April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Caveolin-1 Regulates Na+/K+-ATPase Activity in the Retinal Pigment Epithelium
Author Affiliations & Notes
  • M. H. Elliott
    Ophthalmology, Univ of Oklahoma Hlth Sci Ctr, Oklahoma City, Oklahoma
    Dean A McGee Eye Institute, Oklahoma City, Oklahoma
  • M. E. McClellan
    Ophthalmology, Univ of Oklahoma Hlth Sci Ctr, Oklahoma City, Oklahoma
    Dean A McGee Eye Institute, Oklahoma City, Oklahoma
  • M. N. A. Mandal
    Ophthalmology, Univ of Oklahoma Hlth Sci Ctr, Oklahoma City, Oklahoma
    Dean A McGee Eye Institute, Oklahoma City, Oklahoma
  • J. D. Ash
    Ophthalmology, Univ of Oklahoma Hlth Sci Ctr, Oklahoma City, Oklahoma
    Dean A McGee Eye Institute, Oklahoma City, Oklahoma
  • B. Nagel
    Dept of Pathology, St Louis Univ Hlth Sci Ctr, St Louis, Missouri
  • J. Ryerse
    Dept of Pathology, St Louis Univ Hlth Sci Ctr, St Louis, Missouri
  • S. J. Fliesler
    Ophthalmology, SUNY-Buffalo Ophthal/VA Medical Ctr, Buffalo, New York
  • Footnotes
    Commercial Relationships  M.H. Elliott, None; M.E. McClellan, None; M.N.A. Mandal, None; J.D. Ash, None; B. Nagel, None; J. Ryerse, None; S.J. Fliesler, None.
  • Footnotes
    Support  NIH RR017703; EY012190; EY007361, and Research to Prevent Blindness, Inc.
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 5970. doi:
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    • Get Citation

      M. H. Elliott, M. E. McClellan, M. N. A. Mandal, J. D. Ash, B. Nagel, J. Ryerse, S. J. Fliesler; Caveolin-1 Regulates Na+/K+-ATPase Activity in the Retinal Pigment Epithelium. Invest. Ophthalmol. Vis. Sci. 2010;51(13):5970.

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

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Abstract

Purpose: : Na+/K+-ATPase in RPE apical microvilli plays a critical role in maintaining the photoreceptor microenvironment. In other cell types, Na+/K+-ATPase can be found in caveolae, where it serves both as an ion transporter and ligand-inducible signal transducer. The α1 subunit of Na+/K+-ATPase contains putative binding motifs for the "scaffolding domain" of the caveolar coat protein, caveolin-1 (Cav-1). We tested the hypothesis that Na+/K+-ATPase interacts with and is regulated by Cav-1 in RPE.

Methods: : Caveolar membranes were isolated from bovine RPE and cofractionation of the Na/K-ATPase with Cav-1 was assayed by quantitative Western analysis. Bovine RPE microsomes were incubated with a peptide corresponding to the Cav-1 scaffolding domain ("Cav-SCD"), with a scrambled control peptide ("Cav-X"), or with no peptide, and total (ouabain-sensitive) Na+/K+-ATPase activity was determined. Separately, microsomes from Cav-1 null (KO) and control mouse eyecups were assayed for Na/K-ATPase activity. Correlative confocal immunohistochemistry of eyes from KO and control mice was performed to localize Na+/K+-ATPase and Cav-1 in RPE. The presence of caveolae in RPE was also assessed by TEM. Gene expression analysis with qRT-PCR validation was performed on KO and control mouse eyecups.

Results: : Western analysis indicated ~60% of the total Na+/K+-ATPase α1 subunit cofractionates with Cav-1-enriched membranes. Na+/K+-ATPase and Cav-1 colocalized to the base of the apical RPE microvilli in both KO and control mice, coincident with ultrastructurally identified caveolae. Total Na+/K+-ATPase activity was increased 1.7-fold in KO RPE microsomes relative to controls; this corresponded to a decrease in affinity for K+, but not for Na+. Incubation of bovine RPE microsomes with Cav-SCD significantly inhibited Na+/K+-ATPase activity (3-fold, compared to Cav-X). In KO eyecups, microarray, qRT-PCR, and Western analyses revealed dramatic up-regulation of FXYD3, a regulator of Na+/K+-ATPase.

Conclusions: : Our results suggest that Cav-1 associates with and is an endogenous regulator of Na+/K+-ATPase activity in RPE. The mechanism of this regulation does not involve changes in Na+/K+-ATPase localization or overall protein expression. Rather, regulation appears to be via interaction of Cav-1 with Na+/K+-ATPase.

Keywords: retinal pigment epithelium • NaK ATPase • ion transporters 
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