title: Raf 1 represses expression of the tight junction protein occludin via activation of the zinc-finger transcription factor slug.
abstract: Although dysregulation of tight junction (TJ) proteins is observed in epithelial malignancy, their participation in epithelial transformation is poorly understood. Recently we demonstrated that expression of oncogenic Raf 1 in Pa4 epithelial cells disrupts TJs and induces an oncogenic phenotype by downregulating expression of the TJ protein, occludin. Here we report the mechanism by which Raf 1 regulates occludin expression. Raf 1 inhibited occludin transcription by repressing a minimal segment of the occludin promoter in concert with upregulation of the transcriptional repressor, Slug without influencing the well-documented transcriptional repressor, Snail. Overexpression of Slug in Pa4 cells recapitulated the effect of Raf 1 on occludin expression, and depletion of Slug by small interfering RNA abrogated the effect of Raf 1 on occludin. Finally, chromatin immunoprecipitation assays and site-directed mutagenesis demonstrated a direct interaction between Slug and an E-box within the minimal Raf 1-responsive segment of the occludin promoter. These findings support a role of Slug in mediating Raf 1-induced transcriptional repression of occludin and subsequent epithelial to mesenchymal transition.
authors.affiliation: Epithelial Pathobiology Research Unit, Department of Pathology, Emory University, Atlanta, GA 30322, USA.
authors.forename: Z
authors.initials: Z
authors.lastname: Wang
authors.forename: P
authors.initials: P
authors.lastname: Wade
authors.forename: K J
authors.initials: KJ
authors.lastname: Mandell
authors.forename: A
authors.initials: A
authors.lastname: Akyildiz
authors.forename: C A
authors.initials: CA
authors.lastname: Parkos
authors.forename: R J
authors.initials: RJ
authors.lastname: Mrsny
authors.forename: A
authors.initials: A
authors.lastname: Nusrat
mesh_terms: D000818:Animals; D047369:Chromatin Immunoprecipitation; D015536:Down-Regulation; D024721:E-Box Elements; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation; D006801:Humans; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008565:Membrane Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D062793:Occludin; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D051381:Rats; D012097:Repressor Proteins / Q000378:metabolism*; D000071250:Snail Family Transcription Factors; D019108:Tight Junctions / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D016335:Zinc Fingers
chemical_list: D004791:Enzyme Inhibitors; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; C568987:Ocln protein, rat; D012097:Repressor Proteins; C000604407:SNAI1 protein, human; C000604450:Snai2 protein, rat; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors; D019908:Proto-Oncogene Proteins c-raf