April 2009
Volume 50, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2009
Expression of KCNQ and KCNE Proteins in Bovine Retinal Pigment Epithelium
Author Affiliations & Notes
  • X. Zhang
    Ophthalmology and Visual Sciences, University of Michigan, Ann arbor, Michigan
  • B. A. Hughes
    Ophthalmology and Visual Sciences and Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan
  • Footnotes
    Commercial Relationships  X. Zhang, None; B.A. Hughes, None.
  • Footnotes
    Support  NIH Grant R01-EY08850, NIH Grant P30-EY07003, and RPB
Investigative Ophthalmology & Visual Science April 2009, Vol.50, 2923. doi:
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      X. Zhang, B. A. Hughes; Expression of KCNQ and KCNE Proteins in Bovine Retinal Pigment Epithelium. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2923.

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Abstract

Purpose: : In previous electrophysiological studies, we identified in freshly isolated bovine and primate retinal pigment epithelial (RPE) cells a sustained outwardly rectifying K+ current with properties resembling those of M-currents, which are mediated by channels composed of pore-forming KCNQ and auxiliary KCNE subunits. We also reported the detection by RT-PCR of transcripts for KCNQ1, KCNQ4, and KCNQ5 in monkey and human RPE and, in monkey RPE, KCNE1, KCNE2, KCNE3, and KCNE4. The objective of this study was to determine whether KCNQ and KCNE proteins are expressed in the RPE.

Methods: : KCNQ and KCNE expression was examined by Western analysis using plasma membrane proteins from bovine RPE and positive control tissues (skeletal muscle and heart) in conjunction with goat anti-KCNQ1 and anti-KCNQ4 antibodies and rabbit anti-KCNQ5, anti-KCNE1, anti-KCNE2, anti-KCNE3, and anti-KCNE4 antibodies. Anti-Na+/K+-ATPase and anti-SAP97 antibodies served as apical and basolateral membrane protein markers, respectively.

Results: : We detected in bovine RPE KCNQ5, KCNE1, KCNE2, and KCNE3, but not KCNQ1, KCNQ4, or KCNE4. We also detected Na+/K+-ATPase and SAP97, indicating that the bovine RPE plasma membrane fraction contained both apical and basolateral membrane proteins. KCNQ1 and KCNQ4 were observed in heart and skeletal muscle, respectively, indicating that the failure to detect these proteins in bovine RPE was not due to the lack of antibody reactivity in this species.

Conclusions: : The results demonstrate that KCNQ5, KCNE1, KCNE2 and KCNE3 proteins are expressed in bovine RPE, but that KCNQ1, KCNQ4, and KCNE4 are not or are present at levels below detection. We conclude that KCNQ5 is a major component of the M-type channel in bovine RPE and that its co-assembly with KCNE1, KCNE2, or KCNE3 may contribute to the channel’s properties.

Keywords: retinal pigment epithelium • electrophysiology: non-clinical 
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