Bartek J, Iggo R, Gannon J, Lane D P
ICRF Molecular Immunochemistry Laboratory, Clare Hall Laboratories, Potters Bar, Herts, UK.
Oncogene. 1990 Jun;5(6):893-9.
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.
通过免疫组织化学、免疫沉淀和cDNA测序分析了11种人乳腺癌细胞系中肿瘤抑制基因p53的表达情况。我们使用一组抗p53单克隆抗体对细胞进行染色，结果发现每一例均存在异常。其中8种细胞系产生一种形式的p53，它能被单克隆抗体PAb240免疫沉淀，但不能被PAb1620免疫沉淀。在小鼠系统中，PAb240仅能免疫沉淀突变型p53。我们使用不对称PCR模板直接对4种PAb240阳性细胞系的p53 cDNA进行测序。所有4种细胞系的p53 RNA均含有错义突变，未检测到野生型序列的表达。每个细胞系中受影响的残基不同，但所有突变均改变了从人到非洲爪蟾保守的氨基酸。这些结果表明，与小鼠系统一样，PAb240抗体能可靠地检测到多种p53突变，且这些突变对p53的结构有共同影响。免疫组织化学数据表明，p53突变是迄今为止在原发性乳腺癌中检测到的最常见的基因改变。
