December 2002
Volume 43, Issue 13
ARVO Annual Meeting Abstract  |   December 2002
Interaction of Lens Connexins with ZO-1
Author Affiliations & Notes
  • NM Kumar
    Cell Biology Univ Illinois-Chicago Chicago IL
  • A Baruch
    Cell Biology Scripps Research Institute La Jolla CA
  • BN G Giepmans
    Division of Cellular Biochemistry The Netherland Cancer Institute Amsterdam Netherlands
  • V Shestopalov
    Ophthalmology Washington University School of Medicine St Louis MO
  • I Dunia
    Institut Jacques Monod CNRS-Universites Paris 6-7 Paris France
  • EL Benedetti
    Institut Jacques Monod CNRS-Universites Paris 6-7 Paris France
  • PA Nielsen
    Cell Biology Scripps Research Institute La Jolla CA
  • Footnotes
    Commercial Relationships   N.M. Kumar, None; A. Baruch, None; B.N.G. Giepmans, None; V. Shestopalov, None; I. Dunia, None; E.L. Benedetti, None; P.A. Nielsen, None. Grant Identification: Danish Medical Research Council(PAN);NIH Grant EY12142 and EY13605(NMK)
Investigative Ophthalmology & Visual Science December 2002, Vol.43, 4800. doi:
  • Views
  • Share
  • Tools
    • Alerts
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      NM Kumar, A Baruch, BN G Giepmans, V Shestopalov, I Dunia, EL Benedetti, PA Nielsen; Interaction of Lens Connexins with ZO-1 . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4800.

      Download citation file:

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

  • Supplements

Abstract: : Purpose: The MAGUK protein ZO-1 has previously been shown to bind Cx43α1 both in vitro and in vivo.The similarity of the most C-terminal residues of the lens connexins Cx46α3 and Cx50α8 to those of Cx43α1, prompted us to examine if ZO-1 is expressed in the mouse lens, and if ZO-1 interacts with the lens gap junctions. Methods: Constructs containing the carboxyl terminal domains of mouse Cx43α1, Cx46α3 or Cx50α8 and the various PDZ domains of ZO-1 were expressed individually in E.coli to produce GST fusion proteins. Mouse lens homogenates were used in pull-down assays with fusion proteins or for co-immunoprecipatation with an anti-ZO-1 antibody. Colocalization of ZO-1 with lens gap junctions was determined by immunofluorescence or EM of SDS freeze fracture labeling of mouse lens sections, using different fluorochromes or different sized gold particles respectively, to visualize anti-connexin and anti-ZO-1 antibodies. Results: ZO-1 transcripts were detected in lens RNA by Northern analysis. By Western analysis, ZO-1 was detected in mouse lens homogenates from the epithelial, cortical and nuclear layers and had the greatest abundance in younger animals. Confocal immunofluorescence analysis of mouse lens sections showed a partial colocalization of ZO-1 with either Cx46α3 or Cx50α8 in lens fiber cells. EM confirmed the observed colocalization on SDS freeze-fracture labeling replicas of mouse differentiating lens fiber plasma membranes. ZO-1 was shown to co-immunoprecipitate with either Cx46α3 or Cx50α8 from mouse lens homogenates. The second PDZ domain of ZO-1 bound both Cx46α3 and Cx50α8 from mouse lens homogenates, where as no binding was observed with the first and third PDZ domains. Mutation of the extreme C-terminal residue of Cx46α3 or Cx50α8 abolished the interaction with ZO-1. Furthermore, using pull-down assays, the C-termini of Cx43α1, Cx46α3 and Cx50α8 when expressed as bacterial GST fusion proteins were demonstrated to bind to the second PDZ domain of ZO-1 fused to a His tag, although with different efficiencies. Conclusion: These results indicate that ZO-1 is expressed in the lens and may interact with lens connexins in a manner similar to the interaction previously described between ZO-1 and Cx43α1. Further studies are needed to elucidate the potential regulation and biological significance of these interactions.

Keywords: 416 gap junctions/coupling • 580 signal transduction • 339 cell adhesions/cell junctions 

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.