May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
The Loss-of-Function Cx50-R205G Mutation Dominantly Alters Wild-Type Lens Connexin Function
Author Affiliations & Notes
  • A. M. DeRosa
    Physiology and Biophysics, State Univ of NY-Stony Brook, Stonybrook, New York
  • C.-H. Xia
    School of Optometry and Vision Science Program, University of California at Berkeley, Berkeley, California
  • X. Gong
    School of Optometry and Vision Science Program, University of California at Berkeley, Berkeley, California
  • T. W. White
    Physiology and Biophysics, State Univ of NY-Stony Brook, Stonybrook, New York
  • Footnotes
    Commercial Relationships  A.M. DeRosa, None; C. Xia, None; X. Gong, None; T.W. White, None.
  • Footnotes
    Support  NIH Grant EY13163, NIH Grant EY13849
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 1522. doi:https://doi.org/
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      A. M. DeRosa, C.-H. Xia, X. Gong, T. W. White; The Loss-of-Function Cx50-R205G Mutation Dominantly Alters Wild-Type Lens Connexin Function. Invest. Ophthalmol. Vis. Sci. 2008;49(13):1522. doi: https://doi.org/.

      Download citation file:


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

      ×
  • Supplements
Abstract

Purpose: : Recent discovery of a novel Cx50 mutation was found to cause an autosomal semi-dominant cataract in mice. Here, we use the dual whole-cell voltage clamp technique in conjunction with the paired Xenopus oocyte assay to analyze the functional consequences of the Cx50-R205G mutation.

Methods: : Murine Cx50 mutated at amino acid 205 (arginine to glycine) was subcloned in pCS2+, expressed in HeLa cells or transcribed in vitro and injected in paired Xenopus oocytes . Protein expression was confirmed, and quantified by immunoblotting and immunohistochemistry. The paired Xenopus oocyte system was used in conjunction with the dual whole-cell voltage clamp technique to analyze differences in macroscopic junctional conductance (Gj), channel kinetics, and voltage gating sensitivity.

Results: : These experiments revealed that the R205G mutant acts as a dominant negative inhibitor of wild-type Cx50 with regard to channel function. Additionally, the Cx50-R205G mutant dominantly altered wild-type Cx46 channel gating when the two connexins were co-expressed in vitro to mimic heteromeric channel formation. Interestingly, the mutant R205G subunits were able to properly localize to the plasma membrane in transfected HeLa cells and showed no discernable reduction in translation compared to the wild-type Cx50 homolog.

Conclusions: : These data define, for the first time, the electrophysiological properties of functional gap junctions formed by the combination of Cx50 or Cx46 and Cx50-R205G mutant proteins. Taken together, these findings reveal that the Cx50-R205G mutation acts as a loss-of-function mutation with a strong dominant inhibition toward wild-type Cx50. Conversely, this mutant is able to interact with wild-type Cx46 to form functional intercellular channels with altered electrophysiological properties, a phenomenon that may play a role in the development of the unique cataract phenotype seen in vivo.

Keywords: cataract • gap junctions/coupling • cell-cell communication 
×
×

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.

×