Reserpine maintains photoreceptor survival in retinal ciliopathy by resolving proteostasis imbalance and ciliogenesis defects

  1. Holly Y Chen
  2. Manju Swaroop
  3. Samantha Papal
  4. Anupam Mondal
  5. Hyun Beom Song
  6. Laura Campello
  7. Gregory Tawa
  8. Florian Regent
  9. Hiroko Shimada
  10. Kunio Nagashima
  11. Natalia de Val
  12. Samuel G Jacobson
  13. Wei Zheng
  14. Anand Swaroop  Is a corresponding author
  1. National Eye Institute, United States
  2. National Center for Advancing Translational Sciences, United States
  3. Frederick National Laboratory for Cancer Research, United States
  4. University of Pennsylvania, United States

Abstract

Ciliopathies manifest from sensory abnormalities to syndromic disorders with multi-organ pathologies, with retinal degeneration a highly penetrant phenotype. Photoreceptor cell death is a major cause of incurable blindness in retinal ciliopathies. To identify drug candidates to maintain photoreceptor survival, we performed an unbiased, high-throughput screening of over 6,000 bioactive small molecules using retinal organoids differentiated from induced pluripotent stem cells (iPSC) of rd16 mouse, which is a model of Leber congenital amaurosis (LCA) type 10 caused by mutations in the cilia-centrosomal gene CEP290. We identified five non-toxic positive hits, including the lead molecule reserpine, which maintained photoreceptor development and survival in rd16 organoids. Reserpine also improved photoreceptors in retinal organoids derived from induced pluripotent stem cells of LCA10 patients and in rd16 mouse retina in vivo. Reserpine-treated patient organoids revealed modulation of signaling pathways related to cell survival/death, metabolism, and proteostasis. Further investigation uncovered dysregulation of autophagy associated with compromised primary cilium biogenesis in patient organoids and rd16 mouse retina. Reserpine partially restored the balance between autophagy and the ubiquitin-proteasome system at least in part by increasing the cargo adaptor p62, resulting in improved primary cilium assembly. Our study identifies effective drug candidates in preclinical studies of CEP290 retinal ciliopathies through cross-species drug discovery using iPSC-derived organoids, highlights the impact of proteostasis in the pathogenesis of ciliopathies, and provides new insights for treatments of retinal neurodegeneration.

Data availability

All data needed to evaluate the conclusions in the paper are present in the paper and/or the Supplementary Materials. RNA-seq data are available through GEO accession #206959.

The following data sets were generated

Article and author information

Author details

  1. Holly Y Chen

    Neurobiology, Neurodegeneration and Repair Laboratory, National Eye Institute, Bethesda, United States
    Competing interests
    Holly Y Chen, Listed as inventor on a patent application related to the small molecules in this study by National Institutes of Health (PCT/US2021/040157).
  2. Manju Swaroop

    National Therapeutics for Rare and Neglected Diseases, National Center for Advancing Translational Sciences, Rockville, United States
    Competing interests
    Manju Swaroop, Listed as inventor on a patent application related to the small molecules in this study by National Institutes of Health (PCT/US2021/040157).
  3. Samantha Papal

    Neurobiology, Neurodegeneration and Repair Laboratory, National Eye Institute, Bethesda, United States
    Competing interests
    Samantha Papal, Listed as inventor on a patent application related to the small molecules in this study by National Institutes of Health (PCT/US2021/040157).
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9417-6215
  4. Anupam Mondal

    Neurobiology, Neurodegeneration and Repair Laboratory, National Eye Institute, Bethesda, United States
    Competing interests
    Anupam Mondal, Listed as inventor on a patent application related to the small molecules in this study by National Institutes of Health (PCT/US2021/040157).
  5. Hyun Beom Song

    Neurobiology, Neurodegeneration and Repair Laboratory, National Eye Institute, Bethesda, United States
    Competing interests
    No competing interests declared.
  6. Laura Campello

    Neurobiology, Neurodegeneration and Repair Laboratory, National Eye Institute, Bethesda, United States
    Competing interests
    No competing interests declared.
  7. Gregory Tawa

    National Therapeutics for Rare and Neglected Diseases, National Center for Advancing Translational Sciences, Rockville, United States
    Competing interests
    Gregory Tawa, Listed as inventor on a patent application related to the small molecules in this study by National Institutes of Health (PCT/US2021/040157).
  8. Florian Regent

    Neurobiology, Neurodegeneration and Repair Laboratory, National Eye Institute, Bethesda, United States
    Competing interests
    No competing interests declared.
  9. Hiroko Shimada

    Neurobiology, Neurodegeneration and Repair Laboratory, National Eye Institute, Bethesda, United States
    Competing interests
    No competing interests declared.
  10. Kunio Nagashima

    Electron Microscopy Laboratory, Frederick National Laboratory for Cancer Research, Frederick, United States
    Competing interests
    No competing interests declared.
  11. Natalia de Val

    Electron Microscopy Laboratory, Frederick National Laboratory for Cancer Research, Frederick, United States
    Competing interests
    No competing interests declared.
  12. Samuel G Jacobson

