TANGO1 membrane helices create a lipid diffusion barrier at curved membranes

  1. Ishier Raote  Is a corresponding author
  2. Andreas M Ernst
  3. Felix Campelo
  4. James E Rothman
  5. Frederic Pincet  Is a corresponding author
  6. Vivek Malhotra  Is a corresponding author
  1. The Barcelona Institute of Science and Technology, Spain
  2. Yale School of Medicine, United States
  3. ICFO - The Institute of Photonic Sciences, Spain
  4. Ecole Normal Superieure, France

Abstract

We have previously shown TANGO1 organises membranes at the interface of the endoplasmic reticulum (ER) and ERGIC/Golgi (Raote et al., 2018). TANGO1 corrals retrograde membranes at ER exit sites to create an export conduit. Here the retrograde membrane is, in itself, an anterograde carrier. This mode of forward transport necessitates a mechanism to prevent membrane mixing between ER and the retrograde membrane. TANGO1 has an unusual membrane helix organisation, composed of one membrane-spanning helix (TM) and another that penetrates the inner leaflet (IM). We have reconstituted these membrane helices in model membranes and shown that TM and IM together reduce the flow of lipids at a region of defined shape. We have also shown that the helices align TANGO1 around an ER exit site. We suggest this is a mechanism to prevent membrane mixing during TANGO1-mediated transfer of bulky secretory cargos from the ER to the ERGIC/Golgi via a tunnel.

Data availability

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

Article and author information

Author details

  1. Ishier Raote

    Centre for Genomic Regulation, The Barcelona Institute of Science and Technology, Barcelona, Spain
    For correspondence
    ishier.raote@crg.eu
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5898-4896
  2. Andreas M Ernst

    Department of Cell Biology, Yale School of Medicine, New Haven, United States
    Competing interests
    No competing interests declared.
  3. Felix Campelo

    ICFO - The Institute of Photonic Sciences, Barcelona, Spain
    Competing interests
    Felix Campelo, Reviewing editor, eLife.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0786-9548
  4. James E Rothman

    Department of Cell Biology, Yale School of Medicine, New Haven, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8653-8650
  5. Frederic Pincet

    Laboratoire de Physique de l'Ecole Normale Supérieure, Ecole Normal Superieure, Paris, France
    For correspondence
    frederic.pincet@ens.fr
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4243-2157
  6. Vivek Malhotra

    Centre for Genomic Regulation, The Barcelona Institute of Science and Technology, Barcelona, Spain
    For correspondence
    vivek.malhotra@crg.eu
    Competing interests
    Vivek Malhotra, Senior editor, eLife.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6198-7943

Funding

Ministerio de Economía y Competitividad (SEV-2012-0208)

  • Ishier Raote
  • Vivek Malhotra

Generalitat de Catalunya (CERCA)

  • Felix Campelo

Ministerio de Economía y Competitividad (BFU2013-44188-P)

  • Ishier Raote
  • Vivek Malhotra

Ministerio de Economía y Competitividad (CSD2009-00016)

  • Ishier Raote
  • Vivek Malhotra

Ministerio de Economía y Competitividad (IJCI-2017-34751)

  • Ishier Raote

National Institutes of Health (R35 GM118084)

  • James E Rothman

Ministerio de Economía y Competitividad (SEV-2015-0522)

  • Felix Campelo

Ministerio de Economía y Competitividad (RYC-2017-22227)

  • Felix Campelo

Fundacio Privada Cellex

  • Felix Campelo

Fundacio Privada Mir-Puig

  • Felix Campelo

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

Reviewing Editor

  1. Adam Linstedt, Carnegie Mellon University, United States

Version history

  1. Received: April 15, 2020
  2. Accepted: May 21, 2020
  3. Accepted Manuscript published: May 26, 2020 (version 1)
  4. Version of Record published: June 2, 2020 (version 2)

Copyright

© 2020, Raote 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

  • 2,873
    Page views
  • 486
    Downloads
  • 20
    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. Ishier Raote
  2. Andreas M Ernst
  3. Felix Campelo
  4. James E Rothman
  5. Frederic Pincet
  6. Vivek Malhotra
(2020)
TANGO1 membrane helices create a lipid diffusion barrier at curved membranes
eLife 9:e57822.
https://doi.org/10.7554/eLife.57822

