The scaffolding protein Flot2 promotes cytoneme-based transport of Wnt3 in gastric cancer

  1. Daniel Routledge
  2. Sally Rogers
  3. Yosuke Ono
  4. Lucy Brunt
  5. Valerie Meniel
  6. Giusy Tornillo
  7. Hassan Ashktorab
  8. Toby Phesse
  9. Steffen Scholpp  Is a corresponding author
  1. University of Exeter, United Kingdom
  2. Cardiff University, United Kingdom
  3. Howard University, United States

Abstract

The Wnt/β-catenin signalling pathway regulates multiple cellular processes during development and many diseases, including cell proliferation, migration, and differentiation. Despite their hydrophobic nature, Wnt proteins exert their function over long distances to induce paracrine signalling. Recent studies have identified several factors involved in Wnt secretion, however, our understanding of how Wnt ligands are transported between cells to interact with their cognate receptors is still debated. Here, we demonstrate that gastric cancer cells utilise cytonemes to transport Wnt3 intercellularly to promote proliferation and cell survival. Furthermore, we identify the membrane-bound scaffolding protein Flotillin-2 (Flot2), frequently overexpressed in gastric cancer, as a modulator of these cytonemes. Together with the Wnt co-receptor and cytoneme initiator Ror2, Flot2 determines the number and length of Wnt3 cytonemes in gastric cancer. Finally, we show that Flotillins are also necessary for Wnt8a cytonemes during zebrafish embryogenesis, suggesting a conserved mechanism for Flotillin-mediated Wnt transport on cytonemes in development and disease.

Data availability

All data generated or analysed during this study are included in the manuscript, supporting files and source files; Supporting Data files and Source Data have been provided to all figures.

Article and author information

Author details

  1. Daniel Routledge

    Living Systems Institute, University of Exeter, Exeter, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  2. Sally Rogers

    Living Systems Institute, University of Exeter, Exeter, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  3. Yosuke Ono

    Living Systems Institute, University of Exeter, Exeter, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  4. Lucy Brunt

    Living Systems Institute, University of Exeter, Exeter, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  5. Valerie Meniel

    The European Cancer Stem Cell Research Institute, Cardiff University, Cardiff, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  6. Giusy Tornillo

    The European Cancer Stem Cell Research Institute, Cardiff University, Cardiff, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  7. Hassan Ashktorab

    Department of Medicine, Howard University, Washington, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Toby Phesse

    The European Cancer Stem Cell Research Institute, Cardiff University, Cardiff, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9568-4916
  9. Steffen Scholpp

    Living Systems Institute, University of Exeter, Exeter, United Kingdom
    For correspondence
    s.scholpp@exeter.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4903-9657

Funding

Medical Research Council (MR/N0137941/1)

  • Daniel Routledge

Medical Research Council (MR/S007970/1)

  • Sally Rogers
  • Steffen Scholpp

Biotechnology and Biological Sciences Research Council (BB/S016295/1)

  • Yosuke Ono
  • Lucy Brunt
  • Steffen Scholpp

Medical Research Council (MR/R026424/1)

  • Valerie Meniel
  • Giusy Tornillo
  • Toby Phesse

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

Reviewing Editor

  1. Julia Christina Gross, Health and Medical University

Ethics

Animal experimentation: Zebrafish care and all experimental procedures were carried out in accordance with the European Communities Council Directive (2010/63/EU) and Animals Scientific Procedures Act (ASPA) 1986. Zebrafish experimental procedures were carried out under personal and project licenses granted by the UK Home Office under ASPA, and ethically approved by the Animal Welfare and Ethical Review Body at the University of Exeter.

Version history

  1. Preprint posted: January 8, 2022 (view preprint)
  2. Received: January 26, 2022
  3. Accepted: August 27, 2022
  4. Accepted Manuscript published: August 30, 2022 (version 1)
  5. Version of Record published: September 8, 2022 (version 2)

Copyright

© 2022, Routledge 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

  • 1,343
    views
  • 299
    downloads
  • 6
    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. Daniel Routledge
  2. Sally Rogers
  3. Yosuke Ono
  4. Lucy Brunt
  5. Valerie Meniel
  6. Giusy Tornillo
  7. Hassan Ashktorab
  8. Toby Phesse
  9. Steffen Scholpp
(2022)
The scaffolding protein Flot2 promotes cytoneme-based transport of Wnt3 in gastric cancer
eLife 11:e77376.
https://doi.org/10.7554/eLife.77376

Share this article

https://doi.org/10.7554/eLife.77376

Further reading

    1. Cancer Biology
    2. Cell Biology
    Linda Zhang, Joanne I Hsu ... Margaret A Goodell
    Research Article

    The DNA damage response is critical for maintaining genome integrity and is commonly disrupted in the development of cancer. PPM1D (protein phosphatase Mg2+/Mn2+-dependent 1D) is a master negative regulator of the response; gain-of-function mutations and amplifications of PPM1D are found across several human cancers making it a relevant pharmacological target. Here, we used CRISPR/Cas9 screening to identify synthetic-lethal dependencies of PPM1D, uncovering superoxide dismutase-1 (SOD1) as a potential target for PPM1D-mutant cells. We revealed a dysregulated redox landscape characterized by elevated levels of reactive oxygen species and a compromised response to oxidative stress in PPM1D-mutant cells. Altogether, our results demonstrate a role for SOD1 in the survival of PPM1D-mutant leukemia cells and highlight a new potential therapeutic strategy against PPM1D-mutant cancers.

    1. Cancer Biology
    Haley R Noonan, Alexandra M Thornock ... Leonard I Zon
    Research Article

    Developmental signaling pathways associated with growth factors such as TGFb are commonly dysregulated in melanoma. Here we identified a human TGFb enhancer specifically activated in melanoma cells treated with TGFB1 ligand. We generated stable transgenic zebrafish with this TGFb Induced Enhancer driving green fluorescent protein (TIE:EGFP). TIE:EGFP was not expressed in normal melanocytes or early melanomas but was expressed in spatially distinct regions of advanced melanomas. Single-cell RNA-sequencing revealed that TIE:EGFP+ melanoma cells down-regulated interferon response while up-regulating a novel set of chronic TGFb target genes. ChIP-sequencing demonstrated that AP-1 factor binding is required for activation of chronic TGFb response. Overexpression of SATB2, a chromatin remodeler associated with tumor spreading, showed activation of TGFb signaling in early melanomas. Confocal imaging and flow cytometric analysis showed that macrophages localize to TIE:EGFP+ regions and preferentially phagocytose TIE:EGFP+ melanoma cells compared to TIE:EGFP- melanoma cells. This work identifies a TGFb induced immune response and demonstrates the need for the development of chronic TGFb biomarkers to predict patient response to TGFb inhibitors.