May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
Homozygous Deletion of Pdlim2 Affects Corneal Epithelial Morphology and Cytoskeletal Organization
Author Affiliations & Notes
  • P. S. Zelenka
    LMDB, National Eye Inst/NIH, Bethesda, Maryland
  • C. Y. Gao
    LMDB, National Eye Inst/NIH, Bethesda, Maryland
  • M. Grusby
    Immunology and Infectious Disease, Harvard School of Public Health, boston, Massachusetts
  • S. Tomarev
    LMDB, National Eye Inst/NIH, Bethesda, Maryland
  • Footnotes
    Commercial Relationships  P.S. Zelenka, None; C.Y. Gao, None; M. Grusby, None; S. Tomarev, None.
  • Footnotes
    Support  NEI Intramural Program Z01 EY 00238-22
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 4304. doi:
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      P. S. Zelenka, C. Y. Gao, M. Grusby, S. Tomarev; Homozygous Deletion of Pdlim2 Affects Corneal Epithelial Morphology and Cytoskeletal Organization. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4304.

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

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Abstract

Purpose: : Pdlim2 is an E3 ubiquitin ligase, which contains an N-terminal PDZ motif and a C-terminal LIM domain. Previous studies have shown that Pdlim2 is expressed at very high levels in the corneal epithelium and interacts with α-actinins, filamin A, myosin VI, and MYH9. This study was undertaken to investigate Pdlim2 function in the cornea by determining its subcellular localization in wild type mouse corneas and by characterizing the corneal epithelial phenotype of Pdlim2 homozygous mice.

Methods: : Pdlim2 subcellular localization was investigated by confocal microscopy of immunostained, whole mounted corneas and by Total Internal Reflection Fluorescence (TIRF) microscopy of GFP-Pdlim2 transfected cells. Corneal epithelial protein expression profiles of wild type and knockout mice were compared by fluorescence difference (Cy3 vs. Cy5) 2D gel electrophoresis. Morphology was compared by light microscopy, and cytoskeletal organization of immunostained samples was determined by confocal microscopy.

Results: : Pdlim2 localized in a punctate pattern along the cell periphery in wild type corneal epithelia. TIRF microscopy demonstrated that GFP-Pdlim2 localized to cell-matrix adhesions and actin attachment sites. Knockout mice showed irregular thickening of the corneal epithelium, calcium deposits, breaks of Bowman’s membrane, and neovascularization between corneal epithelium and stroma. Confocal images of knockout corneas showed irregular actin cytoskeleton near the cell periphery in basal corneal epithelial cells. 2D fluorescence gel analysis revealed that destrin expression in Pdlim2 knockout is reduced more than 5 fold relative to wild type.

Conclusions: : The corneal phenotype of the Pdlim2 knockout mimics that of Corn1/Corn12J mutants, caused by mutations of the actin-depolymerizing factor, destrin, which is downregulated in Pdlim2 knockouts. These findings suggest that Pdlim2 may play a role in cytoskeletal regulation in corneal epithelial cells, possibly by regulating destrin function.

Keywords: cornea: epithelium • cytoskeleton • neovascularization 
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