title: Alterations in transcription factor-binding activities to fibronectin promoter during aging of vascular endothelial cells.
abstract: Previously, we showed that the expression of the fibronectin (FN) gene is enhanced during aging of human endothelial cells and fibroblasts. To elucidate the mechanism, we explored binding proteins to the FN promoter. The promoter contains sites for the general transcription factors: CAAT-binding transcription factor (CTF), promoter-specific transcription factor-1 (Sp1), and transcription factor-IID (TFIID). The promoter also contains sites for inducible transcription factors, cAMP-responsive element (CRE)-binding protein (CREB), and Activator protein 2 (AP-2). We synthesized 10 different oligonucleotides for these and other potential transcription factor-binding sites. Using these oligonucleotides, we searched for binding proteins in young and old endothelial cells by electrophoretic mobility shift and supershift assays. Our results showed that AP-1 decreased with aging, but Sp1 and CREB1 were unaffected. However, decreased binding activities to CRE at positions -170 and -415 were shown in old cells. This could be explained by the decrease of AP-1 because these CREs bound not only CREB1 but also AP-1. Moreover, we observed that the binding activities of TFIID, CTF, and binding proteins to -40, -120 and -260 regions increased. These differential changes may cause the enhancement of FN expression in senescent cells.
authors.affiliation: National Institute of Bioscience and Human Technology, Agency of Industrial Science and Technology, Ibaraki, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Kumazaki
authors.forename: Y
authors.initials: Y
authors.lastname: Mitsui
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D002478:Cells, Cultured; D016922:Cellular Senescence; D016384:Consensus Sequence; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D004586:Electrophoresis; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology*; D005353:Fibronectins / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D011401:Promoter Regions, Genetic; D016385:TATA Box; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D005353:Fibronectins; D018808:Transcription Factor AP-1; D014157:Transcription Factors