July 2018
Volume 59, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2018
SLC4A11 extracellular Loop 3 in corneal endothelial cell adhesion, FECD pathology and therapeutics
Author Affiliations & Notes
  • Darpan Malhotra
    Biochemistry , University of Alberta, Edmonton, Alberta, Canada
  • Martin Jung
    Medizinische Biochemie und Molekularbiologie, Universität des Saarlandes, Homburg, Saarland, Germany
  • Sergei Noskov
    Department of Biological Sciences, University of Calgary, Calgary, Alberta, Canada
  • Richard Zimmermann
    Medizinische Biochemie und Molekularbiologie, Universität des Saarlandes, Homburg, Saarland, Germany
  • Joseph Roman Casey
    Biochemistry , University of Alberta, Edmonton, Alberta, Canada
  • Footnotes
    Commercial Relationships   Darpan Malhotra, None; Martin Jung, None; Sergei Noskov, None; Richard Zimmermann, None; Joseph Casey, None
  • Footnotes
    Support  CIHR Grant
Investigative Ophthalmology & Visual Science July 2018, Vol.59, 4433. doi:https://doi.org/
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Darpan Malhotra, Martin Jung, Sergei Noskov, Richard Zimmermann, Joseph Roman Casey; SLC4A11 extracellular Loop 3 in corneal endothelial cell adhesion, FECD pathology and therapeutics. Invest. Ophthalmol. Vis. Sci. 2018;59(9):4433. doi: https://doi.org/.

      Download citation file:


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

      ×
  • Supplements
Abstract

Purpose : Mutations in SLC4A11 cause some cases of Fuchs endothelial corneal dystrophy (FECD) and congenital hereditary endothelial dystrophy (CHED). We found that SLC4A11 contributes to corneal endothelial cell (CEC) adhesion to its underlying basement membrane, Descemet’s membrane (DM). Loss of the adhesion role may explain the pathophysiology of these dystrophies. Here, we identify the SLC4A11 region involved in CEC adhesion and its interaction partner in DM. We further test SLC4A11-extracellular loop 3 (EL3) as a potential therapeutic target to treat FECD and CHED using chimeric proteins.

Methods : HEK293 cells expressing SLC4A11 or control cells were pre-incubated with antibody targeting a region of SLC4A11-EL3 and tested in cell adhesion assays (CAA) (n=12). Human, bovine and murine SLC4A11 were tested in CAA (n=8). We created an energy-minimized 3D molecular model of SLC4A11-EL3 refined by Replica-Exchange Molecular Dynamics Simulations to map the FECD mutations. We constructed a small chimeric protein (15 kDa) of Glycophorin-A precursor and SLC4A11-EL3 (GPA-EL3) as a potential therapeutic for viral delivery, tested for expression and further in CAA (n=8). SLC4A11 interaction with DM components, COL8A1 and COL8A2, was assessed and the binding region were determined using peptide arrays. One-way and two-way ANOVA was used for statistical analysis.

Results : Anti-EL3 antibody prevented SLC4A11-mediated cell adhesion (p<0.0001), without any effect on control cells. Pre-immune serum, used as a negative control, had no effect on cell adhesion. Bovine and murine SLC4A11 promoted cell adhesion to similar levels as their human ortholog when compared to vector controls (p<0.0001). The 3D molecular model of EL3 mapped the three FECD mutations to a cluster where they may disrupt the interaction by causing conformational changes in EL3. GPA-EL3 chimera expressed as a dimer in the HEK293 cells and promoted cell-adhesion to similar levels as full length SLC4A11 (p<0.0001). COL8A1and COL8A2 were confirmed as SLC4A11 interactors using peptide arrays and their binding regions were identified.

Conclusions : SLC4A11-EL3 contributes to anchoring CECs to DM. Disease mutations in EL3 disrupt SLC4A11-mediated cell adhesion. We propose loss of SLC4A11-mediated cell adhesion as a mechanism of FECD pathophysiology. GPA-EL3 chimera has the potential to be tested as a therapeutic via gene delivery to alleviate FECD pathology.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.

×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×