May 2005
Volume 46, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2005
Over–Expression of Pax6 Retards Cell Cycle Progression and Reduces Cell Migration Rate in Corneal Epithelial Cells
Author Affiliations & Notes
  • J. Ouyang
    Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, FL
    Physiology/Biophysics, University of Miami School of Medicine, Miami, FL
  • S.–Y. Cheng
    Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, FL
  • L.–K. Yeh
    Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, FL
  • C.–Y. Liu
    Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, FL
  • M.E. Fini
    Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, FL
  • Footnotes
    Commercial Relationships  J. Ouyang, None; S. Cheng, None; L. Yeh, None; C. Liu, None; M.E. Fini, None.
  • Footnotes
    Support  EY014801,EY12486,Walter G. Ross Foundation, RPB
Investigative Ophthalmology & Visual Science May 2005, Vol.46, 2595. doi:
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    • Get Citation

      J. Ouyang, S.–Y. Cheng, L.–K. Yeh, C.–Y. Liu, M.E. Fini; Over–Expression of Pax6 Retards Cell Cycle Progression and Reduces Cell Migration Rate in Corneal Epithelial Cells . Invest. Ophthalmol. Vis. Sci. 2005;46(13):2595.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Abstract: : Purpose: Transcription factor Pax6 resides at the top of a regulatory hierarchy controlling formation of the eye during embryological development. Pax6 is also expressed in several tissues of the adult eye. The goal of this study was to learn more about the role of Pax6 in the corneal epithelium. Methods: Pax6 dosage is critical for proper formation of the eye. Thus to study its role in corneal epithelial cells, we increased its expression level. A rabbit corneal epithelial cell line, SIRC, was used to establish stable transformant clones carrying a mouse wild–type Pax6 full–length coding region or a mutant Pax6, lacking the c–terminal transactivation domain. The full–length mPax6 or mutant mPax6 transgene was regulated using a doxycycline–based (tet–on) inducible gene expression system. Expression of Pax6 protein or truncated Pax6 protein was induced by adding doxycycline to the culture medium. A cell growth curve, cell cycle profile, cell proliferation index, cell migration, and apoptosis was studied. In all experiments, induced and non–induced tetO–Pax6 or tetO–delta286 transformed SIRC cells were compared. Results: Pax6 over–expression has been reported to retard or inhibit the cell cycle in lens cells and neurons. We found that over–expression of Pax6 protein also retards the SIRC cell cycle. Consistent with this, retinoblastoma tumor suppression protein (pRB) was up–regulated as judged by immuno–precipitation analysis. In contrast, truncated Pax6 protein without the transactivation domain did not affect the cell cycle. Pax6 over–expression induced apoptosis in a small fraction of cells. Western blotting experiments suggested that this effect was mediated via a caspase–3 independent pathway. In addition, we previously reported that Pax6 levels and DNA binding activity increase at the epithelial front migrating to heal a corneal wound, and migration is faster in "small eye" mice with decreased Pax6 dosage. Consistent with this, over–expression of wild–type but not mutant Pax6 inhibited the rate of SIRC cell migration. Conclusions: These results suggest that normal dosage of Pax6 protein plays a pivotal role in the maintenance of corneal epithelial cell function. The feature of cell cycle retardation in Pax6 over–expressing cells is consistent with the notion that Pax6 may control the rate of epithelial turnover and repair and contribute to the slow cycling of stem cells. The effects on cell migration are consistent with a proposed role in controlling corneal re–epithelialization.

Keywords: cornea: epithelium • gene/expression • cornea: basic science 
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