Comparative study of PRPH2 D2 loop mutants reveals divergent disease mechanism in rods and cones.


Mutations in the photoreceptor-specific tetraspanin gene peripherin-2 (PRPH2) lead to widely varying forms of retinal degeneration ranging from retinitis pigmentosa to macular dystrophy. Both inter- and intra-familial phenotypic heterogeneity has led to much interest in uncovering the complex pathogenic mechanisms of PRPH2-associated disease. Majority of disease-causing mutations in PRPH2 reside in the second intradiscal loop, wherein seven cysteines control protein folding and oligomerization. Here, we utilize knockin models to evaluate the role of three D2 loop cysteine mutants (Y141C, C213Y and C150S), alone or in combination. We elucidated how these mutations affect PRPH2 properties, including oligomerization and subcellular localization, and contribute to disease processes. Results from our structural, functional and molecular studies revealed that, in contrast to our understanding from prior investigations, rods are highly affected by PRPH2 mutations interfering with oligomerization and not merely by the haploinsufficiency associated with these mutations. On the other hand, cones are less affected by the toxicity of the mutant protein and significantly reduced protein levels, suggesting that knockdown therapeutic strategies may sustain cone functionality for a longer period. This observation provides useful data to guide and simplify the current development of effective therapeutic approaches for PRPH2-associated diseases that combine knockdown with high levels of gene supplementation needed to generate prolonged rod improvement.





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Publication Info

Ikelle, Larissa, Mustafa Makia, Tylor Lewis, Ryan Crane, Mashal Kakakhel, Shannon M Conley, James R Birtley, Vadim Y Arshavsky, et al. (2023). Comparative study of PRPH2 D2 loop mutants reveals divergent disease mechanism in rods and cones. Cellular and molecular life sciences : CMLS, 80(8). p. 214. 10.1007/s00018-023-04851-3 Retrieved from

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Vadim Y Arshavsky

Helena Rubinstein Foundation Distinguished Professor of Ophthalmology

The Biology and Pathophysiology of Vertebrate Photoreceptor Cells

Research conducted in our laboratory is dedicated to understanding how vision is performed on the molecular level. Most of our work is centered on vertebrate photoreceptor cells, which are sensory neurons responsible for light detection in the eye. Photoreceptors capture photons, produce an electrical signal, and transmit this information to the secondary neurons in the retina, and ultimately to the brain, through modulation of their synaptic release. 

The main experimental direction of our laboratory is to elucidate the cellular processes responsible for building the light-sensitive organelle of photoreceptor cells, called the outer segment, and for populating this organelle with proteins supporting its structure and conducting visual signaling. Of particular interest is the mechanism by which outer segments form their “disc” membrane stacks providing vast membrane surfaces for efficient photon capture. Outer segment membranes are continuously renewed throughout the lifetime of a photoreceptor, with new discs added to the outer segment base and old discs phagocytosed at the tip by the retinal pigment epithelium. As a result, the entire mammalian outer segment is replaced with new discs over the course of 8-10 days. One of the central goals of our current studies is to elucidate the signaling pathway that acts as a “control center” to initiate the formation of each new disc with the strikingly regular frequency of approximately 80 times per day. 

Our second major research direction explores a connection between understanding the basic function of rods and cones and practical, translational ideas aiming to ameliorate retinal degeneration caused by mutations in critical photoreceptor-specific proteins. Several years ago, we found that photoreceptors bearing a broad spectrum of disease-associated mutations suffer from a common cellular stress factor, proteasomal overload, i.e. insufficient ability of the ubiquitin-proteasome system to process misfolded and/or mislocalized proteins produced in these cells. Our more recent data demonstrate that the enhancement of protein degradation machinery in these cells causes a remarkable delay in the progression of photoreceptor degeneration. We continue investigating photoreceptor proteostasis in further mechanistic depth and seek optimal strategies to employ proteasomal activation as a means to ameliorate or cure inherited blindness. 

During our studies, we explore high-end applications of mass spectrometry-based proteomics and were the first laboratory adopting several advanced proteomic approaches to vision research. Of particular significance are the applications of so-called “label-free” quantitative proteomics for simultaneous elucidation of multiple protein distributions among different compartments of the photoreceptor cells and for identification of unique protein components of various photoreceptor membranes. Using label-free proteomics, we demonstrated that a small protein PRCD (progressive rod and cone degeneration) is a unique component of photoreceptor discs and subsequently identified several novel unique components of the plasma membrane enclosing the rod outer segment. Most recently, we adopted a highly efficient and accurate methodology for simultaneous absolute quantification of several dozen proteins, termed MS Western. This method allowed us to determine the precise molar ratio amongst all major functional and structural proteins residing in the light-sensitive outer segments of photoreceptor cells.

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