title: Doxorubicin and 5-fluorouracil induced accumulation and transcriptional activity of p53 are independent of the phosphorylation at serine 15 in MCF-7 breast cancer cells.
abstract: The chemotherapeutic agents doxorubicin (dox) or 5-fluorouracil (5FU) are used to treat cancer cells as they cause irreparable DNA damage, inducing these aberrant cells to undergo cell death. The mediator of this process is presumed to be in part the tumor suppressor p53 which regulates genes involved in DNA repair and cell death. When MCF-7 breast cancer cells are treated with these drugs, we observed that the level of p53 and the p53 negative regulator, Mdm2, increased, as seen by others. But contrary to some reports, we observed minimal phosphorylation of p53 at serine 15 in MCF-7 cells after drug treatment. Interestingly, we determined that there was differential regulation of the kinases ATM and Chk2 with the drug treatments, likely the cause for the lack of phosphorylation of p53. We found a dramatic drop in p53 DNA binding affinity for p21 and other gene response elements (RE) after drug treatment. To determine if the p53 that accumulated in the drug treated cells was functionally active, we monitored changes in the protein products of two p53-regulated genes following drug treatment with and without the addition of a p53-specific siRNA. In response to 5FU, both p21 and Mdm2 proteins increased and that increase was alleviated if a p53-specific siRNA was added. This effect was not seen with the addition of dox. Thus, the phosphorylation at serine 15 is not necessary for the functional activation of this transcription factor. We propose a new model for the regulation of p53, Mdm2, and MdmX after drug treatment.
authors.affiliation: Department of Biological Sciences; Binghamton University; Binghamton, NY USA.
authors.forename: Matthew T
authors.initials: MT
authors.lastname: Balmer
authors.affiliation: Department of Biological Sciences; Binghamton University; Binghamton, NY USA.
authors.forename: Ryan D
authors.initials: RD
authors.lastname: Katz
authors.affiliation: Department of Biological Sciences; Binghamton University; Binghamton, NY USA.
authors.forename: Si
authors.initials: S
authors.lastname: Liao
authors.affiliation: Department of Biological Sciences; Binghamton University; Binghamton, NY USA.
authors.forename: James S
authors.initials: JS
authors.lastname: Goodwine
authors.affiliation: Department of Biological Sciences; Binghamton University; Binghamton, NY USA.
authors.forename: Susannah
authors.initials: S
authors.lastname: Gal
mesh_terms: D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D064007:Ataxia Telangiectasia Mutated Proteins / Q000378:metabolism; D018797:Cell Cycle Proteins; D064447:Checkpoint Kinase 2 / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D004317:Doxorubicin / Q000494:pharmacology*; D005260:Female; D005472:Fluorouracil / Q000494:pharmacology*; D006801:Humans; D061986:MCF-7 Cells; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism; D012694:Serine / Q000378:metabolism*; D014158:Transcription, Genetic; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*
chemical_list: D000903:Antibiotics, Antineoplastic; D000964:Antimetabolites, Antineoplastic; C493956:CDKN1A protein, human; D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; C482775:MDM4 protein, human; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D016159:Tumor Suppressor Protein p53; D012694:Serine; D004317:Doxorubicin; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; D064447:Checkpoint Kinase 2; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C578634:CHEK2 protein, human; D005472:Fluorouracil
keywords: 5-fluorouracil; ATM; Chk2; DNA binding; MCF-7; Mdm2; MdmX; doxorubicin; p53; posttranslational modification