The long term objectives of the research program are to understand protein structure/function relationships in retinal photoreceptors, and to translate this understanding to the treatment of patients with retinitis pigmentosa and related disorders. In humans, CNGB1 gene defects cause hereditary retinal degeneration. We have therefore focused on the murine Cngbl locus that expresses, exclusively in the rod outer segment (ROS), two relatively abundant proteins: plasma membrane localized 2-subunit of the cGMP-gated cation channel and cytoplasmic glutamic acid-proline rich polypeptide (GARP-2). The rod cGMP-gated cation channel plays a critical role in vision by regulating the flow of calcium and sodium ions into the rod cell. In addition, both the 2-subunit and GARP-2 interact with peripherin-2/rds, a disk membrane protein, and thus may play essential structural roles. To determine the contribution of the proteins in ROS, we generated a homozygous Cngbl photoreceptor null (5' KO), and are analyzing the mice using light and EM level analysis, ERG and single cell recordings, and Western analysis. Homozygous 5' KO mice show a significant reduction (>30 fold) in light sensitivity and attenuated b-wave response, but are functional prior to the onset of retinal degeneration. Surprisingly, in the 59 KO not only is the 2-subunit absent but the channel 1-subunit is also greatly reduced in abundance in ROS. We hypothesize that the GARP region has an important structural role in the photoreceptor and that GARP-2, like the 2-subunit has also an important functional role. We will directly examine the structural and functional roles of the GARP region of the beta subunit and of GARP-2 by targeted deletion of GARP-2 expression and introduction of transgenes into 59 KO and 2-subunit knockout mice (39 KO). The proposed studies will help to establish the function of the Cngbl encoded GARP region and of GARP-2 in the photoreceptor, and hence provide important insights into the mechanisms whereby CNGB1 gene defects cause hereditary retinal degeneration. The studies may also yield new targets for intervention as possible treatments for certain forms of hereditary retina degeneration. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
1R01EY018143-01A1
Application #
7372754
Study Section
Special Emphasis Panel (ZRG1-CB-G (90))
Program Officer
Mariani, Andrew P
Project Start
2008-01-01
Project End
2011-12-31
Budget Start
2008-01-01
Budget End
2008-12-31
Support Year
1
Fiscal Year
2008
Total Cost
$362,500
Indirect Cost
Name
University of Alabama Birmingham
Department
Physiology
Type
Schools of Optometry/Ophthalmol
DUNS #
063690705
City
Birmingham
State
AL
Country
United States
Zip Code
35294
DeRamus, Marci L; Stacks, Delores A; Zhang, Youwen et al. (2017) GARP2 accelerates retinal degeneration in rod cGMP-gated cation channel ?-subunit knockout mice. Sci Rep 7:42545
Ding, Xi-Qin; Thapa, Arjun; Ma, Hongwei et al. (2016) The B3 Subunit of the Cone Cyclic Nucleotide-gated Channel Regulates the Light Responses of Cones and Contributes to the Channel Structural Flexibility. J Biol Chem 291:8721-34
Chakraborty, Dibyendu; Conley, Shannon M; Pittler, Steven J et al. (2016) Role of RDS and Rhodopsin in Cngb1-Related Retinal Degeneration. Invest Ophthalmol Vis Sci 57:787-97
Chakraborty, Dibyendu; Conley, Shannon M; DeRamus, Marci L et al. (2015) Varying the GARP2-to-RDS Ratio Leads to Defects in Rim Formation and Rod and Cone Function. Invest Ophthalmol Vis Sci 56:8187-98
Sarfare, Shanta; McKeown, Alex S; Messinger, Jeffrey et al. (2014) Overexpression of rod photoreceptor glutamic acid rich protein 2 (GARP2) increases gain and slows recovery in mouse retina. Cell Commun Signal 12:67
Lu, Rongwen; Levy, Alexander M; Zhang, Qiuxiang et al. (2013) Dynamic near-infrared imaging reveals transient phototropic change in retinal rod photoreceptors. J Biomed Opt 18:106013
Zhang, Youwen; Rubin, Glen R; Fineberg, Naomi et al. (2012) Age-related changes in Cngb1-X1 knockout mice: prolonged cone survival. Doc Ophthalmol 124:163-75
Lu, Rong-Wen; Curcio, Christine A; Zhang, Youwen et al. (2012) Investigation of the hyper-reflective inner/outer segment band in optical coherence tomography of living frog retina. J Biomed Opt 17:060504
Gilliam, Jared C; Chang, Juan T; Sandoval, Ivette M et al. (2012) Three-dimensional architecture of the rod sensory cilium and its disruption in retinal neurodegeneration. Cell 151:1029-41
Zhang, Qiu-Xiang; Zhang, Youwen; Lu, Rong-Wen et al. (2012) Comparative intrinsic optical signal imaging of wild-type and mutant mouse retinas. Opt Express 20:7646-54

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