can protect the p53 wild-type conformation at 37°C. Thus, activation of the DNA binding function of p53 is not synonymous with protection from thermal denaturation, and therefore, both of these pathways may be used in cells to control the physiological activity of p53. The protection of p53 conformation from heat denaturation by interacting proteins suggests a novel mechanism by which p53 function could be regulated in vivo.

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Section: Resultsmentioning

confidence: 99%

Allosteric Regulation of the Thermostability and DNA Binding Activity of Human p53 by Specific Interacting Proteins

Hansen¹,

Hupp

Lane

1996

Journal of Biological Chemistry

101

118

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“…Analysis of monoclonal antibodies produced against human p53 demonstrated a strong bias in the epitope recognised by these monoclonal antibodies. Most of them recognised epitopes localised in the amino terminus of p53 (Wade-Evans & Jenkins, 1985;Vojtesek et al, 1992;Bartek et al, 1993;Legros et al, 1993). Furthermore, we showed that p53 antibodies in sera of animals hyperimmunised with human p53 recognised epitopes similar to those identified in a previous work (Schlichtholz et al, 1992; Y. Legros & T. Soussi, submitted for publication).…”

Section: Discussionmentioning

confidence: 99%

Analyses of p53 antibodies in sera of patients with lung carcinoma define immunodominant regions in the p53 protein

Schlichtholz

Trédaniel²,

Lubin³

et al. 1994

Br J Cancer

134

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Summary Antibodies specific for human p53 were analysed in sera of lung cancer patients. We detected p53 antibodies in the sera of 24% (10/42) of patients with lung carcinoma. The distribution was as follows: 4/9 small-cell lung carcinomas (SCLCs), 2/18 squamous cell lung carcinomas (SCCs), 2/10 adenocarcinomas (ADCs) and 2/5 large-cell lung carcinomas (LCCs). p53 antibodies were always present at the time of diagnosis and did not appear during progression of the disease. Using an original peptide-mapping procedure, we precisely localised the p53 epitopes recognised by p53 antibodies. Immunodominant epitopes reacting with antibodies were localised in the amino and carboxy termini of the protein, similar to those found in breast carcinoma patients or in animals immunised with p53. In light of these data, we suggest that p53 antibodies occur via a self-immunisation process that is the consequence of p53 accumulation in tumour cells. p53 antibodies were also detected in two patients without detected malignant disease. One of these patients died 6 months later of lung carcinoma, suggesting that p53 antibodies may be a precocious marker of p53 alteration.

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“…Aliquots containing equal amounts of TCA-insoluble radioactivity were first precleared, incubated with anti-P53 monoclonal antibodies or a control MAb, MOPC2 1, and then immunoprecipitated using protein ASepharose, which was followed by SDS-polyacrylamide gel electrophoresis on a 10% gel essentially as described (15). Anti-p53 monoclonal antibodies used were PAb 1801 (Oncogene Science, Manhasset, NY) (16) (16,18).…”

Section: Methodsmentioning

confidence: 99%

Identification of intronic point mutations as an alternative mechanism for p53 inactivation in lung cancer.

Takahashi¹,

D’Amico²,

Chiba³

et al. 1990

J. Clin. Invest.

102

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The p53 gene initially was thought to be an oncogene, but recent evidence suggests that wild-type p53 can function as a tumor suppressor gene in lung, colon, and breast cancer as well as less common malignancies. This study reports the first identification of intronic point mutations as a mechanism for inactivation of the p53 tumor suppressor gene. Abnormally sized p53 mRNAs found in a small cell and a non-small cell lung cancer cell line were characterized by sequence analysis of cDNA/PCR products, the RNase protection assay and immunoprecipitation. These mRNAs were found to represent aberrant splicing leading to the production of abnormal or no p53 protein. Sequence analysis of genomic DNA revealed that a point mutation at the splice acceptor site in the third intron or the splice donor site in the seventh intron accounts for the abnormal mRNA splicing. In one patient the same intronic point mutation was found in the tumor cell line derived from a bone marrow metastasis and in multiple liver metastases but not in normal DNA, indicating that it occurred as a somatic event before the development of these metastases. These findings further support the role of inactivation of the p53 gene in the pathogenesis of lung cancer and indicate the role of intronic point mutation in this process. (J. Clin Invest. 1990. 86:363-369).

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Precise epitope mapping of the murine transformation-associated protein, p53.

Cited by 131 publications

References 28 publications

Allosteric Regulation of the Thermostability and DNA Binding Activity of Human p53 by Specific Interacting Proteins

Allosteric Regulation of the Thermostability and DNA Binding Activity of Human p53 by Specific Interacting Proteins

Analyses of p53 antibodies in sera of patients with lung carcinoma define immunodominant regions in the p53 protein

Identification of intronic point mutations as an alternative mechanism for p53 inactivation in lung cancer.

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