Further reading

    1. Cell Biology
    2. Medicine
    Thao DV Le, Dianxin Liu ... Julio E Ayala
    Research Article Updated

    The canonical target of the glucagon-like peptide-1 receptor (GLP-1R), Protein Kinase A (PKA), has been shown to stimulate mechanistic Target of Rapamycin Complex 1 (mTORC1) by phosphorylating the mTOR-regulating protein Raptor at Ser791 following β-adrenergic stimulation. The objective of these studies is to test whether GLP-1R agonists similarly stimulate mTORC1 via PKA phosphorylation of Raptor at Ser791 and whether this contributes to the weight loss effect of the therapeutic GLP-1R agonist liraglutide. We measured phosphorylation of the mTORC1 signaling target ribosomal protein S6 in Chinese Hamster Ovary cells expressing GLP-1R (CHO-Glp1r) treated with liraglutide in combination with PKA inhibitors. We also assessed liraglutide-mediated phosphorylation of the PKA substrate RRXS*/T* motif in CHO-Glp1r cells expressing Myc-tagged wild-type (WT) Raptor or a PKA-resistant (Ser791Ala) Raptor mutant. Finally, we measured the body weight response to liraglutide in WT mice and mice with a targeted knock-in of PKA-resistant Ser791Ala Raptor. Liraglutide increased phosphorylation of S6 and the PKA motif in WT Raptor in a PKA-dependent manner but failed to stimulate phosphorylation of the PKA motif in Ser791Ala Raptor in CHO-Glp1r cells. Lean Ser791Ala Raptor knock-in mice were resistant to liraglutide-induced weight loss but not setmelanotide-induced (melanocortin-4 receptor-dependent) weight loss. Diet-induced obese Ser791Ala Raptor knock-in mice were not resistant to liraglutide-induced weight loss; however, there was weight-dependent variation such that there was a tendency for obese Ser791Ala Raptor knock-in mice of lower relative body weight to be resistant to liraglutide-induced weight loss compared to weight-matched controls. Together, these findings suggest that PKA-mediated phosphorylation of Raptor at Ser791 contributes to liraglutide-induced weight loss.

    1. Cell Biology
    2. Developmental Biology
    Simon Schneider, Andjela Kovacevic ... Hubert Schorle
    Research Article

    Cylicins are testis-specific proteins, which are exclusively expressed during spermiogenesis. In mice and humans, two Cylicins, the gonosomal X-linked Cylicin 1 (Cylc1/CYLC1) and the autosomal Cylicin 2 (Cylc2/CYLC2) genes, have been identified. Cylicins are cytoskeletal proteins with an overall positive charge due to lysine-rich repeats. While Cylicins have been localized in the acrosomal region of round spermatids, they resemble a major component of the calyx within the perinuclear theca at the posterior part of mature sperm nuclei. However, the role of Cylicins during spermiogenesis has not yet been investigated. Here, we applied CRISPR/Cas9-mediated gene editing in zygotes to establish Cylc1- and Cylc2-deficient mouse lines as a model to study the function of these proteins. Cylc1 deficiency resulted in male subfertility, whereas Cylc2-/-, Cylc1-/yCylc2+/-, and Cylc1-/yCylc2-/- males were infertile. Phenotypical characterization revealed that loss of Cylicins prevents proper calyx assembly during spermiogenesis. This results in decreased epididymal sperm counts, impaired shedding of excess cytoplasm, and severe structural malformations, ultimately resulting in impaired sperm motility. Furthermore, exome sequencing identified an infertile man with a hemizygous variant in CYLC1 and a heterozygous variant in CYLC2, displaying morphological abnormalities of the sperm including the absence of the acrosome. Thus, our study highlights the relevance and importance of Cylicins for spermiogenic remodeling and male fertility in human and mouse, and provides the basis for further studies on unraveling the complex molecular interactions between perinuclear theca proteins required during spermiogenesis.