): Defects in the human RDS gene result in nearly a dozen clinically distinct retinal diseases - all of which impair central and/or peripheral vision progressively. The RDS gene product, peripherin/rds, is an integral membrane protein required for the formation of the photoreceptor outer segment (OS); however, neither its function at the molecular level, nor its role in retinal pathophysiology is currently understood. We hypothesize that inherited defects in peripherin/rds can affect particular protein domains to preferentially inhibit photoreceptor OS disk morphogenesis, stacking, or shedding - and thereby generate heterogeneous disease phenotypes.
Specific aim 1 will test the hypothesis that polytopic protein domains in peripherin/rds contribute differentially to its biosynthesis, subcellular localization, and ability to promote in vitro membrane fusion (fusogenicity), by using an insertional mutagenesis approach combined with cell-free and heterologous expression systems. These studies will determine if, and how, mutations in individual regions of peripherin/rds selectively affect particular aspects of protein structure and/or in vitro function.
Specific aim 2 will test the hypothesis that peripherin/rds fusogenicity is required in vivo for OS disk shedding. A heterologous expression system will be used to design a partial loss-of-function mutant that is defective only in its ability to catalyze membrane fusion. Light and electron microscopic analyses will determine whether disk shedding is inhibited in mouse rod photoreceptors expressing this transgene for """"""""non-fusogenic"""""""" peripherin/rds.
Specific aim 3 will test the hypothesis that peripherin/rds plays a role in maintaining the integrity of outer segment disk stacks. A heterologous expression system will be used to design an otherwise wild-type peripherin/rds variant that contains a highly-specific Factor Xa protease cleavage site; this variant will be expressed as a transgene to rescue photoreceptors in the retinal degeneration slow (rds) mouse (rds does not express peripherin/rds). In vitro Factor Xa proteolysis combined with electron microscopic and Western blot analyses will be used to determine whether specific cleavage of this peripherin/rds variant disrupts the disk stack organization of transgenic mouse photoreceptor OSs. In sum, this research program is directed towards establishing the normal function(s) of peripherin/rds in OS renewal, to begin a molecular description of the etiologies behind a wide variety of progressive blinding diseases that result from defects in RDS.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY013246-02
Application #
6498366
Study Section
Visual Sciences C Study Section (VISC)
Program Officer
Mariani, Andrew P
Project Start
2001-02-05
Project End
2006-01-31
Budget Start
2002-02-01
Budget End
2003-01-31
Support Year
2
Fiscal Year
2002
Total Cost
$248,500
Indirect Cost
Name
Oakland University
Department
Type
Organized Research Units
DUNS #
City
Rochester
State
MI
Country
United States
Zip Code
48309
Milstein, Michelle L; Kimler, Victoria A; Ghatak, Chiranjib et al. (2017) An inducible amphipathic helix within the intrinsically disordered C terminus can participate in membrane curvature generation by peripherin-2/rds. J Biol Chem 292:7850-7865
Wahl, Silke; Magupalli, Venkat Giri; Dembla, Mayur et al. (2016) The Disease Protein Tulp1 Is Essential for Periactive Zone Endocytosis in Photoreceptor Ribbon Synapses. J Neurosci 36:2473-93
Wright, Zachary C; Singh, Ratnesh K; Alpino, Ryan et al. (2016) ARL3 regulates trafficking of prenylated phototransduction proteins to the rod outer segment. Hum Mol Genet 25:2031-2044
Goldberg, Andrew F X; Moritz, Orson L; Williams, David S (2016) Molecular basis for photoreceptor outer segment architecture. Prog Retin Eye Res 55:52-81
Khattree, Nidhi; Ritter, Linda M; Goldberg, Andrew F X (2013) Membrane curvature generation by a C-terminal amphipathic helix in peripherin-2/rds, a tetraspanin required for photoreceptor sensory cilium morphogenesis. J Cell Sci 126:4659-70
Sharma, Yagya V; Cojocaru, Radu I; Ritter, Linda M et al. (2012) Protective gene expression changes elicited by an inherited defect in photoreceptor structure. PLoS One 7:e31371
Ritter, Linda M; Khattree, Nidhi; Tam, Beatrice et al. (2011) In situ visualization of protein interactions in sensory neurons: glutamic acid-rich proteins (GARPs) play differential roles for photoreceptor outer segment scaffolding. J Neurosci 31:11231-43
Kirschman, Lindsay T; Kolandaivelu, Saravanan; Frederick, Jeanne M et al. (2010) The Leber congenital amaurosis protein, AIPL1, is needed for the viability and functioning of cone photoreceptor cells. Hum Mol Genet 19:1076-87
Goldberg, Andrew F X; Ritter, Linda M; Khattree, Nidhi et al. (2007) An intramembrane glutamic acid governs peripherin/rds function for photoreceptor disk morphogenesis. Invest Ophthalmol Vis Sci 48:2975-86
Goldberg, Andrew F X (2006) Role of peripherin/rds in vertebrate photoreceptor architecture and inherited retinal degenerations. Int Rev Cytol 253:131-75

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