Raf 1 represses expression of the tight junction protein occludin via activation of the zinc-finger transcription factor slug.
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.
Wang
Z
Z
Epithelial Pathobiology Research Unit, Department of Pathology, Emory University, Atlanta, GA 30322, USA.
Wade
P
P
Mandell
K J
KJ
Akyildiz
A
A
Parkos
C A
CA
Mrsny
R J
RJ
Nusrat
A
A
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
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