title: Increased mdm-2 expression in a p53-independent manner blocks UV-induced cell cycle arrest and apoptosis in human osteosarcoma cells.
abstract: DNA damage results in an increase in P53 levels, which is required to initiate a P53-mediated cell cycle arrest and/or apoptosis. P53 and MDM-2 form a feedback control loop: while P53 can transactivate the MDM-2 gene, high levels of MDM-2 inhibit P53 transactivation as well as promote rapid degradation of P53. In the present study, we investigated the interaction between endogenous MDM-2 and P53 following UV-induced DNA damage in an MDM-2 overexpression cell line. A human osteosarcoma cell line (OsACL, which contains wild-type P53 and overexpresses MDM-2 protein) was used in this study. Here we show that following UV treatment, P53 levels increased in the OsACL cells despite the presence of high-level endogenous MDM-2; however, CAT assays using a P53 reporter system revealed that this P53 was transcriptionally inactive. Although P53 transactivation was inhibited, MDM-2 levels rose markedly following UV irradiation. Northern blot analysis revealed that the increase in MDM-2 protein levels was a result of increased levels of MDM-2 mRNA, possibly due to increased transcription. Cell cycle analysis revealed that OsACL cells were markedly resistant to UV-induced apoptosis. Transfection of OsACL cells with an anti-sense MDM-2 plasmid dowregulated MDM-2 expression and increased UV-induced apoptosis. In conclusion, MDM-2 overexpression can block UV-induced cell cycle arrest and apoptosis by inhibiting P53 transcriptional activity. Furthermore, increased expression of MDM-2 in OsACL cells following UV irradiation appears to be related to P53-independent mechanisms.
authors: Department of Surgery, University of Louisville, James Graham Brown Cancer Center, Louisville, KY 40202, USA.
authors: Yan Bin
authors: YB
authors: Dong
authors: Hai Liang
authors: HL
authors: Yang
authors: Mary Jane
authors: MJ
authors: Elliott
authors: Kelly M
authors: KM
authors: McMasters
mesh_terms: D000818:Animals; D017209:Apoptosis; D015152:Blotting, Northern; D015153:Blotting, Western; D002453:Cell Cycle; D045744:Cell Line, Tumor; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D004249:DNA Damage; D004307:Dose-Response Relationship, Radiation; D015536:Down-Regulation; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells; D009687:Nuclear Proteins; D016376:Oligonucleotides, Antisense / Q000737:chemistry / Q000494:pharmacology; D012516:Osteosarcoma / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis*; D051736:Proto-Oncogene Proteins c-mdm2; D012333:RNA, Messenger / Q000378:metabolism; D013997:Time Factors; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis*; D014466:Ultraviolet Rays; D015854:Up-Regulation
chemical_list: D009687:Nuclear Proteins; D016376:Oligonucleotides, Antisense; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; D015500:Chloramphenicol O-Acetyltransferase; C497206:MDM2 protein, human; C497207:Mdm2 protein, mouse; D051736:Proto-Oncogene Proteins c-mdm2