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

Li Qun Jia, Motonobu Osada, Chikashi Ishioka, Makio Gamo, Shuntaro Ikawa, Takao Suzuki, Hideki Shimodaira, Tomohito Niitani, Toshio Kudo, Mitoshi Akiyama, Narimiti Kimura, Mitsuyoshi Matsuo, Hiroshi Mizusawa, Noriho Tanaka, Hideki Koyama, Masayoshi Namba, Ryunosuke Kanamaru, Toshio Kuroki

Research output: Contribution to journalArticlepeer-review

157 Citations (Scopus)

Abstract

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 GALl 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.

Original languageEnglish
Pages (from-to)243-253
Number of pages11
JournalMolecular Carcinogenesis
Volume19
Issue number4
DOIs
Publication statusPublished - 1997 Sep 26

Keywords

  • Carcinogenesis
  • Immortalization
  • Temperature- sensitive mutation
  • Tumor suppressor gene
  • p53

ASJC Scopus subject areas

  • Molecular Biology
  • Cancer Research

Fingerprint Dive into the research topics of 'Screening the p53 status of human cell lines using a yeast functional assay'. Together they form a unique fingerprint.

Cite this