Structural basis of phosphatidylcholine recognition by the C2-domain of cytosolic phospholipase A2α

  1. Yoshinori Hirano
  2. Yong-Guang Gao
  3. Daniel J Stephenson
  4. Ngoc T Vu
  5. Lucy Malinina
  6. Dhirendra K Simanshu
  7. Charles E Chalfant
  8. Dinshaw J Patel
  9. Rhoderick E Brown  Is a corresponding author
  1. Memorial Sloan Kettering Cancer Center, United States
  2. University of Minnesota, United States
  3. University of South Florida, United States
  4. Virginia Commonwealth University, United States

Abstract

Ca2+-stimulated translocation of cytosolic phospholipase A2α (cPLA2α) to the Golgi induces arachidonic acid production, the rate-limiting step in pro-inflammatory eicosanoid synthesis. Structural insights into the cPLA2α preference for phosphatidylcholine (PC)-enriched membranes have remained elusive. Here, we report cPLA2α C2-domain structure (2.2Å resolution) containing bound 1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC) and Ca2+ ions. Two Ca2+ are complexed at locations previously reported for lipid-free C2-domain. One of these Ca2+ along with a third Ca2+ bridge the C2-domain to the DHPC phosphate group, which also interacts with Asn65. Tyr96 plays a key role in lipid headgroup recognition via cation-π interaction with the PC trimethylammonium group. Mutagenesis analyses confirm Tyr96 and Asn65 function in PC binding selectivity by C2-domain and regulation of cPLA2α activity. The differing DHPC-binding mode of cPLA2α C2-domain, compared to phosphatidylserine or phosphatidylinositol 4,5-bisphosphate binding by other C2-domains, expands and deepens knowledge of lipid-binding mechanisms mediated by C2-domains.

Data availability

Diffraction data have been deposited in PDB under the accession code 6IEJ

The following data sets were generated

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Author details

  1. Yoshinori Hirano

    Structural Biology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9888-1616
  2. Yong-Guang Gao

    Hormel Institute, University of Minnesota, Austin, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9359-4252
  3. Daniel J Stephenson

    Department of Cell Biology, Microbiology, and Molecular Biology, University of South Florida, Tampa, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5698-3400
  4. Ngoc T Vu

    Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, Richmond, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Lucy Malinina

    Hormel Institute, University of Minnesota, Austin, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7973-1831
  6. Dhirendra K Simanshu

    Structural Biology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9717-4618
  7. Charles E Chalfant

    Department of Cell Biology, Microbiology, and Molecular Biology, University of South Florida, Tampa, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5844-5235
  8. Dinshaw J Patel

    Structural Biology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
  9. Rhoderick E Brown

    Hormel Institute, University of Minnesota, Austin, United States
    For correspondence
    reb@umn.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7337-3604

Funding

National Institutes of Health (HL125353)

  • Rhoderick E Brown

Ministry of Education, Culture, Sports, Science, and Technology

  • Yoshinori Hirano

U.S. Department of Veterans Affairs (I BX001792)

  • Charles E Chalfant

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

Reviewing Editor

  1. Arun Radhakrishnan, University of Texas Southwestern Medical Center, United States

Publication history

  1. Received: December 28, 2018
  2. Accepted: May 3, 2019
  3. Accepted Manuscript published: May 3, 2019 (version 1)
  4. Version of Record published: June 5, 2019 (version 2)
  5. Version of Record updated: November 29, 2021 (version 3)

Copyright

© 2019, Hirano 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. Yoshinori Hirano
  2. Yong-Guang Gao
  3. Daniel J Stephenson
  4. Ngoc T Vu
  5. Lucy Malinina
  6. Dhirendra K Simanshu
  7. Charles E Chalfant
  8. Dinshaw J Patel
  9. Rhoderick E Brown
(2019)
Structural basis of phosphatidylcholine recognition by the C2-domain of cytosolic phospholipase A2α
eLife 8:e44760.
https://doi.org/10.7554/eLife.44760

Further reading

    1. Structural Biology and Molecular Biophysics
    Huan-Huan Hu et al.
    Research Article Updated

    Phosphoribosyl pyrophosphate (PRPP) is a key intermediate in the biosynthesis of purine and pyrimidine nucleotides, histidine, tryptophan, and cofactors NAD and NADP. Abnormal regulation of PRPP synthase (PRPS) is associated with human disorders, including Arts syndrome, retinal dystrophy, and gouty arthritis. Recent studies have demonstrated that PRPS can form filamentous cytoophidia in eukaryotes. Here, we show that PRPS forms cytoophidia in prokaryotes both in vitro and in vivo. Moreover, we solve two distinct filament structures of E. coli PRPS at near-atomic resolution using Cryo-EM. The formation of the two types of filaments is controlled by the binding of different ligands. One filament type is resistant to allosteric inhibition. The structural comparison reveals conformational changes of a regulatory flexible loop, which may regulate the binding of the allosteric inhibitor and the substrate ATP. A noncanonical allosteric AMP/ADP binding site is identified to stabilize the conformation of the regulatory flexible loop. Our findings not only explore a new mechanism of PRPS regulation with structural basis, but also propose an additional layer of cell metabolism through PRPS filamentation.