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A. V. Matveev, A. Quiambao, J. Fitzgerald, X.-Q. Ding; Native Cone Photoreceptor Cyclic Nucleotide-Gated Channel Is a Heterotetrameric Complex With Potential Distinct Modulation Mechanisms. Invest. Ophthalmol. Vis. Sci. 2008;49(13):2413. doi: https://doi.org/.
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Cone vision mediated by photoreceptor cyclic nucleotide-gated (CNG) channel activation is essential for central and color vision and visual acuity. Mutations in genes encoding the cone CNG channel subunits, CNGA3 and CNGB3, have been linked to various forms of achromatopsia and progressive cone dystrophy in humans. This study investigates the biochemical characteristics of native cone CNG channels, using the cone-dominant retina in mice deficient in the transcription factor neural retina leucine zipper (Nrl).
Expression and photoreceptor localization of CNG channel and cone Na+/Ca2+-K+ exchanger (NCKX2) in the mouse retinas were examined by immunoblotting and immunolabeling of the retinal sections. Co-immunoprecipitation was used to detect the inter-subunit interaction and the association between the cone CNG channel and NCKX2. Chemical cross-linking using thiol-specific and amino-specific cross-linkers were used to analyze the channel complexes and the affinity binding was used to determine association of the cone CNG channel with the calmodulin.
Abundant expression of CNGA3 and CNGB3, but no rod CNG channel expression, was detected in Nrl-/- retina. Blue and red/green cone localization of the channel was shown in the retinas of wild type and Nrl-/- mice. Co-immunoprecipitation detected a direct interaction between CNGA3 and CNGB3 but association between the channel and NCKX2 was not evident. Chemical cross-linking readily generated products at sizes consistent with oligomers of the channel complexes, ranging from dimeric to tetrameric complexes. Unlike the modulation of rod CNG channel, calmodulin was shown to bind to CNGA3 but not CNGB3.
This study provides the first biochemical evidence showing the inter-subunit interaction and the presence of hetero-tetrameric complexes of the native cone CNG channel in retina. The observations of calmodulin binding to CNGA3 but not CNGB3 and lack of association between CNGA3 and NCKX2 may implicate a distinct mechanism of CNG channel modulation and Ca2+ homeostasis in cones.
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