Distinct regions of H. pylori's bactofilin CcmA regulate protein-protein interactions to control helical cell shape

  1. Sophie R Sichel
  2. Benjamin P Bratton
  3. Nina Reda Salama  Is a corresponding author
  1. Fred Hutchinson Cancer Research Center, United States
  2. Vanderbilt University Medical Center, United States

Abstract

The helical shape of H. pylori cells promotes robust stomach colonization, however, how the helical shape of H. pylori cells is determined is unresolved. Previous work identified helical-cell-shape-promoting protein complexes containing a peptidoglycan-hydrolase (Csd1), a peptidoglycan precursor synthesis enzyme (MurF), a non-enzymatic homologue of Csd1 (Csd2), non-enzymatic transmembrane proteins (Csd5 and Csd7), and a bactofilin (CcmA). Bactofilins are highly conserved, spontaneously polymerizing cytoskeletal bacterial proteins. We sought to understand CcmA's function in generating the helical shape of H. pylori cells. Using CcmA deletion analysis, in vitro polymerization, and in vivo co-immunoprecipitation experiments we identified that the bactofilin domain and N-terminal region of CcmA are required for helical cell shape and the bactofilin domain of CcmA is sufficient for polymerization and interactions with Csd5 and Csd7. We also found that CcmA's N-terminal region inhibits interaction with Csd7. Deleting the N-terminal region of CcmA increases CcmA-Csd7 interactions and destabilizes the peptidoglycan-hydrolase Csd1. Using super-resolution microscopy, we found that Csd5 recruits CcmA to the cell envelope and promotes CcmA enrichment at the major helical axis. Thus, CcmA helps organize cell-shape-determining proteins and peptidoglycan synthesis machinery to coordinate cell wall modification and synthesis, promoting the curvature required to build a helical cell.

Data availability

Data generated or analysed during this study are included in the manuscript and supporting file; Source Data files have been provided for Figures 2, 5, 6 and 7.Microscopy data are available at BioImage Archive and accession code is S-BIAD462

The following data sets were generated

Article and author information

Author details

  1. Sophie R Sichel

    Division of Human Biology, Fred Hutchinson Cancer Research Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Benjamin P Bratton

    Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1128-2560
  3. Nina Reda Salama

    Division of Human Biology, Fred Hutchinson Cancer Research Center, Seattle, United States
    For correspondence
    nsalama@fredhutch.org
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2762-1424

Funding

National Institute of Allergy and Infectious Diseases (F31 AI152331)

  • Sophie R Sichel

National Institute of Allergy and Infectious Diseases (R01 AI136946)

  • Nina Reda Salama

National Institute of General Medical Sciences (T32 GM95421)

  • Sophie R Sichel

GO-MAP Graduat Opportunity Program Research Assistantship Award (Sophie Sichel)

  • Sophie R Sichel

VUMC Discovery Scholars in Health and Medicine Program (Benjamin Bratton)

  • Benjamin P Bratton

National Cancer Institute (P31 CA015704)

  • Nina Reda Salama

Audacious Project (Institute for Protein Design)

  • Sophie R Sichel

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

Reviewing Editor

  1. Petra Anne Levin, Washington University in St. Louis, United States

Version history

  1. Preprint posted: April 5, 2022 (view preprint)
  2. Received: May 9, 2022
  3. Accepted: September 7, 2022
  4. Accepted Manuscript published: September 8, 2022 (version 1)
  5. Version of Record published: September 23, 2022 (version 2)

Copyright

© 2022, Sichel 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

  • 1,341
    views
  • 219
    downloads
  • 12
    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. Sophie R Sichel
  2. Benjamin P Bratton
  3. Nina Reda Salama
(2022)
Distinct regions of H. pylori's bactofilin CcmA regulate protein-protein interactions to control helical cell shape
eLife 11:e80111.
https://doi.org/10.7554/eLife.80111

Share this article

https://doi.org/10.7554/eLife.80111

Further reading

    1. Immunology and Inflammation
    2. Microbiology and Infectious Disease
    Ffion R Hammond, Amy Lewis ... Philip M Elks
    Research Article

    Tuberculosis is a major global health problem and is one of the top 10 causes of death worldwide. There is a pressing need for new treatments that circumvent emerging antibiotic resistance. Mycobacterium tuberculosis parasitises macrophages, reprogramming them to establish a niche in which to proliferate, therefore macrophage manipulation is a potential host-directed therapy if druggable molecular targets could be identified. The pseudokinase Tribbles1 (Trib1) regulates multiple innate immune processes and inflammatory profiles making it a potential drug target in infections. Trib1 controls macrophage function, cytokine production, and macrophage polarisation. Despite wide-ranging effects on leukocyte biology, data exploring the roles of Tribbles in infection in vivo are limited. Here, we identify that human Tribbles1 is expressed in monocytes and is upregulated at the transcript level after stimulation with mycobacterial antigen. To investigate the mechanistic roles of Tribbles in the host response to mycobacteria in vivo, we used a zebrafish Mycobacterium marinum (Mm) infection tuberculosis model. Zebrafish Tribbles family members were characterised and shown to have substantial mRNA and protein sequence homology to their human orthologues. trib1 overexpression was host-protective against Mm infection, reducing burden by approximately 50%. Conversely, trib1 knockdown/knockout exhibited increased infection. Mechanistically, trib1 overexpression significantly increased the levels of proinflammatory factors il-1β and nitric oxide. The host-protective effect of trib1 was found to be dependent on the E3 ubiquitin kinase Cop1. These findings highlight the importance of Trib1 and Cop1 as immune regulators during infection in vivo and suggest that enhancing macrophage TRIB1 levels may provide a tractable therapeutic intervention to improve bacterial infection outcomes in tuberculosis.

    1. Microbiology and Infectious Disease
    2. Physics of Living Systems
    Chi Zhang, Rongjing Zhang, Junhua Yuan
    Research Article

    Bacteria in biofilms secrete potassium ions to attract free swimming cells. However, the basis of chemotaxis to potassium remains poorly understood. Here, using a microfluidic device, we found that Escherichia coli can rapidly accumulate in regions of high potassium concentration on the order of millimoles. Using a bead assay, we measured the dynamic response of individual flagellar motors to stepwise changes in potassium concentration, finding that the response resulted from the chemotaxis signaling pathway. To characterize the chemotactic response to potassium, we measured the dose–response curve and adaptation kinetics via an Förster resonance energy transfer (FRET) assay, finding that the chemotaxis pathway exhibited a sensitive response and fast adaptation to potassium. We further found that the two major chemoreceptors Tar and Tsr respond differently to potassium. Tar receptors exhibit a biphasic response, whereas Tsr receptors respond to potassium as an attractant. These different responses were consistent with the responses of the two receptors to intracellular pH changes. The sensitive response and fast adaptation allow bacteria to sense and localize small changes in potassium concentration. The differential responses of Tar and Tsr receptors to potassium suggest that cells at different growth stages respond differently to potassium and may have different requirements for potassium.