Expression and stability of p53 protein in normal human mammary epithelial cells.
To examine the p53 protein expressed in human mammary epithelial cell strains grown in vitro we first established the presence of only wild-type p53 and lack of any missense mutations in a representative normal mammary epithelial cell strain, 76N, by cloning and sequencing the entire coding region of the p53 mRNA. The p53 protein expressed in this cell strain and a number of similarly derived normal mammary epithelial cell strains was compared with mammary epithelial tumor cell lines having known p53 mutations and with fibroblasts derived from normal mammary tissue. In all cell strains, immunoprecipitation from metabolically labelled cells using monoclonal antibodies (mAb) PAb 1801 and PAb 122 revealed easily detectable p53 protein. Surprisingly, mAb PAb 1620 (wild type-specific) and PAb 240 (mutant-specific) each immunoprecipitated p53 protein from both normal and tumor cells. Furthermore, p53 protein in normal mammary epithelial cells was shown to be markedly more stable (half-life of approximately 3 h) compared to that in mammary fibroblasts or rodent fibroblasts (half-life < 30 min). Immunocytochemistry with PAb 1801 showed detectable p53 protein in normal mammary epithelial cells with a predominantly nuclear staining; however, p53 protein was undetectable in fibroblasts by immunocytochemistry. Together, these results reveal unusual features of wild-type p53 protein in normal human mammary epithelial cells, suggesting a cell type-specific regulation of its expression and function.
Delmolino
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Department of Radiation Oncology, New England Medical Center Hospitals, Boston, MA.
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D000911:Antibodies, Monoclonal; D001483:Base Sequence; D001940:Breast; D002460:Cell Line; D003001:Cloning, Molecular; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D016158:Genes, p53; D006801:Humans; D007124:Immunoenzyme Techniques; D007150:Immunohistochemistry; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D016159:Tumor Suppressor Protein p53 / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics
D000911:Antibodies, Monoclonal; D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53