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Youwen Zhang, Minh-Thu Nguyen, Hongjun Wei, Timothy W. Kraft, Steven J. Pittler; Characterization of Transgenic Mice Overexpressing Cngb1 Locus Encoded Garp1 in the Retina. Invest. Ophthalmol. Vis. Sci. 2011;52(14):53.
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© ARVO (1962-2015); The Authors (2016-present)
The Cngb1 locus encodes the rod cGMP-gated cation channel β-subunit and two alternatively spliced glutamic acid-rich proteins, Garp1 and Garp2. We previously showed that knock-out (KO) of all three proteins (Cngb1-X1) exhibits ectopic disk morphogenesis, loss of structural integrity and progressive retinal degeneration. Overexpression of Garp2 (Garp2-Tg) also effects disk morphogenesis and alters phototransduction kinetics. Transgenic mice overexpressing Garp1 were created to understand its structural and functional roles in the retina.
A murine Garp1 transgene construct was generated containing the entire Garp1 protein coding sequence under control of a 4.4 kb mouse rhodopsin promoter. Four transgenic lines were confirmed by PCR-based genotyping, and expression was confirmed and quantified by Western blot analysis in two lines. Light microscopy, OCT, fundus imaging, immunofluorescence, and ERG analyses were performed.
Garp1 levels were at least 80 fold greater than wild type in two lines. Using a murine Garp1 specific antibody rod outer segments (ROS) were strongly labeled and the outer plexiform layer was less intensely but consistently labeled. In WT mice Garp1 labeling was not above background levels. Morphologically the transgenic retina appeared normal with no apparent ROS thickening or shorterning as observed in Garp2-Tg mice. Dark and light-adapted ERG analyses in 2-3 month old transgenic mice indicated normal retina function.
High level overexpression of Garp1 in the rod photoreceptor in WT mice does not appear to cause light-level morphological or gross functional changes in the retina.
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