Differential gene expression in p53-mediated apoptosis-resistant vs. apoptosis-sensitive tumor cell lines.
Induction of wild-type p53 in the ECV-304 bladder carcinoma cell line by infection with a p53 recombinant adenovirus (Ad5CMV-p53) resulted in extensive apoptosis and eventual death of nearly all of the cells. As a strategy to determine the molecular events important to p53-mediated apoptosis in these transformed cells, ECV-304 cells were selected for resistance to p53 by repeated infections with Ad5CMV-p53. We compared the expression of 5,730 genes in p53-resistant (DECV) and p53-sensitive ECV-304 cells by reverse transcription-PCR, Northern blotting, and DNA microarray analysis. The expression of 480 genes differed by 2-fold or more between the two p53-infected cell lines. A number of potential targets for p53 were identified that play roles in cell cycle regulation, DNA repair, redox control, cell adhesion, apoptosis, and differentiation. Proline oxidase, a mitochondrial enzyme involved in the proline/pyrroline-5-carboxylate redox cycle, was up-regulated by p53 in ECV but not in DECV cells. Pyrroline-5-carboxylate (P5C), a proline-derived metabolite generated by proline oxidase, inhibited the proliferation and survival of ECV-304 and DECV cells and induced apoptosis in both cell lines. A recombinant proline oxidase protein tagged with a green fluorescent protein at the amino terminus localized to mitochondria and induced apoptosis in p53-null H1299 non-small cell lung carcinoma cells. The results directly implicate proline oxidase and the proline/P5C pathway in p53-induced growth suppression and apoptosis.
Maxwell
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Department of Pathology and Laboratory Medicine, Texas A&M University Health Science Center, College Station, TX 77843-1114, USA. s-maxwell@tamu.edu
Davis
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GE
D000256:Adenoviridae; D017209:Apoptosis / Q000502:physiology*; D001119:Arginase / Q000235:genetics; D018432:Drug Resistance, Multiple; D005696:Galactosidases / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D011394:Proline Oxidase / Q000235:genetics; D011994:Recombinant Proteins / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D001749:Urinary Bladder Neoplasms
D008164:Luminescent Proteins; D011994:Recombinant Proteins; D016159:Tumor Suppressor Protein p53; D049452:Green Fluorescent Proteins; D011394:Proline Oxidase; D005696:Galactosidases; D001119:Arginase