Retinal photoreceptor cells maintain specific composition of the photosensitive membranes by continuous membrane renewal. In patients with retinal genetic disorders this process is often compromised by mutations in rhodopsin or factors that regulate its intracellular trafficking. For example the C-terminal domain of rhodopsin is a frequent target for mutations that cause severe forms of autosomal dominant retinitis pigmentosa (ADRP) while mutations in regulators of small GTPases of the ras and rab super family cause choroideremia and X- linked RP3. The long-term goal is to delineate the molecular mechanisms that regulate polarized delivery of rhodopsin and its associated proteins to the rod outer segment. Based on our preliminary studies, we propose to investigate interactions between rhodopsin, small GTPases and their regulatory proteins and phospholipids that in normal cells bring about the correct membrane sorting and delivery to the ROS. We will apply biochemical, morphological and cell biological tools to analyze photoreceptor biosynthetic membranes under normal conditions and under normal conditions that correspond to those found in retinal diseases.
The specific aims of this proposal are: 1) To determine the role of rhodopsin in regulation of its post-Golgi trafficking. 2) To elucidate the role of small GTPases, their regulators such as Retinitis Pigmentosa GTPase Regulator (Rpgr), and post-Golgi membrane specific proteins in rhodopsin trafficking. 3) To elucidate the role of lipids in the polarized membrane delivery to the ROS. 4) To determine the mechanism of global regulation of membrane delivery to the ROS by extracellular signals and secondary messengers. Rhodopsin trafficking will be monitored in vivo and in an established cell-free system that reconstitutes sorting of rhodopsin into specific post-Golgi carrier membranes. Biochemical analyses of retinal subcellular factions will be complemented by confocal and electron microscopy. To model conditions found in retinal diseases membrane renewal will be perturbed by adding antibodies, inhibitors, peptides and mutant proteins, or by depleting proteins or lipids that participate in intracellular sorting and trafficking. Our research goals are to provide better understanding of the molecular mechanisms that govern cell polarity and the health of retinal photoreceptors. With this knowledge we will be able to design and develop new approaches to the treatments of retinal diseases.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
1R01EY012421-01
Application #
2752324
Study Section
Visual Sciences C Study Section (VISC)
Project Start
1999-01-01
Project End
2002-12-31
Budget Start
1999-01-01
Budget End
1999-12-31
Support Year
1
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Ophthalmology
Type
Schools of Medicine
DUNS #
791277940
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Wang, Jing; Fresquez, Theresa; Kandachar, Vasundhara et al. (2017) The Arf GEF GBF1 and Arf4 synergize with the sensory receptor cargo, rhodopsin, to regulate ciliary membrane trafficking. J Cell Sci 130:3975-3987
Wang, Jing; Deretic, Dusanka (2015) The Arf and Rab11 effector FIP3 acts synergistically with ASAP1 to direct Rabin8 in ciliary receptor targeting. J Cell Sci 128:1375-85
Vetter, Melanie; Wang, Jing; Lorentzen, Esben et al. (2015) Novel topography of the Rab11-effector interaction network within a ciliary membrane targeting complex. Small GTPases 6:165-73
Wang, Jing; Deretic, Dusanka (2014) Molecular complexes that direct rhodopsin transport to primary cilia. Prog Retin Eye Res 38:1-19
Deretic, Dusanka (2013) Crosstalk of Arf and Rab GTPases en route to cilia. Small GTPases 4:70-7
Wang, Jing; Morita, Yoshiko; Mazelova, Jana et al. (2012) The Arf GAP ASAP1 provides a platform to regulate Arf4- and Rab11-Rab8-mediated ciliary receptor targeting. EMBO J 31:4057-71
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
Deretic, Dusanka; Wang, Jing (2012) Molecular assemblies that control rhodopsin transport to the cilia. Vision Res 75:5-10
Ward, Heather H; Brown-Glaberman, Ursa; Wang, Jing et al. (2011) A conserved signal and GTPase complex are required for the ciliary transport of polycystin-1. Mol Biol Cell 22:3289-305
Mazelova, Jana; Ransom, Nancy; Astuto-Gribble, Lisa et al. (2009) Syntaxin 3 and SNAP-25 pairing, regulated by omega-3 docosahexaenoic acid, controls the delivery of rhodopsin for the biogenesis of cilia-derived sensory organelles, the rod outer segments. J Cell Sci 122:2003-13

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