title: Ectopic Mitf in the embryonic chick retina by co-transfection of β-catenin and Otx2.
abstract: OBJECTIVE
  Development of the retinal pigment epithelium (RPE) is controlled by intrinsic and extrinsic regulators including orthodenticle homeobox 2 (Otx2) and the Wnt/β-catenin pathway, respectively. Otx2 and β-catenin are necessary for the expression of the RPE key regulator microphthalmia-associated transcription factor (Mitf); however, neither factor is sufficient to promote Mitf expression in vivo. The study was conducted to determine whether Otx2 and β-catenin act in a combinatorial manner and tested whether co-expression in the presumptive chick retina induces ectopic Mitf expression.
  
  
  METHODS
  The sufficiency of Wnt/β-catenin activation and/or Otx2 expression to induce RPE-specific gene expression was examined in chick optic vesicle explant cultures or in the presumptive neural retina using in ovo-electroporation. Luciferase assays were used to examine the transactivation potentials of Otx2 and β-catenin on the Mitf-D enhancer and autoregulation of the Mitf-D and Otx2T0 enhancers.
  
  
  RESULTS
  In optic vesicles explant cultures, RPE-specific gene expression was activated by lithium chloride, a Wnt/β-catenin agonist. However, in vivo, Mitf was induced only in the presumptive retina if both β-catenin and Otx2 are co-expressed. Furthermore, both Mitf and Otx2 can autoregulate their own enhancers in vitro.
  
  
  CONCLUSIONS
  The present study provides evidence that β-catenin and Otx2 are sufficient, at least in part, to convert retinal progenitor cells into presumptive RPE cells expressing Mitf. Otx2 may act as a competence factor that allows RPE specification in concert with additional RPE-promoting factors such as β-catenin.
authors.affiliation: Department of Ophthalmology and Visual Sciences, Moran Eye Center, University of Utah, Salt Lake City, Utah 84132, USA.
authors.forename: Peter D
authors.initials: PD
authors.lastname: Westenskow
authors.forename: Jon B
authors.initials: JB
authors.lastname: McKean
authors.forename: Fumi
authors.initials: F
authors.lastname: Kubo
authors.forename: Shinichi
authors.initials: S
authors.lastname: Nakagawa
authors.forename: Sabine
authors.initials: S
authors.lastname: Fuhrmann
mesh_terms: D000818:Animals; D002452:Cell Count; D002454:Cell Differentiation; D002478:Cells, Cultured; D002642:Chick Embryo; D018274:Electroporation; D019084:Fluorescent Antibody Technique, Indirect; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D051739:Microphthalmia-Associated Transcription Factor / Q000235:genetics*; D018613:Microscopy, Confocal; D051857:Otx Transcription Factors / Q000235:genetics*; D010957:Plasmids; D012160:Retina / Q000196:embryology*; D055351:Retinal Neurons / Q000166:cytology; D055213:Retinal Pigment Epithelium / Q000166:cytology* / Q000378:metabolism; D015533:Transcriptional Activation; D014162:Transfection; D051153:Wnt Proteins / Q000235:genetics; D051176:beta Catenin / Q000235:genetics*
chemical_list: D051739:Microphthalmia-Associated Transcription Factor; D051857:Otx Transcription Factors; D051153:Wnt Proteins; D051176:beta Catenin
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