1. Cell Biology

Endoplasmic reticulum tubules limit the size of misfolded protein condensates

  1. Smriti Parashar
  2. Ravi Chidambaram
  3. Shuliang Chen
  4. Christina R Liem
  5. Eric Griffis
  6. Gerard G Lambert
  7. Nathan C Shaner
  8. Matthew Wortham
  9. Jesse C Hay
  10. Susan Ferro-Novick  Is a corresponding author
  1. University of California, San Diego, United States
  2. University of Montana, United States
https://doi.org/10.7554/eLife.71642
Share this article
Cite this article
  1. Smriti Parashar
  2. Ravi Chidambaram
  3. Shuliang Chen
  4. Christina R Liem
  5. Eric Griffis
  6. Gerard G Lambert
  7. Nathan C Shaner
  8. Matthew Wortham
  9. Jesse C Hay
  10. Susan Ferro-Novick
(2021)
Endoplasmic reticulum tubules limit the size of misfolded protein condensates
eLife 10:e71642.
https://doi.org/10.7554/eLife.71642
  2. Download BibTeX
  3. Download .RIS

Abstract

The endoplasmic reticulum (ER) is composed of sheets and tubules. Here we report that the COPII coat subunit, SEC24C, works with the long form of the tubular ER-phagy receptor, RTN3, to target dominant-interfering mutant proinsulin Akita puncta to lysosomes. When the delivery of Akita puncta to lysosomes was disrupted, large puncta accumulated in the ER. Unexpectedly, photobleach analysis indicated that Akita puncta behaved as condensates and not aggregates, as previously suggested. Akita puncta enlarged when either RTN3 or SEC24C were depleted, or when ER sheets were proliferated by either knocking out Lunapark or overexpressing CLIMP63. Other ER-phagy substrates that are segregated into tubules behaved like Akita, while a substrate (type I procollagen) that is degraded by the ER-phagy sheets receptor, FAM134B, did not. Conversely, when ER tubules were augmented in Lunapark knock-out cells by overexpressing reticulons, ER-phagy increased and the number of large Akita puncta were reduced. Our findings imply that segregating cargos into tubules has two beneficial roles. First, it localizes mutant misfolded proteins, the receptor and SEC24C to the same ER domain. Second, physically restraining condensates within tubules, before they undergo ER-phagy, prevents them from enlarging and impacting cell health.

Data availability

All data generated or analyzed during this study are included in the manuscript and supporting files.

Article and author information

Author details

  1. Smriti Parashar

    Department of Cellular and Molecular Medicine, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.

  2. Ravi Chidambaram

    Department of Cellular and Molecular Medicine, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.

  3. Shuliang Chen

    Department of Cellular and Molecular Medicine, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.

  4. Christina R Liem

    Division of Biological Sciences, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.

  5. Eric Griffis

    Nikon Imaging Center, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.

  6. Gerard G Lambert

    Department of Neurosciences, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.

  7. Nathan C Shaner

    Department of Neurosciences, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.

  8. Matthew Wortham

    Department of Neurosciences, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.

  9. Jesse C Hay

    Division of Biological Sciences and Center for Structural & Functional Neuroscience, University of Montana, Missoula, United States
    Competing interests
    The authors declare that no competing interests exist.

  10. Susan Ferro-Novick

    Department of Cellular and Molecular Medicine, University of California, San Diego, San Diego, United States
    For correspondence
    sfnovick@ucsd.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8714-7352

Funding

National Institute of General Medical Sciences (5R35GM131681)

  • Susan Ferro-Novick

National Science Foundation (1707352)

  • Nathan C Shaner

The Pathways in Biological Science Graduate Training Program

  • Christina R Liem

National Institute of Neurological Disorders and Stroke (RO1NS117440)

  • Susan Ferro-Novick

National Institute of Diabetes and Digestive and Kidney Diseases (R01DK068471)

  • Matthew Wortham

National Institute of General Medical Sciences (2R15GM106323)

  • Jesse C Hay

National Institute of General Medical Sciences (R01GM109984)

  • Nathan C Shaner

National Institute of General Medical Sciences (R01GM121944)

  • Nathan C Shaner

National Institute of Neurological Disorders and Stroke (U01NS099709)

  • Nathan C Shaner

National Eye Institute (R21EY030716)

  • Nathan C Shaner

National Institute of Neurological Disorders and Stroke (U01NS113294)

  • Nathan C Shaner

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

Reviewing Editor

  1. Randy Schekman, Howard Hughes Medical Institute, University of California, Berkeley, United States

Publication history

  1. Received: June 25, 2021
  2. Accepted: August 31, 2021
  3. Accepted Manuscript published: September 1, 2021 (version 1)
  4. Version of Record published: October 1, 2021 (version 2)

Copyright

© 2021, Parashar et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 4,072
    Page views
  • 631
    Downloads
  • 13
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

