September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Identification of Protein-Protein Interactors of The Corneal Dystrophy-Causing Protein, SLC4A11
Author Affiliations & Notes
  • Nada Ahmed Alshumaimeri
    Biochemistry, University of Alberta, Edmonton, Alberta, Canada
  • Joseph R Casey
    Biochemistry, University of Alberta, Edmonton, Alberta, Canada
  • Footnotes
    Commercial Relationships   Nada Alshumaimeri, None; Joseph Casey, None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 4802. doi:
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      Nada Ahmed Alshumaimeri, Joseph R Casey; Identification of Protein-Protein Interactors of The Corneal Dystrophy-Causing Protein, SLC4A11. Invest. Ophthalmol. Vis. Sci. 2016;57(12):4802.

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

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Purpose : SLC4A11 is a membrane transport protein found at the basolateral surface of corneal endothelial cells. Mutations of SLC4A11 cause Endothelial Corneal Dystrophies (ECD), specifically Fuchs ECD and Congenital Hereditary Endothelial Dystrophy (CHED), both marked by endothelial cell loss and corneal stromal edema. We hypothesize that protein interactors of SLC4A11 influence the function and localization of SLC4A11. Identifying SLC4A11-protein interactors will lead us to understand the full physiological function of SLC4A11.

Methods : The Membrane Yeast two Hybrid (MYTH) technique was used to identify SLC4A11-protein interactors. A cDNA library was constructed from bovine cornea mRNA and cloned into the prey vector (pPR3-N). Yeast were transformed with human SLC4A11 cDNA cloned in bait vector (pCMBV). The ability of cotransformed yeast to grow on auxotrophic selective plates indicated colonies where SLC4A11 (Bait) interacted with prey protein. Candidate interacting proteins were further tested for their ability to interact with SLC4A11 by co-immunoprecipitation when expressed in HEK293 cells and in corneal lysates. In addition, co-localization of the interactors was assessed in corneal tissue, using confocal immunofluorescence.

Results : We screened 59 x 106 total transformants. 431 clones grew as interactors. 273 clones remaining as interactors after confirmatory screening were then sequenced. This left 8 confirmed, unique candidate SLC4A11 interactors. One of the promising hits is the transmembrane protein (TM254) that appeared several times.

Conclusions : The identified TM254 protein has an unknown function. Since TM254 is related to Glycophorin, a chaperone protein for Band 3 protein (AE1/SLC4A1) we hypothesize that TM254 acts to chaperone the folding and trafficking of SLC4A11 from the site of biosynthesis (endoplasmic reticulum) to the plasma membrane. One other potential interactor is involved in cell-matrix interaction that might explain the loss of endothelial cells in ECDs.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.


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