Retinal degenerative diseases are often characterized by abnormalities in photoreceptor structure. Central to our understanding of photoreceptor health and disease is a fundamental knowledge regarding how photoreceptor outer segments are formed and what processes contribute to their structural stability. Peripherin/rds (p/rds), encoded by the rds gene, is essential for the formation, organization and maintenance of outer segments. In humans, any one of over 150 mutations within this gene results in a broad variety of late onset progressive retinal dystrophies characterized by abnormal photoreceptor structure. The pathogenic mechanism(s) underlying p/rds mediated retinal dystrophies are unknown, however new insight has been gained in our understanding of p/rds function based upon our recent structural studies identifying the C- terminal domain of this protein as homologous to structures called intrinsically disordered domains (IDD). An unusually large number of IDD containing proteins are associated with human diseases, including cancer, cardiovascular disease, amyloidosis, neurodegenerative diseases and diabetes. IDDs are random structures which are stabilized through interactions with one or with several binding partners. Structural flexibility imparts multi-functionality to the protein and binding partners act as regulators of function. We anticipate that deciphering the function of the IDD C-terminal domain of p/rds is critical to understanding the mechanism of OS formation in photoreceptors and how mutations in this region contribute to rod or cone dominant dystrophies. Hence the primary goals of this project are to test the hypothesis that the IDD p/rds C-terminal domain plays a direct role in the biogenesis of rod OS and in stabilizing OS structure. We will define the role(s) of the C-terminal domain of p/rds in OS morphogenesis and stability and understand the physiological implications of binary (direct) protein binding to this domain. A comprehensive strategy involving in vivo mouse models and in vitro biophysical approaches is proposed.
In specific Aim #1 a using in vivo electroporation to introduce genetically modified rds into the developing mouse retina we will determine how truncation and functional mutations in the p/rds C-terminus modulate p/rds trafficking as well as the establishment and maintenance of OS structure. A primary objective of this aim is to map the p/rds localization signal in mouse. Furthermore, we will visualize the interaction between p/rds and its known protein binding partners in vivo and assess how inhibition of these interactions affects, OS formation using bi- molecular fluorescence complementation assays. In addition, we will assess how disease linked p/rds C- terminal mutations alter p/rds targeting, assembly and maintenance of structure and correlate how folding of the intrinsically disordered C-terminal domain in these mutants may alter binding partner affinity. The proposed studies are expected to provide significant mechanistic insights into peripherin/rds function and a better understanding of both normal biology of the retina and retinal degenerations associated with mutations in p/rds. An understanding of the function of p/rds in the establishment and maintenance of photoreceptor outer segments should provide clues to an overall understanding of tetraspanin protein function and the role of intrinsically disordered domains in cellular processes and disease pathogenesis. These studies will provide the underpinnings for the design of therapeutic strategies to treat late-onset retinal degenerations, a growing problem in an aging population.

Public Health Relevance

Mutations in photoreceptor peripherin/rds are one of the most common causes of blindness and impaired vision in the elderly. Specifically, they are the most prevalent cause of autosomal dominant retinitis pigmentosa (adRP) in the United States and Northern Europe;over 150 mutations within this gene result in a broad variety of late onset progressive retinal dystrophies characterized by abnormal photoreceptor structure. We will determine how peripherin/rds mutations contribute to photoreceptor dysfunction by examining the role of structurally unique regions of this protein in photoreceptor formation using in vitro methods and mouse models of human disease. Our studies should reveal novel insights into the pathogenesis of peripherin/rds mediated retinal dystrophies and form the underpinnings for the development of therapeutic strategies to treat these diseases.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY010420-16
Application #
8247084
Study Section
Biology and Diseases of the Posterior Eye Study Section (BDPE)
Program Officer
Neuhold, Lisa
Project Start
1994-08-01
Project End
2014-03-31
Budget Start
2012-04-01
Budget End
2014-03-31
Support Year
16
Fiscal Year
2012
Total Cost
$374,220
Indirect Cost
$136,620
Name
University of Pennsylvania
Department
Biochemistry
Type
Schools of Dentistry
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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