Genetic and immunochemical analysis of mutant p53 in human breast cancer cell lines.
The expression of the tumour suppressor gene p53 was analysed in 11 human breast cancer cell lines by immunohistochemistry, immunoprecipitation and cDNA sequencing. We used a panel of anti-p53 monoclonal antibodies for cell staining and found abnormalities in every case. Eight of the cell lines produce a form of p53 which can be immunoprecipitated by the monoclonal antibody PAb240 but not by PAb1620. In the murine system PAb240 only immunoprecipitates mutant p53. We sequenced p53 cDNA directly from four of the PAb240 positive cell lines using asymmetric PCR templates. All four contained missense mutations in p53 RNA, with no detectable expression of the wild type sequence. Different residues were affected in each cell line, but all the mutations changed amino acids conserved from man to Xenopus. These results imply that as in the murine system, the PAb240 antibody reliably detects a wide variety of p53 mutations and that these mutations have a common effect on the structure of p53. Immunohistochemical data suggest that p53 mutation is the commonest genetic alteration so far detected in primary breast cancer.
ICRF Molecular Immunochemistry Laboratory, Clare Hall Laboratories, Potters Bar, Herts, UK.
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Bartek
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Iggo
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Gannon
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Lane
D000595:Amino Acid Sequence; D000906:Antibodies / Q000276:immunology; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D002285:Carcinoma, Intraductal, Noninfiltrating / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D004273:DNA, Neoplasm / Q000235:genetics / Q000378:metabolism; D000939:Epitopes / Q000276:immunology; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D008969:Molecular Sequence Data; D009154:Mutation; D015513:Oncogene Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011233:Precipitin Tests; D012334:RNA, Neoplasm / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured / Q000378:metabolism / Q000473:pathology; D016159:Tumor Suppressor Protein p53
D000906:Antibodies; D004273:DNA, Neoplasm; D000939:Epitopes; D015513:Oncogene Proteins; D010750:Phosphoproteins; D012334:RNA, Neoplasm; D016159:Tumor Suppressor Protein p53