Structural analysis of the Legionella pneumophila Dot/Icm Type IV Secretion System Core Complex

  1. Clarissa L Durie
  2. Michael J Sheedlo
  3. Jeong Min Chung
  4. Brenda G Byrne
  5. Min Su
  6. Thomas Knight
  7. Michele Swanson  Is a corresponding author
  8. D Borden Lacy  Is a corresponding author
  9. Melanie D Ohi  Is a corresponding author
  1. University Of Michigan, United States
  2. Vanderbilt University Medical Center, United States
  3. University of Michigan, United States

Abstract

Legionella pneumophila is an opportunistic pathogen that causes the potentially fatal pneumonia Legionnaires' Disease. This infection and subsequent pathology require the Dot/Icm Type IV Secretion System (T4SS) to deliver effector proteins into host cells. Compared to prototypical T4SSs, the Dot/Icm assembly is much larger, containing ~27 different components including a core complex reported to be composed of five proteins: DotC, DotD, DotF, DotG, and DotH. Using single particle cryo-electron microscopy (cryo-EM), we report reconstructions of the core complex of the Dot/Icm T4SS that includes a symmetry mismatch between distinct structural features of the outer membrane cap (OMC) and periplasmic ring (PR). We present models of known core complex proteins, DotC, DotD, and DotH, and two structurally similar proteins within the core complex, DotK and Lpg0657. This analysis reveals the stoichiometry and contact interfaces between the key proteins of the Dot/Icm T4SS core complex and provides a framework for understanding a complex molecular machine.

Data availability

All cryo-EM data included in this manuscript are available through the Electron Microscopy Data Bank (EMD-22068, EMD-22069, EMD-22070 and EMD-22071). All models that were constructed from these data are available via the Protein Data Bank (PDB 6x62, 6x64, 6x65, and 6x66).

The following data sets were generated

Article and author information

Author details

  1. Clarissa L Durie

    Life Sciences Institute, University Of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4027-4386
  2. Michael J Sheedlo

    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:" 0000-0002-3185-1727
  3. Jeong Min Chung

    Life Sciences Institute, University of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4285-8764
  4. Brenda G Byrne

    Department of Microbiology & Immunology, University of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Min Su

    Life Sciences Institute, University of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Thomas Knight

    Department of Microbiology & Immunology, University of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Michele Swanson

    Department of Microbiology & Immunology, University of Michigan, Ann Arbor, United States
    For correspondence
    mswanson@umich.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2542-0266
  8. D Borden Lacy

    Pathology, Microbiology and Immunology; Biochemistry, Vanderbilt University Medical Center, Nashville, United States
    For correspondence
    borden.lacy@vanderbilt.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2273-8121
  9. Melanie D Ohi

    Life Sciences Institute, University of Michigan, Ann Arbor, United States
    For correspondence
    mohi@umich.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1750-4793

Funding

National Institute of Allergy and Infectious Diseases (F32 AI150027-01)

  • Clarissa L Durie

National Institute of General Medical Sciences (S10OD020011)

  • Melanie D Ohi

National Institute of Allergy and Infectious Diseases (2T32DK007673)

  • Michele Swanson

National Institute of Allergy and Infectious Diseases (R01AI118932)

  • Melanie D Ohi

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

Copyright

© 2020, Durie 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. Clarissa L Durie
  2. Michael J Sheedlo
  3. Jeong Min Chung
  4. Brenda G Byrne
  5. Min Su
  6. Thomas Knight
  7. Michele Swanson
  8. D Borden Lacy
  9. Melanie D Ohi
(2020)
Structural analysis of the Legionella pneumophila Dot/Icm Type IV Secretion System Core Complex
eLife 9:e59530.
https://doi.org/10.7554/eLife.59530

Share this article

https://doi.org/10.7554/eLife.59530

Further reading

    1. Microbiology and Infectious Disease
    Dawid S Zyla
    Insight

    A combination of imaging techniques reveals how herpes simplex virus type 1 assembles within infected cells, highlighting the roles of essential viral proteins in viral assembly and exit.

    1. Microbiology and Infectious Disease
    Dhaval Ghone, Edward L Evans ... Aussie Suzuki
    Research Article

    Virion Infectivity Factor (Vif) of the Human Immunodeficiency Virus type 1 (HIV-1) targets and degrades cellular APOBEC3 proteins, key regulators of intrinsic and innate antiretroviral immune responses, thereby facilitating HIV-1 infection. While Vif’s role in degrading APOBEC3G is well-studied, Vif is also known to cause cell cycle arrest, but the detailed nature of Vif’s effects on the cell cycle has yet to be delineated. In this study, we employed high-temporal resolution single-cell live imaging and super-resolution microscopy to monitor individual cells during Vif-induced cell cycle arrest. Our findings reveal that Vif does not affect the G2/M boundary as previously thought. Instead, Vif triggers a unique and robust pseudo-metaphase arrest, distinct from the mild prometaphase arrest induced by Vpr. During this arrest, chromosomes align properly and form the metaphase plate, but later lose alignment, resulting in polar chromosomes. Notably, Vif, unlike Vpr, significantly reduces the levels of both Protein Phosphatase 1 (PP1) and 2 A (PP2A) at kinetochores, which regulate chromosome-microtubule interactions. These results unveil a novel role for Vif in kinetochore regulation that governs the spatial organization of chromosomes during mitosis.