Soluble MAC is primarily released from MAC-resistant bacteria that potently convert complement component C5

Abstract

The Membrane Attack Complex (MAC or C5b-9) is an important effector of the immune system to kill invading microbes. MAC formation is initiated when complement enzymes on the bacterial surface convert complement component C5 into C5b. Although the MAC is a membrane-inserted complex, soluble forms of MAC (sMAC, or terminal complement complex (TCC)) are often detected in sera of patients suffering from infections. Consequently, sMAC has been proposed as a biomarker, but it remains unclear when and how it is formed during infections. Here, we studied mechanisms of MAC formation on different Gram-negative and Gram-positive bacteria and found that sMAC is primarily formed in human serum by bacteria resistant to MAC-dependent killing. Surprisingly, C5 was converted into C5b more potently by MAC-resistant compared to MAC-sensitive Escherichia coli strains. In addition, we found that MAC precursors are released from the surface of MAC-resistant bacteria during MAC assembly. Although release of MAC precursors from bacteria induced lysis of bystander human erythrocytes, serum regulators vitronectin (Vn) and clusterin (Clu) can prevent this. Combining size exclusion chromatography with mass spectrometry profiling, we show that sMAC released from bacteria in serum is a heterogeneous mixture of complexes composed of C5b-8, up to 3 copies of C9 and multiple copies of Vn and Clu. Altogether, our data provide molecular insight into how sMAC is generated during bacterial infections. This fundamental knowledge could form the basis for exploring the use of sMAC as biomarker.

Data availability

All relevant data supporting the findings of this manuscript have been added in the main manuscript and supplemental information. Supporting source data (for figures 1 - 6a and supplements) have been uploaded to Dryad Digital Repository (doi:10.5061/dryad.g4f4qrfsd). The MS data (in figure 6, figure 6 supplement and figure 7) have been deposited to the ProteomeXchange partner MassIVE database and assigned the identifier MSV000088560 (doi:10.25345/C5QW00).

The following data sets were generated

Article and author information

Author details

  1. Dennis J Doorduijn

    Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, Netherlands
    Competing interests
    The authors declare that no competing interests exist.
  2. Marie V Lukassen

    Biomolecular Mass Spectrometry and Proteomics, Utrecht University, Utrecht, Netherlands
    Competing interests
    The authors declare that no competing interests exist.
  3. Marije FL van 't Wout

    Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, Netherlands
    Competing interests
    The authors declare that no competing interests exist.
  4. Vojtech Franc

    Biomolecular Mass Spectrometry and Proteomics, Utrecht University, Utrecht, Netherlands
    Competing interests
    The authors declare that no competing interests exist.
  5. Maartje Ruyken

    Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, Netherlands
    Competing interests
    The authors declare that no competing interests exist.
  6. Bart W Bardoel

    Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, Netherlands
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6450-277X
  7. Albert JR Heck

    Biomolecular Mass Spectrometry and Proteomics, Utrecht University, Utrecht, Netherlands
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2405-4404
  8. Suzan HM Rooijakkers

    Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, Netherlands
    For correspondence
    s.h.m.rooijakkers@umcutrecht.nl
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-4102-0377

Funding

European Research Council ((639209-ComBact)

  • Suzan HM Rooijakkers

Nederlandse Organisatie voor Wetenschappelijk Onderzoek (Aspasia)

  • Suzan HM Rooijakkers

Utrecht Molecular Immunology HUB (eSTIMATE)

  • Suzan HM Rooijakkers

Netherlands Proteomics Centre (184.034.019)

  • Albert JR Heck

Independent Research Fund Denmark (9036-00007B)

  • Marie V Lukassen

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

Reviewing Editor

  1. Jameel Iqbal, DaVita Labs, United States

Version history

  1. Preprint posted: December 15, 2021 (view preprint)
  2. Received: February 1, 2022
  3. Accepted: August 1, 2022
  4. Accepted Manuscript published: August 10, 2022 (version 1)
  5. Version of Record published: August 24, 2022 (version 2)

Copyright

© 2022, Doorduijn 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. Dennis J Doorduijn
  2. Marie V Lukassen
  3. Marije FL van 't Wout
  4. Vojtech Franc
  5. Maartje Ruyken
  6. Bart W Bardoel
  7. Albert JR Heck
  8. Suzan HM Rooijakkers
(2022)
Soluble MAC is primarily released from MAC-resistant bacteria that potently convert complement component C5
eLife 11:e77503.
https://doi.org/10.7554/eLife.77503

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https://doi.org/10.7554/eLife.77503

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