title: Minimal essential region for krüppel-like factor 5 expression and the regulation by specificity protein 3-GC box binding.
abstract: Krüppel-like factor 5 (KLF5) transcriptionally controls the proliferation-differentiation balance of epithelium and is overexpressed in carcinomas. Although genomic region modifying KLF5 expression is widespread in different types of cells, the region that commonly regulates basal expression of the genes across cell-types is uncertain. In this study we determined the minimal essential region for the expression and its regulatory transcription factors using oral carcinoma cells. A reporter assay defined a 186bp region downstream of the transcription start site and a cluster of six GC boxes (GC1-GC6) as the minimal essential region. Mutation in the GC1 or GC6 regions but not other GC boxes significantly decreased the reporter expression. The decrease by the GC1 mutation was reproduced in the 2kbp full-length promoter, but not by the GC6 mutation. Additionally, specificity proteins (Sp) that can be expressed in epithelial cells and bind GC box, Sp3 co-localized with KLF5 in oral epithelium and carcinomas and chromatin immunoprecipitation analyses showed Sp3 as the prime GC1-binding protein. Inhibition of Sp-GC box binding by mithramycin A and knockdown of Sp3 by the short interfering RNA decreased expression of the reporter gene and endogenous KLF5. These data demonstrate that a 186bp region is the minimal essential region and that Sp3-GC1 binding is essential to the basal expression of KLF5.
authors.affiliation: Department of Biochemistry, The Nippon Dental University School of Life Dentistry at Tokyo, Tokyo, Japan. Electronic address: leschang@tky.ndu.ac.jp.
authors.forename: Nozomi
authors.initials: N
authors.lastname: Mihara
authors.affiliation: Department of Biochemistry, The Nippon Dental University School of Life Dentistry at Tokyo, Tokyo, Japan. Electronic address: tchiba@tky.ndu.ac.jp.
authors.forename: Tadashige
authors.initials: T
authors.lastname: Chiba
authors.affiliation: Department of Biochemistry, The Nippon Dental University School of Life Dentistry at Tokyo, Tokyo, Japan. Electronic address: kyama@tky.ndu.ac.jp.
authors.forename: Kosuke
authors.initials: K
authors.lastname: Yamaguchi
authors.affiliation: Department of Biochemistry, The Nippon Dental University School of Life Dentistry at Tokyo, Tokyo, Japan. Electronic address: sudoh@tky.ndu.ac.jp.
authors.forename: Haruka
authors.initials: H
authors.lastname: Sudo
authors.affiliation: Division of Oral Diagnosis, Dental and Maxillofacial Radiology and Oral Pathology Diagnostic Services, The Nippon Dental University Hospital, Tokyo, Japan. Electronic address: h-yagishita@tky.ndu.ac.jp.
authors.forename: Hisao
authors.initials: H
authors.lastname: Yagishita
authors.affiliation: Department of Biochemistry, The Nippon Dental University School of Life Dentistry at Tokyo, Tokyo, Japan. Electronic address: kimai@tky.ndu.ac.jp.
authors.forename: Kazushi
authors.initials: K
authors.lastname: Imai
mesh_terms: D002294:Carcinoma, Squamous Cell / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006801:Humans; D051741:Kruppel-Like Transcription Factors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D051705:Sp3 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D024363:Transcription Initiation Site
chemical_list: C492214:KLF5 protein, human; D051741:Kruppel-Like Transcription Factors; C497121:SP3 protein, human; D016329:Sp1 Transcription Factor; C108209:SP1 protein, human; D051705:Sp3 Transcription Factor
keywords: Cancer; Gene transcription; Transcription factor; Transcription promoter