July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Pharmacological interference in the VCP/ERAD/proteasome axis rescues photoreceptor degeneration in RhP23H
Author Affiliations & Notes
  • Merve Sen
    Centre for Ophthalmology, University Eye Hospital Tübingen, Tübingen, Germany
    Graduate Training Centre of Neuroscience, Eberhard Karls University of Tübingen, Tübingen, Germany
  • Oksana Kutsyr
    Department of Physiology, Genetics and Microbiology, University of Alicante, Alicante, Spain
  • Sylvia Bolz
    Centre for Ophthalmology, University Eye Hospital Tübingen, Tübingen, Germany
  • Tsui-Fen Chou
    Department of Pediatrics, Harbor-UCLA Medical Center and Los Angeles Biomedical Research Institute, California, United States
  • Ray Deshaies
    Research, Amgen Inc., California, United States
  • Blanca Arango-Gonzalez
    Centre for Ophthalmology, University Eye Hospital Tübingen, Tübingen, Germany
  • Marius Ueffing
    Centre for Ophthalmology, University Eye Hospital Tübingen, Tübingen, Germany
  • Footnotes
    Commercial Relationships   Merve Sen, None; Oksana Kutsyr, None; Sylvia Bolz, None; Tsui-Fen Chou, None; Ray Deshaies, Amgen (E), Cleave Biosciences (I); Blanca Arango-Gonzalez, None; Marius Ueffing, None
  • Footnotes
    Support  H2020-MSCA-ITN-2016 OcuTher-722717; FFB Grant PPA-0717-0719-RAD; Pro-Re/Projekt/Ueffing.1-2015; Pro-Re/Projekt/Ueffing.1-2015/17
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 4881. doi:https://doi.org/
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    • Get Citation

      Merve Sen, Oksana Kutsyr, Sylvia Bolz, Tsui-Fen Chou, Ray Deshaies, Blanca Arango-Gonzalez, Marius Ueffing; Pharmacological interference in the VCP/ERAD/proteasome axis rescues photoreceptor degeneration in RhP23H. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4881. doi: https://doi.org/.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : One of the most prevalent mutations in rhodopsin (Rh) causing autosomal dominant Retinitis Pigmentosa (adRP) RhP23H, results in its misfolding and its retention in the Endoplasmic Reticulum (ER). Valosin-containing protein (VCP) is an ATP driven molecular checkpoint at the ER before these proteins are released from the ER towards plasma membrane. Misfolded proteins are selectively identified, retained at the ER and cleared by a process called ER-associated degradation (ERAD). ERAD overload due to RhP23H misfolding causes cell death. As several pathways regulating protein homeostasis are associated with ERAD, we tested their specific impact on photoreceptor (PR) degeneration through pharmacological inhibition.

Methods : Organotypic cultures from RhP23H retinas of transgenic rats at PN9 cultured for 6 days were treated with different concentrations of pathway specific inhibitors: NMS-873 interferes with quality control mechanisms by VCP inhibition; the proteasome inhibitor Bortezomib (BO) suppresses proteasomal degradation; the mannosidase inhibitor Kifunensine (KIF) blocks processing of glycoproteins, and Geldanamycin (GA) inhibits the protein-folding mechanism in the ER. PR cell survival was assessed by counting cell rows and measuring the percentage of TUNEL-positive cells. Rh distribution was checked using a specific antibody.

Results : VCP inhibitor NMS-873 caused strong and dose-dependent protection at 5µM (TUNEL; treated: 2,4%±0,2, control: 6,76%±0,6, P<0,002). Proteasome inhibition by BO significantly reduced the number of dying cells at 10µM (TUNEL; treated: 3,13%±0,3, control: 5,42%±0,6, P<0,05). The number of PR cell rows in the ONL was increased by both, NMS-873 (5µM; treated: 9,6±0,3, control: 6,06±0,6, P<0,05), and BO (10µM; treated: 9,37±0,6, control: 7,6±0,1, P<0,002). Only VCP inhibitor corrected aberrant inner segment distribution of Rh shifting it to physiological outer segment (OS). In contrary, KIF and GA turned out to accelerate PR cell degeneration.

Conclusions : Modulation of the VCP/ERAD/proteasome axis by inhibiting VCP and proteasome rescues PR cells in RhP23H. Inhibition of VCP activity establishes proper transport of Rh to the OS. These results strongly suggest that the VCP/ERAD/proteasome axis acts as a critical checkpoint in PR homeostasis whereas other pathways in ERAD that control protein integrity or proper folding of proteins do not.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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