title: Ionizing radiation down-regulates p53 protein in primary Egr-1-/- mouse embryonic fibroblast cells causing enhanced resistance to apoptosis.
abstract: In this study, we sought to investigate the mechanism of the proapoptotic function of Egr-1 in relation to p53 status in normal isogenic cell backgrounds by using primary MEF cells established from homozygous (Egr-1(-/-)) and heterozygous (Egr-1(+/-)) Egr-1 knock-out mice. Ionizing radiation caused significantly enhanced apoptosis in Egr-1(+/-) cells (22.8%; p < 0.0001) when compared with Egr-1(-/-) cells (3.5%). Radiation elevated p53 protein in Egr-1(+/-) cells in 3-6 h. However, in Egr-1(-/-) cells, the p53 protein was down-regulated 1 h after radiation and was completely degraded at the later time points. Radiation elevated the p53-CAT activity in Egr-1(+/-) cells but not in Egr-1(-/-) cells. Interestingly, transient overexpression of EGR-1 in p53(-/-) MEF cells caused marginal induction of radiation-induced apoptosis when compared with p53(+/+) MEF cells. Together, these results indicate that Egr-1 may transregulate p53, and both EGR-1 and p53 functions are essential to mediate radiation-induced apoptosis. Rb, an Egr-1 target gene, forms a trimeric complex with p53 and MDM2 to prevent MDM2-mediated p53 degradation. Low levels of Rb including hypophosphorylated forms were observed in Egr-1(-/-) MEF cells before and after radiation when compared with the levels observed in Egr-1(+/-) cells. Elevated amounts of the p53-MDM2 complex and low amounts of Rb-MDM-2 complex were observed in Egr-1(-/-) cells after radiation. Because of a reduction in Rb binding to MDM2 and an increase in MDM2 binding with p53, p53 is directly degraded by MDM2, and this leads to inactivation of the p53-mediated apoptotic pathway in Egr-1(-/-) MEF cells. Thus, the proapoptotic function of Egr-1 may involve the mediation of Rb protein that is essential to overcome the antiapoptotic function of MDM2 on p53.
authors.affiliation: Department of Radiation Medicine and Markey Cancer Center, University of Kentucky, Lexington, Kentucky 40536, USA.
authors.forename: A
authors.initials: A
authors.lastname: Das
authors.forename: D
authors.initials: D
authors.lastname: Chendil
authors.forename: S
authors.initials: S
authors.lastname: Dey
authors.forename: M
authors.initials: M
authors.lastname: Mohiuddin
authors.forename: M
authors.initials: M
authors.lastname: Mohiuddin
authors.forename: J
authors.initials: J
authors.lastname: Milbrandt
authors.forename: V M
authors.initials: VM
authors.lastname: Rangnekar
authors.forename: M M
authors.initials: MM
authors.lastname: Ahmed
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics; D002478:Cells, Cultured; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000096:biosynthesis / Q000235:genetics; D003587:Cytomegalovirus / Q000235:genetics; D004268:DNA-Binding Proteins / Q000172:deficiency / Q000378:metabolism*; D015536:Down-Regulation / Q000528:radiation effects; D051766:Early Growth Response Protein 1; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D005822:Genetic Vectors; D017874:Immediate-Early Proteins; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D018345:Mice, Knockout; D009687:Nuclear Proteins; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000235:genetics; D051736:Proto-Oncogene Proteins c-mdm2; D012333:RNA, Messenger / Q000096:biosynthesis; D011839:Radiation, Ionizing; D016160:Retinoblastoma Protein / Q000378:metabolism; D014157:Transcription Factors / Q000172:deficiency / Q000378:metabolism*; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; C406792:BFAR protein, human; D002352:Carrier Proteins; C493957:Cdkn1a protein, mouse; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004268:DNA-Binding Proteins; D051766:Early Growth Response Protein 1; C486109:Egr1 protein, mouse; D017874:Immediate-Early Proteins; D008565:Membrane Proteins; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D016160:Retinoblastoma Protein; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; C497207:Mdm2 protein, mouse; D051736:Proto-Oncogene Proteins c-mdm2