Screening the p53 status of human cell lines using a yeast functional assay

We have screened the p53 status of 156 human cell lines, including 142 tumor cell lines from 27 different tumor types and 14 cell lines from normal tissues by using functional analysis of separated alleles in yeast. This assay enables us to score wild‐type p53 expression on the basis of the ability of expressed p53 to transactivate the reporter gene HIS3 via the p53‐responsive GAL1 promoter in Saccharomyces cerevisiae. Of 142 tumor cell lines, at least 104 lines (73.2%) were found to express the mutated p53 gene: 94 lines (66.2%) were mutated in both alleles, three lines (2.1%) were heterozygous, and no p53 cDNA was amplified from seven lines (4.9%). Of the 14 cell lines originating from normal tissues, all the transformed or immortalized cell lines expressed mutant p53 only. Yeast cells expressing mutant p53 derived from 94 cell lines were analyzed for temperature‐sensitive growth. p53 cDNA from eight cell lines showed p53‐dependent temperature‐sensitive growth, growing at 30°C but not at 37°C. Four temperature‐sensitive p53 mutations were isolated: CAT→CGT at codon 214 (H214R), TAC→TGC at codon 234 (Y234C), GTG→ATG at codon 272 (V272M), and GAG→AAG (E285K). Functionally wild‐type p53 was detected in 38 tumor cell lines (26.8%) and all of the diploid fibroblasts at early and late population doubling levels. These results strongly support the previous findings that p53 inactivation is one of the most frequent genetic events that occurs during carcinogenesis and immortalization. Mol. Carcinog. 19:243–253, 1997. © 1997 Wiley‐Liss, Inc.

[1]  A. Fattaey,et al.  An Adenovirus Mutant That Replicates Selectively in p53- Deficient Human Tumor Cells , 1996, Science.

[2]  W. Hong,et al.  Retrovirus–mediated wild–type P53 gene transfer to tumors of patients with lung cancer. , 1996, Nature Medicine.

[3]  C. Ishioka,et al.  Mutation in p53 and de‐regulation of p53‐related gene expression in three human cell lines immortalized with 4‐nitroquinoline 1‐oxide or 60Co gamma rays , 1996, International journal of cancer.

[4]  J. Foncin.  The true Western diet , 1996, Nature Medicine.

[5]  L. Holmberg,et al.  The p53 gene in breast cancer: prognostic value of complementary DNA sequencing versus immunohistochemistry. , 1996, Journal of the National Cancer Institute.

[6]  W. Yung,et al.  Adenovirus-mediated transfer of the p53 gene produces rapid and generalized death of human glioma cells via apoptosis. , 1996, Cancer research.

[7]  R. Ueda,et al.  Prognostic significance of abnormal p53 accumulation in primary, resected non-small-cell lung cancers. , 1996, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[8]  M. Oren,et al.  Specific loss of apoptotic but not cell‐cycle arrest function in a human tumor derived p53 mutant. , 1996, The EMBO journal.

[9]  David E. Housman,et al.  Hypoxia-mediated selection of cells with diminished apoptotic potential in solid tumours , 1996, Nature.

[10]  E. Shaulian,et al.  Induction of apoptosis in HeLa cells by trans-activation-deficient p53. , 1995, Genes & development.

[11]  H. Nguyen,et al.  Cytotoxic effects of adenovirus-mediated wild-type p53 protein expression in normal and tumor mammary epithelial cells. , 1995, Clinical cancer research : an official journal of the American Association for Cancer Research.

[12]  U. Moll,et al.  Wild-type p53 protein undergoes cytoplasmic sequestration in undifferentiated neuroblastomas but not in differentiated tumors. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[13]  P. Chappuis,et al.  A simple p53 functional assay for screening cell lines, blood, and tumors. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[14]  S. Friend,et al.  Mutational analysis of the carboxy-terminal portion of p53 using both yeast and mammalian cell assays in vivo. , 1995, Oncogene.

[15]  D. Thorley-Lawson,et al.  A novel form of Epstein-Barr virus latency in normal B cells in vivo , 1995, Cell.

[16]  John Calvin Reed,et al.  Tumor suppressor p53 is a direct transcriptional activator of the human bax gene , 1995, Cell.

[17]  J. Roth,et al.  In vivo molecular therapy with p53 adenovirus for microscopic residual head and neck squamous carcinoma. , 1995, Cancer research.

[18]  K. Vousden,et al.  Transcriptional activation by p53 correlates with suppression of growth but not transformation , 1994, Cell.

[19]  S C Chen,et al.  Accumulation of nuclear p53 and tumor progression in bladder cancer. , 1994, The New England journal of medicine.

[20]  B. Quesnel,et al.  p53 Mutations Are Associated With Resistance to Chemotherapy and Short Survival in Hematologic Malignancies , 1994 .

[21]  H. Shepard,et al.  Development and characterization of recombinant adenoviruses encoding human p53 for gene therapy of cancer. , 1994, Human gene therapy.

[22]  R. Iggo,et al.  A rapid PCR fidelity assay. , 1994, Nucleic acids research.

[23]  T. Soussi,et al.  Analysis of the most representative tumour‐derived p53 mutants reveals that changes in protein conformation are not correlated with loss of transactivation or inhibition of cell proliferation. , 1994, The EMBO journal.

