Differential requirements for cyclase-associated protein (CAP) in actin-dependent processes of Toxoplasma gondii

  1. Alex Hunt
  2. Matthew Robert Geoffrey Russell
  3. Jeanette Wagener
  4. Robyn Kent
  5. Romain Carmeille
  6. Christopher J Peddie
  7. Lucy Collinson
  8. Aoife Heaslip
  9. Gary E Ward
  10. Moritz Treeck  Is a corresponding author
  1. The Francis Crick Institute, United Kingdom
  2. University of Vermont, United States
  3. University of Connecticut, United States

Abstract

Toxoplasma gondii contains a limited subset of actin binding proteins. Here we show that the putative actin regulator cyclase-associated protein (CAP) is present in two different isoforms and its deletion leads to significant defects in some but not all actin dependent processes. We observe defects in cell-cell communication, daughter cell orientation and the juxtanuclear accumulation of actin, but only modest defects in synchronicity of division and no defect in the replication of the apicoplast. 3D electron microscopy reveals that loss of CAP results in a defect in formation of a normal central residual body, but parasites remain connected within the vacuole. This dissociates synchronicity of division and parasite rosetting and reveals that establishment and maintenance of the residual body may be more complex than previously thought. These results highlight the different spatial requirements for F-actin regulation in Toxoplasma which appear to be achieved by partially overlapping functions of actin regulators.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for Figures 4, 5, 6, 8 and 10. Data Availability: Raw data for FIB SEM supporting movies will be uploaded to EMPIAR. Raw data for FIB SEM supporting movies have been deposited to EMPIAR.

Article and author information

Author details

  1. Alex Hunt

    Signalling in Apicomplexan Parasites Laboratory, The Francis Crick Institute, London, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7431-7156
  2. Matthew Robert Geoffrey Russell

    Electron Microscopy Science Technology Platform, The Francis Crick Institute, London, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-4608-7669
  3. Jeanette Wagener

    Signalling in Apicomplexan Parasites Laboratory, The Francis Crick Institute, London, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7227-4348
  4. Robyn Kent

    Department of Microbiology and Molecular Genetics, University of Vermont, Burlington, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Romain Carmeille

    Department of Molecular and Cell Biology, University of Connecticut, Storrs, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Christopher J Peddie

    Electron Microscopy Science Technology Platform, The Francis Crick Institute, London, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  7. Lucy Collinson

    Electron Microscopy Science Technology Platform, The Francis Crick Institute, London, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  8. Aoife Heaslip

    Department of Molecular and Cell Biology, University of Connecticut, Storrs, United States
    Competing interests
    The authors declare that no competing interests exist.
  9. Gary E Ward

    Deaprtment of Microbiology and Molecular Genetics, University of Vermont, Burlington, United States
    Competing interests
    The authors declare that no competing interests exist.
  10. Moritz Treeck

    Signalling in Apicomplexan Parasites Laboratory, The Francis Crick Institute, London, United Kingdom
    For correspondence
    moritz.treeck@crick.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9727-6657

Funding

Francis Crick Institute (FC001189)

  • Lucy Collinson

NIH Office of the Director (AI121885)

  • Romain Carmeille
  • Aoife Heaslip

NIH Office of the Director (AI139201)

  • Robyn Kent
  • Gary E Ward

Francis Crick Institute (FC001999)

  • Matthew Robert Geoffrey Russell
  • Christopher J Peddie
  • Lucy Collinson

NIH Office of the Director (AI137767)

  • Robyn Kent
  • Gary E Ward

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

Ethics

Animal experimentation: All experiments were performed in accordance with UK Home Office regulations (PPL 80/2616) and approved by the ethical review panel at the Francis Crick Institute.

Copyright

© 2019, Hunt 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

  • 2,027
    views
  • 300
    downloads
  • 46
    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. Alex Hunt
  2. Matthew Robert Geoffrey Russell
  3. Jeanette Wagener
  4. Robyn Kent
  5. Romain Carmeille
  6. Christopher J Peddie
  7. Lucy Collinson
  8. Aoife Heaslip
  9. Gary E Ward
  10. Moritz Treeck
(2019)
Differential requirements for cyclase-associated protein (CAP) in actin-dependent processes of Toxoplasma gondii
eLife 8:e50598.
https://doi.org/10.7554/eLife.50598

Share this article

https://doi.org/10.7554/eLife.50598

Further reading

    1. Microbiology and Infectious Disease
    Mehak Zahoor Khan, Debbie M Hunt ... Vinay Kumar Nandicoori
    Research Article

    Mycobacterium tuberculosis’s (Mtb) autarkic lifestyle within the host involves rewiring its transcriptional networks to combat host-induced stresses. With the help of RNA sequencing performed under various stress conditions, we identified that genes belonging to Mtb sulfur metabolism pathways are significantly upregulated during oxidative stress. Using an integrated approach of microbial genetics, transcriptomics, metabolomics, animal experiments, chemical inhibition, and rescue studies, we investigated the biological role of non-canonical L-cysteine synthases, CysM and CysK2. While transcriptome signatures of RvΔcysM and RvΔcysK2 appear similar under regular growth conditions, we observed unique transcriptional signatures when subjected to oxidative stress. We followed pool size and labelling (34S) of key downstream metabolites, viz. mycothiol and ergothioneine, to monitor L-cysteine biosynthesis and utilization. This revealed the significant role of distinct L-cysteine biosynthetic routes on redox stress and homeostasis. CysM and CysK2 independently facilitate Mtb survival by alleviating host-induced redox stress, suggesting they are not fully redundant during infection. With the help of genetic mutants and chemical inhibitors, we show that CysM and CysK2 serve as unique, attractive targets for adjunct therapy to combat mycobacterial infection.

    1. Microbiology and Infectious Disease
    2. Physics of Living Systems
    Tingting Yang, Marko S Chavez ... Mohamed Y El-Naggar
    Research Article

    Filamentous multicellular cable bacteria perform centimeter-scale electron transport in a process that couples oxidation of an electron donor (sulfide) in deeper sediment to the reduction of an electron acceptor (oxygen or nitrate) near the surface. While this electric metabolism is prevalent in both marine and freshwater sediments, detailed electronic measurements of the conductivity previously focused on the marine cable bacteria (Candidatus Electrothrix), rather than freshwater cable bacteria, which form a separate genus (Candidatus Electronema) and contribute essential geochemical roles in freshwater sediments. Here, we characterize the electron transport characteristics of Ca. Electronema cable bacteria from Southern California freshwater sediments. Current–voltage measurements of intact cable filaments bridging interdigitated electrodes confirmed their persistent conductivity under a controlled atmosphere and the variable sensitivity of this conduction to air exposure. Electrostatic and conductive atomic force microscopies mapped out the characteristics of the cell envelope’s nanofiber network, implicating it as the conductive pathway in a manner consistent with previous findings in marine cable bacteria. Four-probe measurements of microelectrodes addressing intact cables demonstrated nanoampere currents up to 200 μm lengths at modest driving voltages, allowing us to quantify the nanofiber conductivity at 0.1 S/cm for freshwater cable bacteria filaments under our measurement conditions. Such a high conductivity can support the remarkable sulfide-to-oxygen electrical currents mediated by cable bacteria in sediments. These measurements expand the knowledgebase of long-distance electron transport to the freshwater niche while shedding light on the underlying conductive network of cable bacteria.