Abstract

Phagocyte oxidase plays an essential role in the first line of host defense against pathogens. It oxidizes intracellular NADPH to reduce extracellular oxygen to produce superoxide anions that participate in pathogen killing. The resting phagocyte oxidase is a heterodimeric complex formed by two transmembrane proteins NOX2 and p22. Despite the physiological importance of this complex, its structure remains elusive. Here we reported the cryo-EM structure of the functional human NOX2-p22 complex in nanodisc in the resting state. NOX2 shows a canonical 6-TM architecture of NOX and p22 has four transmembrane helices. M3, M4, and M5 of NOX2 and M1 and M4 helices of p22 are involved in the hetero-dimer formation. DH domain of NOX2 in the resting state is not optimally docked onto the transmembrane domain (TMD), leading to inefficient electron transfer and NADPH binding. Structural analysis suggests that the cytosolic factors might activate the NOX2-p22 complex by stabilizing the dehydrogenase domain (DH) in a productive docked conformation.

Data availability

Cryo-EM maps and atomic coordinate of the NOX2-p22-7D5-TP1170 complex have been deposited in the EMDB and PDB under the ID codes EMDB: EMD-34389 and PDB: 8GZ3.

The following data sets were generated

Article and author information

Author details

  1. Rui Liu

    Institute of Molecular Medicine, Peking University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3758-6493
  2. Kangcheng Song

    Institute of Molecular Medicine, Peking University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7932-2202
  3. Jing-Xiang Wu

    Institute of Molecular Medicine, Peking University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9851-0065
  4. Xiao-Peng Geng

    Institute of Molecular Medicine, Peking University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  5. Liming Zheng

    College of Chemistry and Molecular Engineering, Peking University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  6. Xiaoyin Gao

    College of Chemistry and Molecular Engineering, Peking University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  7. Hailin Peng

    College of Chemistry and Molecular Engineering, Peking University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  8. Lei Chen

    Institute of Molecular Medicine, Peking University, Beijing, China
    For correspondence
    chenlei2016@pku.edu.cn
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7619-8311

Funding

National Key Research and Development Program of China (2022YFA1300088)

  • Lei Chen

National Natural Science Foundation of China (91957201,31870833,31821091,52021006)

  • Hailin Peng
  • Lei Chen

Beijing National Laboratory for Molecular Sciences (BNLMS-CXTD-202001)

  • Hailin Peng

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

Reviewing Editor

  1. Axel T Brunger, Stanford University School of Medicine, Howard Hughes Medical Institute, United States

Publication history

  1. Received: September 27, 2022
  2. Preprint posted: October 4, 2022 (view preprint)
  3. Accepted: November 21, 2022
  4. Accepted Manuscript published: November 22, 2022 (version 1)
  5. Version of Record published: November 30, 2022 (version 2)

Copyright

© 2022, Liu 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

  • 815
    Page views
  • 191
    Downloads
  • 4
    Citations

Article citation count generated by polling the highest count across the following sources: PubMed Central, Crossref, 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. Rui Liu
  2. Kangcheng Song
  3. Jing-Xiang Wu
  4. Xiao-Peng Geng
  5. Liming Zheng
  6. Xiaoyin Gao
  7. Hailin Peng
  8. Lei Chen
(2022)
Structure of human phagocyte NADPH oxidase in the resting state
eLife 11:e83743.
https://doi.org/10.7554/eLife.83743

Further reading

    1. Immunology and Inflammation
    Yu Li, Pablo Guaman Tipan ... Lauren IR Ehrlich
    Research Article

    Central tolerance ensures autoreactive T cells are eliminated or diverted to the regulatory T cell lineage, thus preventing autoimmunity. To undergo central tolerance, thymocytes must enter the medulla to test their TCRs for autoreactivity against the diverse self-antigens displayed by antigen presenting cells (APCs). While CCR7 is known to promote thymocyte medullary entry and negative selection, our previous studies implicate CCR4 in these processes, raising the question of whether CCR4 and CCR7 play distinct or redundant roles in central tolerance. Here, synchronized positive selection assays, 2-photon timelapse microscopy, and quantification of TCR-signaled apoptotic thymocytes, demonstrate that CCR4 and CCR7 promote medullary accumulation and central tolerance of distinct post-positive selection thymocyte subsets in mice. CCR4 is upregulated within hours of positive selection signaling and promotes medullary entry and clonal deletion of immature post-positive selection thymocytes. In contrast, CCR7 is expressed several days later and is required for medullary localization and negative selection of mature thymocytes. In addition, CCR4 and CCR7 differentially enforce self-tolerance, with CCR4 enforcing tolerance to self-antigens presented by activated APCs, which express CCR4 ligands. Our findings show that CCR7 expression is not synonymous with medullary localization and support a revised model of central tolerance in which CCR4 and CCR7 promote early and late stages of negative selection, respectively, via interactions with distinct APC subsets.

    1. Biochemistry and Chemical Biology
    2. Immunology and Inflammation
    Minato Hirano, Gaddiel Galarza-Muñoz ... Mariano A Garcia-Blanco
    Research Article

    Genes associated with increased susceptibility to multiple sclerosis (MS) have been identified, but their functions are incompletely understood. One of these genes codes for the RNA helicase DExD/H-Box Polypeptide 39B (DDX39B), which shows genetic and functional epistasis with interleukin-7 receptor-α gene (IL7R) in MS-risk. Based on evolutionary and functional arguments, we postulated that DDX39B enhances immune tolerance thereby decreasing MS risk. Consistent with such a role we show that DDX39B controls the expression of many MS susceptibility genes and important immune-related genes. Among these we identified Forkhead Box P3 (FOXP3), which codes for the master transcriptional factor in CD4+/CD25+ T regulatory cells. DDX39B knockdown led to loss of immune-regulatory and gain of immune-effector expression signatures. Splicing of FOXP3 introns, which belong to a previously unrecognized type of introns with C-rich polypyrimidine tracts, was exquisitely sensitive to DDX39B levels. Given the importance of FOXP3 in autoimmunity, this work cements DDX39B as an important guardian of immune tolerance.