Transcriptional activation of the carboxylesterase 2 gene by the p53 pathway.
The p53 tumor suppressor is an important regulator of cellular response to chemotherapeutic agents by virtue of the protein's ability, upon activation by phosphorylation, to transcriptionally activate a number of genes involved in cell proliferation, apoptosis, and metabolism. Transcriptome analysis following introduction of a constitutively active form of p53 (p53T18D/S20D) into colon carcinoma cell lines identified transcriptional activation of the carboxylesterase 2 (CES-2) gene, which is involved in drug metabolism. We examined whether p53 activated by the DNA-damaging drug 5-fluorouracil (5-FU) also induces CES-2 expression. Our experiments showed that 5-FU induced CES-2 expression in two colon carcinoma cell lines that express wild-type p53 (HCT116 p53(+/+) and RKO) but not in five lines that are p53-null (HCT116 p53(-/- )) or express mutated p53 (HT29, KM12C, KM12SM, and KM12L4A). Sequence analysis revealed a putative p53-binding element in the first intron of CES-2 that differed from consensus by one nucleotide. A reporter gene assay showed that the luciferase construct with the p53-binding element responded to 5-FU treatment, whereas the reporter construct without the binding element did not. Chromatin immunoprecipitation assay confirmed that p53 bound the CES-2 fragment containing the p53-binding element after 5-FU treatment, whereas p21 binding to p53 was present with or without chemotherapy. Knockdown of expression of CES-2 and p53 by small interference RNA in RKO and HCT116 p53(+/+) cells attenuated the anti-proliferation effects of CPT11. These results taken together show that activated p53 directly regulates CES-2 expression via a p53-binding site, representing a novel mechanism through which the p53 pathway modulates drug metabolism. In addition, the degree of homology in the p53-binding element may determine the strength of p53 regulation.
Choi
Woonyoung
W
Department of Pathology, The University of Texas MD Anderson Cancer Center, Houston, Texas 77030, USA.
Cogdell
David
D
Feng
Yumei
Y
Hamilton
Stanley R
SR
Zhang
Wei
W
D000230:Adenocarcinoma / Q000235:genetics*; D043182:Carboxylesterase / Q000235:genetics*; D045744:Cell Line, Tumor; D002843:Chromatin / Q000235:genetics; D003110:Colonic Neoplasms / Q000235:genetics*; D016384:Consensus Sequence; D004249:DNA Damage; D017931:DNA Primers; D004273:DNA, Neoplasm / Q000187:drug effects / Q000235:genetics; D005472:Fluorouracil / Q000494:pharmacology; D006801:Humans; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015533:Transcriptional Activation; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000235:genetics* / Q000378:metabolism
D002843:Chromatin; D017931:DNA Primers; D004273:DNA, Neoplasm; D016159:Tumor Suppressor Protein p53; C517204:CES2 protein, human; D043182:Carboxylesterase; D005472:Fluorouracil