A novel mode of Capping Protein-regulation by Twinfilin

  1. Adam B Johnston
  2. Denise M Hilton
  3. Patrick McConnell
  4. Britney Johnson
  5. Meghan T Harris
  6. Avital Simone
  7. Gaya K Amarasinghe
  8. John A Cooper
  9. Bruce L Goode  Is a corresponding author
  1. Brandeis University, United States
  2. Washington University in St Louis, United States

Abstract

Cellular actin assembly is controlled at the barbed ends of actin filaments, where capping protein (CP) limits polymerization. Twinfilin is a conserved in vivo binding partner of CP, yet the significance of this interaction has remained a mystery. Here, we discover that the C-terminal tail of Twinfilin harbors a CP-interacting (CPI) motif, identifying it as a novel CPI-motif protein. Twinfilin and the CPI-motif protein CARMIL have overlapping binding sites on CP. Further, Twinfilin binds competitively with CARMIL to CP, protecting CP from barbed-end displacement by CARMIL. Twinfilin also accelerates dissociation of the CP inhibitor V-1, restoring CP to an active capping state. Knockdowns of Twinfilin and CP each cause similar defects in cell morphology, and elevated Twinfilin expression rescues defects caused by CARMIL hyperactivity. Together, these observations define Twinfilin as the first 'pro-capping' ligand of CP and lead us to propose important revisions to our understanding of the CP regulatory cycle.

Data availability

All datasets associated with this article are included in the manuscript and supporting files.

Article and author information

Author details

  1. Adam B Johnston

    Department of Biology, Rosenstiel Basic Medical Science Research Center, Brandeis University, Waltham, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Denise M Hilton

    Department of Biology, Rosenstiel Basic Medical Science Research Center, Brandeis University, Waltham, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1577-1855
  3. Patrick McConnell

    Department of Biochemistry and Molecular Biophysics, Washington University in St Louis, St Louis, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Britney Johnson

    Department of Pathology and Immunology, Washington University in St Louis, St Louis, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Meghan T Harris

    Department of Biology, Rosenstiel Basic Medical Science Research Center, Brandeis University, Waltham, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Avital Simone

    Department of Biology, Rosenstiel Basic Medical Science Research Center, Brandeis University, Waltham, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Gaya K Amarasinghe

    Department of Pathology and Immunology, Washington University in St Louis, St Louis, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. John A Cooper

    Department of Biochemistry and Molecular Biophysics, Washington University in St Louis, St Louis, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0933-4571
  9. Bruce L Goode

    Department of Biology, Rosenstiel Basic Medical Science Research Center, Brandeis University, Waltham, United States
    For correspondence
    goode@brandeis.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6443-5893

Funding

National Institutes of Health (R01 GM063691)

  • Bruce L Goode

Defense Threat Reduction Agency (HDTRA1-16-1-0033)

  • Gaya K Amarasinghe

National Institutes of Health (R35 GM118171)

  • John A Cooper

National Science Foundation (MRSEC DMR-1420382)

  • Bruce L Goode

National Institutes of Health (P01 AI120943)

  • Gaya K Amarasinghe

National Institutes of Health (R01 AI123926)

  • Gaya K Amarasinghe

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

Copyright

© 2018, Johnston 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.

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. Adam B Johnston
  2. Denise M Hilton
  3. Patrick McConnell
  4. Britney Johnson
  5. Meghan T Harris
  6. Avital Simone
  7. Gaya K Amarasinghe
  8. John A Cooper
  9. Bruce L Goode
(2018)
A novel mode of Capping Protein-regulation by Twinfilin
eLife 7:e41313.
https://doi.org/10.7554/eLife.41313

Share this article

https://doi.org/10.7554/eLife.41313

Further reading

    1. Cell Biology
    2. Developmental Biology
    Jeet H Patel, Mary C Mullins
    Insight

    Disease-causing mutations in the signaling protein BMP4 impair its secretion, but only when it is made as a homodimer.

    1. Cell Biology
    2. Genetics and Genomics
    Róża K Przanowska, Yuechuan Chen ... Anindya Dutta
    Research Article

    The six-subunit ORC is essential for the initiation of DNA replication in eukaryotes. Cancer cell lines in culture can survive and replicate DNA replication after genetic inactivation of individual ORC subunits, ORC1, ORC2, or ORC5. In primary cells, ORC1 was dispensable in the mouse liver for endo-reduplication, but this could be explained by the ORC1 homolog, CDC6, substituting for ORC1 to restore functional ORC. Here, we have created mice with a conditional deletion of ORC2, which does not have a homolog. Although mouse embryo fibroblasts require ORC2 for proliferation, mouse hepatocytes synthesize DNA in cell culture and endo-reduplicate in vivo without ORC2. Mouse livers endo-reduplicate after simultaneous deletion of ORC1 and ORC2 both during normal development and after partial hepatectomy. Since endo-reduplication initiates DNA synthesis like normal S phase replication these results unequivocally indicate that primary cells, like cancer cell lines, can load MCM2-7 and initiate replication without ORC.