    Department of Ophthalmology, University of Pennsylvania, Philadelphia, United States
    Competing interests
    No competing interests declared.
  13. Wei Zheng

    National Therapeutics for Rare and Neglected Diseases, National Center for Advancing Translational Sciences, Rockville, United States
    Competing interests
    Wei Zheng, Listed as inventor on a patent application related to the small molecules in this study by National Institutes of Health (PCT/US2021/040157).
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1034-0757
  14. Anand Swaroop

    Neurobiology, Neurodegeneration and Repair Laboratory, National Eye Institute, Bethesda, United States
    For correspondence
    swaroopa@nei.nih.gov
    Competing interests
    Anand Swaroop, Listed as inventor on a patent application related to the small molecules in this study by National Institutes of Health (PCT/US2021/040157).
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1975-1141

Funding

National Eye Institute (Z01EY000546)

  • Anand Swaroop

National Eye Institute (Z01EY000450)

  • Anand Swaroop

National Center for Advancing Translational Sciences (ZIATR000018-06)

  • Wei Zheng

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Zhongjie Fu, Boston Children's Hospital, United States

Ethics

Animal experimentation: All animal procedures were approved by the Animal Care and Use committee of the National Eye Institutes (Animal study protocol NEI-650) and adhered to ARVO Statement for the Use of Animals in Ophthalmic and Vision Research.

Version history

  1. Received: September 2, 2022
  2. Preprint posted: September 18, 2022 (view preprint)
  3. Accepted: March 23, 2023
  4. Accepted Manuscript published: March 28, 2023 (version 1)
  5. Version of Record published: April 21, 2023 (version 2)

Copyright

This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.

Metrics

  • 1,798
    views
  • 397
    downloads
  • 15
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Holly Y Chen
  2. Manju Swaroop
  3. Samantha Papal
  4. Anupam Mondal
  5. Hyun Beom Song
  6. Laura Campello
  7. Gregory Tawa
  8. Florian Regent
  9. Hiroko Shimada
  10. Kunio Nagashima
  11. Natalia de Val
  12. Samuel G Jacobson
  13. Wei Zheng
  14. Anand Swaroop
(2023)
Reserpine maintains photoreceptor survival in retinal ciliopathy by resolving proteostasis imbalance and ciliogenesis defects
eLife 12:e83205.
https://doi.org/10.7554/eLife.83205

Share this article

https://doi.org/10.7554/eLife.83205

Further reading

    1. Biochemistry and Chemical Biology
    Boglarka Zambo, Evelina Edelweiss ... Gergo Gogl
    Research Article

    Truncation of the protein-protein interaction SH3 domain of the membrane remodeling Bridging Integrator 1 (BIN1, Amphiphysin 2) protein leads to centronuclear myopathy. Here, we assessed the impact of a set of naturally observed, previously uncharacterized BIN1 SH3 domain variants using conventional in vitro and cell-based assays monitoring the BIN1 interaction with dynamin 2 (DNM2) and identified potentially harmful ones that can be also tentatively connected to neuromuscular disorders. However, SH3 domains are typically promiscuous and it is expected that other, so far unknown partners of BIN1 exist besides DNM2, that also participate in the development of centronuclear myopathy. In order to shed light on these other relevant interaction partners and to get a holistic picture of the pathomechanism behind BIN1 SH3 domain variants, we used affinity interactomics. We identified hundreds of new BIN1 interaction partners proteome-wide, among which many appear to participate in cell division, suggesting a critical role of BIN1 in the regulation of mitosis. Finally, we show that the identified BIN1 mutations indeed cause proteome-wide affinity perturbation, signifying the importance of employing unbiased affinity interactomic approaches.

    1. Biochemistry and Chemical Biology
    2. Chromosomes and Gene Expression
    Ramona Weber, Chung-Te Chang
    Research Article

    Recent findings indicate that the translation elongation rate influences mRNA stability. One of the factors that has been implicated in this link between mRNA decay and translation speed is the yeast DEAD-box helicase Dhh1p. Here, we demonstrated that the human ortholog of Dhh1p, DDX6, triggers the deadenylation-dependent decay of inefficiently translated mRNAs in human cells. DDX6 interacts with the ribosome through the Phe-Asp-Phe (FDF) motif in its RecA2 domain. Furthermore, RecA2-mediated interactions and ATPase activity are both required for DDX6 to destabilize inefficiently translated mRNAs. Using ribosome profiling and RNA sequencing, we identified two classes of endogenous mRNAs that are regulated in a DDX6-dependent manner. The identified targets are either translationally regulated or regulated at the steady-state-level and either exhibit signatures of poor overall translation or of locally reduced ribosome translocation rates. Transferring the identified sequence stretches into a reporter mRNA caused translation- and DDX6-dependent degradation of the reporter mRNA. In summary, these results identify DDX6 as a crucial regulator of mRNA translation and decay triggered by slow ribosome movement and provide insights into the mechanism by which DDX6 destabilizes inefficiently translated mRNAs.