A single regulator NrtR controls bacterial NAD+ homeostasis via its acetylation

  1. Rongsui Gao
  2. Wenhui Wei
  3. Bachar H Hassan
  4. Jun Li
  5. Jiao-Yu Deng
  6. Youjun Feng  Is a corresponding author
  1. Zhejiang University School of Medicine, China
  2. Stony Brook University, United States
  3. Zhejiang University of Technology, China
  4. Chinese Academy of Sciences, China

Abstract

Nicotinamide adenine dinucleotide (NAD+) is an indispensable cofactor in all domains of life, the homeostasis of which requires tight regulation. Here we report that a Nudix-related transcriptional factor, designated MsNrtR (MSMEG_3198), controls the de novo pathway of NAD+ biosynthesis in M. smegmatis, a non-tuberculosis Mycobacterium. The integrated evidence in vitro and in vivo confirms that MsNrtR is an auto-repressor, and negatively controls the de novo NAD+ biosynthetic pathway. Binding of MsNrtR cognate DNA is finely mapped, which can be disrupted by an ADP-ribose intermediate. Unexpectedly, we discover that the acetylation of MsNrtR at Lysine 134 participates in the homeostasis of intra-cellular NAD+ level in M. smegmatis. Furthermore, we demonstrate that NrtR acetylation proceeds via the non-enzymatic acetyl-phosphate (AcP) route rather than the enzymatic Pat/CobB pathway. In addition, the acetylation of NrtR also occurs in its paralogs of Gram-positive bacterium Streptococcus and Gram-negative bacterium Vibrio, suggesting a common mechanism of post-translational modification in the context of NAD+ homeostasis. Together, it represents a first paradigm for the recruitment of acetylated NrtR to regulate bacterial central NAD+ metabolism.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files.

Article and author information

Author details

  1. Rongsui Gao

    Department of Pathogen Biology and Microbiology, Zhejiang University School of Medicine, Hangzhou, China
    Competing interests
    The authors declare that no competing interests exist.
  2. Wenhui Wei

    Department of Pathogen Biology and Microbiology, Zhejiang University School of Medicine, Hangzhou, China
    Competing interests
    The authors declare that no competing interests exist.
  3. Bachar H Hassan

    Stony Brook University, Stony Brook, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Jun Li

    College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou, China
    Competing interests
    The authors declare that no competing interests exist.
  5. Jiao-Yu Deng

    Key Laboratory of Agricultural and Environmental Microbiology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
    Competing interests
    The authors declare that no competing interests exist.
  6. Youjun Feng

    Department of Pathogen Biology and Microbiology, Zhejiang University School of Medicine, Hangzhou, China
    For correspondence
    fengyj@zju.edu.cn
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8083-0175

Funding

National Natural Science Foundation of China (31830001)

  • Youjun Feng

National Key R&D Program of China (2017YFD0500202)

  • Youjun Feng

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

Reviewing Editor

  1. Bavesh D Kana, University of the Witwatersrand, South Africa

Publication history

  1. Received: September 4, 2019
  2. Accepted: October 4, 2019
  3. Accepted Manuscript published: October 9, 2019 (version 1)
  4. Accepted Manuscript updated: October 10, 2019 (version 2)
  5. Accepted Manuscript updated: October 10, 2019 (version 3)
  6. Version of Record published: October 18, 2019 (version 4)

Copyright

© 2019, Gao 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

  • 1,812
    Page views
  • 351
    Downloads
  • 10
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

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. Rongsui Gao
  2. Wenhui Wei
  3. Bachar H Hassan
  4. Jun Li
  5. Jiao-Yu Deng
  6. Youjun Feng
(2019)
A single regulator NrtR controls bacterial NAD+ homeostasis via its acetylation
eLife 8:e51603.
https://doi.org/10.7554/eLife.51603

Further reading

    1. Microbiology and Infectious Disease
    Leire Aguinagalde Salazar, Maurits A den Boer ... Suzan HM Rooijakkers
    Research Article

    Streptococcus pneumoniae is the leading cause of community-acquired pneumonia and an important cause of childhood mortality. Despite the introduction of successful vaccines, the global spread of both non-vaccine serotypes and antibiotic-resistant strains reinforces the development of alternative therapies against this pathogen. One possible route is the development of monoclonal antibodies (mAbs) that induce killing of bacteria via the immune system. Here, we investigate whether mAbs can be used to induce killing of pneumococcal serotypes for which the current vaccines show unsuccessful protection. Our study demonstrates that when human mAbs against pneumococcal capsule polysaccharides (CPS) have a poor capacity to induce complement activation, a critical process for immune protection against pneumococci, their activity can be strongly improved by hexamerization-enhancing mutations. Our data indicate that anti-capsular antibodies may have a low capacity to form higher-order oligomers (IgG hexamers) that are needed to recruit complement component C1. Indeed, specific point mutations in the IgG-Fc domain that strengthen hexamerization strongly enhance C1 recruitment and downstream complement activation on encapsulated pneumococci. Specifically, hexamerization-enhancing mutations E430G or E345K in CPS6-IgG strongly potentiate complement activation on S. pneumoniae strains that express capsular serotype 6 (CPS6), and the highly invasive serotype 19A strain. Furthermore, these mutations improve complement activation via mAbs recognizing CPS3 and CPS8 strains. Importantly, hexamer-enhancing mutations enable mAbs to induce strong opsonophagocytic killing by human neutrophils. Finally, passive immunization with CPS6-IgG1-E345K protected mice from developing severe pneumonia. Altogether, this work provides an important proof of concept for future optimization of antibody therapies against encapsulated bacteria.

    1. Microbiology and Infectious Disease
    Ashelyn E Sidders, Katarzyna M Kedziora ... Brian P Conlon
    Research Article Updated

    Antibiotic tolerance and antibiotic resistance are the two major obstacles to the efficient and reliable treatment of bacterial infections. Identifying antibiotic adjuvants that sensitize resistant and tolerant bacteria to antibiotic killing may lead to the development of superior treatments with improved outcomes. Vancomycin, a lipid II inhibitor, is a frontline antibiotic for treating methicillin-resistant Staphylococcus aureus and other Gram-positive bacterial infections. However, vancomycin use has led to the increasing prevalence of bacterial strains with reduced susceptibility to vancomycin. Here, we show that unsaturated fatty acids act as potent vancomycin adjuvants to rapidly kill a range of Gram-positive bacteria, including vancomycin-tolerant and resistant populations. The synergistic bactericidal activity relies on the accumulation of membrane-bound cell wall intermediates that generate large fluid patches in the membrane leading to protein delocalization, aberrant septal formation, and loss of membrane integrity. Our findings provide a natural therapeutic option that enhances vancomycin activity against difficult-to-treat pathogens, and the underlying mechanism may be further exploited to develop antimicrobials that target recalcitrant infection.