Robo recruitment of the Wave Regulatory Complex plays an essential and conserved role in midline repulsion

  1. Karina Chaudhari
  2. Madhavi Gorla
  3. Chao Chang
  4. Artur Kania
  5. Greg J Bashaw  Is a corresponding author
  1. University of Pennsylvania, United States
  2. Institut de recherches cliniques de Montréal (IRCM), Canada

Abstract

The Roundabout (Robo) guidance receptor family induces axon repulsion in response to its ligand Slit by inducing local cytoskeletal changes; however, the link to the cytoskeleton and the nature of these cytoskeletal changes are poorly understood. Here, we show that the heteropentameric Scar/Wave Regulatory Complex (WRC) which drives Arp2/3-induced branched actin polymerization, is a direct effector of Robo signaling. Biochemical evidence shows that Slit triggers WRC recruitment to the Robo receptor's WIRS motif. In Drosophila embryos, mutants of the WRC enhance Robo1-dependent midline crossing defects. Additionally, mutating Robo1's WIRS motif significantly reduces receptor activity in rescue assays in vivo, and CRISPR-Cas9 mutagenesis shows that the WIRS motif is essential for endogenous Robo1 function. Finally, axon guidance assays in mouse dorsal spinal commissural axons and gain-of-function experiments in chick embryos demonstrate that the WIRS motif is also required for Robo1 repulsion in mammals. Together, our data support an essential conserved role for the WIRS-WRC interaction in Robo1-mediated axon repulsion.

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. Karina Chaudhari

    Department of Neuroscience, Perelman School of Medicine, University of Pennsylvania, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3533-3027
  2. Madhavi Gorla

    Department of Neuroscience, Perelman School of Medicine, University of Pennsylvania, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Chao Chang

    Anatomy and Cell Biology, Institut de recherches cliniques de Montréal (IRCM), Montreal, Canada
    Competing interests
    The authors declare that no competing interests exist.
  4. Artur Kania

    Anatomy and Cell Biology, Institut de recherches cliniques de Montréal (IRCM), Montreal, Canada
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5209-2520
  5. Greg J Bashaw

    Department of Neuroscience, Perelman School of Medicine, University of Pennsylvania, Philadelphia, United States
    For correspondence
    gbashaw@pennmedicine.upenn.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6146-0962

Funding

National Institutes of Health (R35 NS097340)

  • Greg J Bashaw

National Science Foundation (IOS-1853719)

  • Greg J Bashaw

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

Ethics

Animal experimentation: This study was performed in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. All of the animals were handled according to approved institutional animal care and use committee (IACUC) protocols of the Perelman School of Medicine at the University of Pennsylvania (Protocol #806216).

Reviewing Editor

  1. Paola Bovolenta, CSIC-UAM, Spain

Publication history

  1. Received: October 29, 2020
  2. Accepted: April 6, 2021
  3. Accepted Manuscript published: April 12, 2021 (version 1)
  4. Version of Record published: May 4, 2021 (version 2)

Copyright

© 2021, Chaudhari 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.

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  1. Karina Chaudhari
  2. Madhavi Gorla
  3. Chao Chang
  4. Artur Kania
  5. Greg J Bashaw
(2021)
Robo recruitment of the Wave Regulatory Complex plays an essential and conserved role in midline repulsion
eLife 10:e64474.
https://doi.org/10.7554/eLife.64474

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