May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
Confocal Fluorescence Microscopy Studies of R120G alphaB-Crystallin and Lens Intermediate Filaments
Author Affiliations & Notes
  • S. Song
    Ophthalmic Research/Surgery, Brigham and Women's Hospital/Harvard Medical School, Boston, Massachusetts
  • J. Liang
    Ophthalmic Research/Surgery, Brigham and Women's Hospital/Harvard Medical School, Boston, Massachusetts
  • B.-F. Liu
    Ophthalmic Research/Surgery, Brigham and Women's Hospital/Harvard Medical School, Boston, Massachusetts
  • L. Chylack, Jr.
    Ophthalmic Research/Surgery, Brigham and Women's Hospital/Harvard Medical School, Boston, Massachusetts
  • Footnotes
    Commercial Relationships  S. Song, None; J. Liang, None; B. Liu, None; L. Chylack, None.
  • Footnotes
    Support  NIH grant (EY13968) & Massachusetts Lions Eye Research Fund
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 5027. doi:https://doi.org/
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    • Get Citation

      S. Song, J. Liang, B.-F. Liu, L. Chylack, Jr.; Confocal Fluorescence Microscopy Studies of R120G alphaB-Crystallin and Lens Intermediate Filaments. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5027. doi: https://doi.org/.

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

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Abstract

Purpose: : The R120G mutation of αB-crystallin is known to cause desmin-related myopathy but the mechanisms underlying the formation of cataract are not clearly established. We speculate that alteration of the protein-protein interaction between R120G αB-crystallin and lens intermediate filament proteins is one of the mechanisms for the congenital cataract.

Methods: : Protein-protein interaction was determined by confocal fluorescence resonance energy transfer (FRET) microscopy using green fluorescence protein (GFP) as the donor and red fluorescence protein (RFP) as the acceptor. Lens intermediate filament (IF) protein gene (vimentin, filensin or CP49) was fused into GFP vector and αB-crystallin (WT or R120G mutant) gene was fused into the RFP vector. The donor-acceptor plasmid pairs of IF-GFP and αB-RFP were co-transfected into HeLa cells. After incubation, confocal fluorescence images of the transfected cells were taken. Protein-protein interaction was evaluated by FRET between the donor and the acceptor.

Results: : The confocal fluorescence images showed that the R120G αB-crystallin expression contained large amounts of protein aggregates with either vimentin, or filensin, or CP49 protein. FRET analyses indicated that R120G αB-crystallin had a significantly greater protein-protein interaction than WT αB-crystallin.

Keywords: crystallins • cytoskeleton • protein structure/function 
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