[24]  S. Friend.  p53: a glimpse at the puppet behind the shadow play. , 1994, Science.

[25]  Stephen J. Elledge,et al.  p53-dependent inhibition of cyclin-dependent kinase activities in human fibroblasts during radiation-induced G1 arrest , 1994, Cell.

[26]  J. Trent,et al.  WAF1, a potential mediator of p53 tumor suppression , 1993, Cell.

[27]  K. Kohn,et al.  Role of the p53 tumor suppressor gene in cell cycle arrest and radiosensitivity of Burkitt's lymphoma cell lines. , 1993, Cancer research.

[28]  S. Friend,et al.  Screening patients for heterozygous p53 mutations using a functional assay in yeast , 1993, Nature genetics.

[29]  D. Housman,et al.  p53-dependent apoptosis modulates the cytotoxicity of anticancer agents , 1993, Cell.

[30]  R. Metcalf,et al.  p53 mutations in human immortalized epithelial cell lines. , 1993, Carcinogenesis.

[31]  R. Metcalf,et al.  p53 gene mutation and integrated hepatitis B viral DNA sequences in human liver cancer cell lines. , 1993, Carcinogenesis.

[32]  S. Y. Zhang,et al.  p53 alterations in human squamous cell carcinomas and carcinoma cell lines. , 1993, The American journal of pathology.

[33]  B. Vogelstein,et al.  A mammalian cell cycle checkpoint pathway utilizing p53 and GADD45 is defective in ataxia-telangiectasia , 1992, Cell.

[34]  T. Sano,et al.  Role of p53 mutations in endocrine tumorigenesis: mutation detection by polymerase chain reaction-single strand conformation polymorphism. , 1992, Cancer research.

[35]  M. Kastan,et al.  Wild-type p53 is a cell cycle checkpoint determinant following irradiation. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[36]  A. Levine,et al.  Two distinct mechanisms alter p53 in breast cancer: mutation and nuclear exclusion. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[37]  H. Westphal,et al.  Abnormal structure and expression of the p53 gene in human ovarian carcinoma cell lines. , 1992, Cancer research.

[38]  Koichi Sugimoto,et al.  Frequent mutations in the p53 gene in human myeloid leukemia cell lines. , 1992, Blood.

[39]  M. Jourdan,et al.  Mutations of the p53 gene in human myeloma cell lines. , 1992, Oncogene.

[40]  N. Tsuchida,et al.  Most human squamous cell carcinomas in the oral cavity contain mutated p53 tumor-suppressor genes. , 1992, Oncogene.

[41]  R. Iggo,et al.  Mammalian p53 can function as a transcription factor in yeast. , 1992, Nucleic acids research.

[42]  T Matozaki,et al.  Missense mutations and a deletion of the p53 gene in human gastric cancer. , 1992, Biochemical and Biophysical Research Communications - BBRC.

[43]  A. Klein-Szanto,et al.  Human pancreatic carcinomas and cell lines reveal frequent and multiple alterations in the p53 and Rb-1 tumor-suppressor genes. , 1992, Oncogene.

[44]  H. Westphal,et al.  Analysis of the p53 gene in human uterine carcinoma cell lines. , 1991, Cancer research.

[45]  S. Hirohashi,et al.  p53 gene mutations in gastric cancer metastases and in gastric cancer cell lines derived from metastases. , 1991, Cancer research.

[46]  R. Metcalf,et al.  Genetic analysis of human esophageal tumors from two high incidence geographic areas: frequent p53 base substitutions and absence of ras mutations. , 1991, Cancer research.

[47]  A. Berrebi,et al.  p53 in chronic myelogenous leukemia in acute phase. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[48]  B. Vogelstein,et al.  p53 mutations in human cancers. , 1991, Science.

[49]  Detection of aberrations of the p53 alleles and the gene transcript in human tumor cell lines by single-strand conformation polymorphism analysis. , 1991, Cancer research.

[50]  Arnold J. Levine,et al.  The E6 oncoprotein encoded by human papillomavirus types 16 and 18 promotes the degradation of p53 , 1990, Cell.

[51]  J. Milner,et al.  Temperature-dependent switching between "wild-type" and "mutant" forms of p53-Val135. , 1990, Journal of molecular biology.

[52]  W. Bodmer,et al.  p53 mutations in colorectal cancer. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[53]  M. Haas,et al.  Frequent mutations in the p53 tumor suppressor gene in human leukemia T-cell lines , 1990, Molecular and cellular biology.

[54]  S. Fields,et al.  Presence of a potent transcription activating sequence in the p53 protein. , 1990, Science.

[55]  G. Lozano,et al.  Transcriptional activation by wild-type but not transforming mutants of the p53 anti-oncogene. , 1990, Science.

[56]  K. Isselbacher,et al.  Abnormal structure and expression of p53 gene in human hepatocellular carcinoma. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[57]  F. Collins,et al.  Mutations in the p53 gene occur in diverse human tumour types , 1989, Nature.

[58]  D. Helfman,et al.  Molecular cloning and in vitro expression of a cDNA clone for human cellular tumor antigen p53 , 1985, Molecular and cellular biology.

[59]  V. Rotter,et al.  Major deletions in the gene encoding the p53 tumor antigen cause lack of p53 expression in HL-60 cells. , 1985, Proceedings of the National Academy of Sciences of the United States of America.