title: E2F1-mediated transcriptional inhibition of the plasminogen activator inhibitor type 1 gene.
abstract: Gene expression of the plasminogen activation system is cell-cycle dependent. Previously, we showed that ectopic expression of E2F1 repressed the plasminogen activator inhibitor type 1 (PAI-1) promoter in a manner dependent on the presence of DNA-binding and transactivation domains of E2F1 but independent of binding to pocket-binding proteins, suggesting a novel mechanism for E2F-mediated negative gene regulation [Koziczak, M., Krek, W. & Nagamine, Y. (2000) Mol. Cell. Biol. 20, 2014-2022]. However, it remains to be seen whether endogenous E2F can exert a similar effect. We report here that down-regulation of PAI-1 gene expression correlates with an increase in endogenous E2F activity. When cells were treated with a cdk2/4-specific inhibitor, which maintains E2F in an inactive state, the decline of serum-induced PAI-1 mRNA levels was suppressed. In mutant U2OS cells expressing a temperature-sensitive retinoblastoma protein (pRB), a shift to a permissive temperature induced PAI-1 mRNA expression. In U2OS cells stably expressing an E2F1-estrogen receptor chimeric protein that could be activated by tamoxifen, PAI-1 gene transcription was markedly reduced by tamoxifen even in the presence of cycloheximide. These results all indicate that endogenous E2F can directly repress the PAI-1 gene. DNase I hypersensitive-site analysis of the PAI-1 promoter suggested the involvement of conformation changes in chromatin structure of the PAI-1 promoter. 5' deletion analysis of the PAI-1 promoter showed that multiple sites were responsible for the E2F negative regulation, some of which were promoter dependent. Interestingly, one of these sites is a p53-binding element.
authors.affiliation: Friedrich Miescher Institute, Basel, Switzerland.
authors.forename: M
authors.initials: M
authors.lastname: Koziczak
authors.forename: H
authors.initials: H
authors.lastname: Müller
authors.forename: K
authors.initials: K
authors.lastname: Helin
authors.forename: Y
authors.initials: Y
authors.lastname: Nagamine
mesh_terms: D000818:Animals; D001665:Binding Sites; D042846:CDC2-CDC28 Kinases; D002453:Cell Cycle; D018797:Cell Cycle Proteins; D002460:Cell Line; D002843:Chromatin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D004247:DNA / Q000235:genetics / Q000378:metabolism; D018983:DNA Footprinting; D004268:DNA-Binding Proteins; D003850:Deoxyribonuclease I / Q000378:metabolism; D015536:Down-Regulation; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D006801:Humans; D009154:Mutation; D017395:Plasminogen Activator Inhibitor 1 / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism; D013552:Swine; D013696:Temperature; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D015854:Up-Regulation
chemical_list: D018797:Cell Cycle Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D017395:Plasminogen Activator Inhibitor 1; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D012097:Repressor Proteins; D016160:Retinoblastoma Protein; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D004247:DNA; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; C495900:CDK4 protein, human; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases; D003850:Deoxyribonuclease I