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. Smriti Parashar
  2. Ravi Chidambaram
  3. Shuliang Chen
  4. Christina R Liem
  5. Eric Griffis
  6. Gerard G Lambert
  7. Nathan C Shaner
  8. Matthew Wortham
  9. Jesse C Hay
  10. Susan Ferro-Novick
(2021)
Endoplasmic reticulum tubules limit the size of misfolded protein condensates
eLife 10:e71642.
https://doi.org/10.7554/eLife.71642

Categories and tags

Further reading

    1. Cell Biology

    Insights into cargo sorting by SNX32 and its role in neurite outgrowth

    Jini Sugatha, Amulya Priya ... Sunando Datta
    Research Article Updated

    Sorting nexins (SNX) are a family of proteins containing the Phox homology domain, which shows a preferential endo-membrane association and regulates cargo sorting processes. Here, we established that SNX32, an SNX-BAR (Bin/Amphiphysin/Rvs) sub-family member associates with SNX4 via its BAR domain and the residues A226, Q259, E256, R366 of SNX32, and Y258, S448 of SNX4 that lie at the interface of these two SNX proteins mediate this association. SNX32, via its PX domain, interacts with the transferrin receptor (TfR) and Cation-Independent Mannose-6-Phosphate Receptor (CIMPR), and the conserved F131 in its PX domain is important in stabilizing these interactions. Silencing of SNX32 leads to a defect in intracellular trafficking of TfR and CIMPR. Further, using SILAC-based differential proteomics of the wild-type and the mutant SNX32, impaired in cargo binding, we identified Basigin (BSG), an immunoglobulin superfamily member, as a potential interactor of SNX32 in SHSY5Y cells. We then demonstrated that SNX32 binds to BSG through its PX domain and facilitates its trafficking to the cell surface. In neuroglial cell lines, silencing of SNX32 leads to defects in neuronal differentiation. Moreover, abrogation in lactate transport in the SNX32-depleted cells led us to propose that SNX32 may contribute to maintaining the neuroglial coordination via its role in BSG trafficking and the associated monocarboxylate transporter activity. Taken together, our study showed that SNX32 mediates the trafficking of specific cargo molecules along distinct pathways.

    1. Cell Biology
    2. Stem Cells and Regenerative Medicine

    Atf3 defines a population of pulmonary endothelial cells essential for lung regeneration

    Terren K Niethamer, Lillian I Levin ... Edward E Morrisey
    Research Article

    Following acute injury, the capillary vascular bed in the lung must be repaired to reestablish gas exchange with the external environment. Little is known about the transcriptional and signaling factors that drive pulmonary endothelial cell (EC) proliferation and subsequent regeneration of pulmonary capillaries, as well as their response to stress. Here, we show that the transcription factor Atf3 is essential for the regenerative response of the mouse pulmonary endothelium after influenza infection. Atf3 expression defines a subpopulation of capillary ECs enriched in genes involved in endothelial development, differentiation, and migration. During lung alveolar regeneration, this EC population expands and increases the expression of genes involved in angiogenesis, blood vessel development, and cellular response to stress. Importantly, endothelial cell-specific loss of Atf3 results in defective alveolar regeneration, in part through increased apoptosis and decreased proliferation in the endothelium. This leads to the general loss of alveolar endothelium and persistent morphological changes to the alveolar niche, including an emphysema-like phenotype with enlarged alveolar airspaces lined with regions that lack vascular investment. Taken together, these data implicate Atf3 as an essential component of the vascular response to acute lung injury that is required for successful lung alveolar regeneration.

    1. Cell Biology
    2. Microbiology and Infectious Disease

    Profiling the bloodstream form and procyclic form Trypanosoma brucei cell cycle using single-cell transcriptomics

    Emma M Briggs, Catarina A Marques ... Keith R Matthews
    Research Article Updated

    African trypanosomes proliferate as bloodstream forms (BSFs) and procyclic forms in the mammal and tsetse fly midgut, respectively. This allows them to colonise the host environment upon infection and ensure life cycle progression. Yet, understanding of the mechanisms that regulate and drive the cell replication cycle of these forms is limited. Using single-cell transcriptomics on unsynchronised cell populations, we have obtained high resolution cell cycle regulated (CCR) transcriptomes of both procyclic and slender BSF Trypanosoma brucei without prior cell sorting or synchronisation. Additionally, we describe an efficient freeze–thawing protocol that allows single-cell transcriptomic analysis of cryopreserved T. brucei. Computational reconstruction of the cell cycle using periodic pseudotime inference allowed the dynamic expression patterns of cycling genes to be profiled for both life cycle forms. Comparative analyses identify a core cycling transcriptome highly conserved between forms, as well as several genes where transcript levels dynamics are form specific. Comparing transcript expression patterns with protein abundance revealed that the majority of genes with periodic cycling transcript and protein levels exhibit a relative delay between peak transcript and protein expression. This work reveals novel detail of the CCR transcriptomes of both forms, which are available for further interrogation via an interactive webtool.