Structural mechanisms of calmodulin activation of Shigella effector OspC3 to ADP-riboxanate caspase-4/11 and block pyroptosis

  • Nat Struct Mol Biol. 2023 Jan 9. doi: 10.1038/s41594-022-00888-3.
Yanjie Hou  #  1 ,  Huan Zeng  #  1  2  3 ,  Zilin Li  3  4 ,  Na Feng  1 ,  Fanyi Meng  1  2 ,  Yue Xu  3  5 ,  Lin Li  3 ,  Feng Shao  6  7  8  9 ,  Jingjin Ding  10  11  12
Affiliations
  • 1. National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.
  • 2. University of Chinese Academy of Sciences, Beijing, China.
  • 3. National Institute of Biological Sciences, Beijing, Beijing, China.
  • 4. Research Unit of Pyroptosis and Immunity, Chinese Academy of Medical Sciences and National Institute of Biological Sciences, Beijing, China.
  • 5. Department of Pathophysiology, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
  • 6. National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China. [email protected].
  • 7. National Institute of Biological Sciences, Beijing, Beijing, China. [email protected].
  • 8. Research Unit of Pyroptosis and Immunity, Chinese Academy of Medical Sciences and National Institute of Biological Sciences, Beijing, China. [email protected].
  • 9. Tsinghua Institute of Multidisciplinary Biomedical Research, Tsinghua University, Beijing, China. [email protected].
  • 10. National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China. [email protected].
  • 11. University of Chinese Academy of Sciences, Beijing, China. [email protected].
  • 12. National Institute of Biological Sciences, Beijing, Beijing, China. [email protected].
  • # Contributed equally.
PMID: 36624349 DOI: 10.1038/s41594-022-00888-3
Abstract

The caspase-4/11-GSDMD Pyroptosis axis recognizes cytosolic lipopolysaccharide for Antibacterial defenses. Shigella flexneri employs an OspC3 effector to block Pyroptosis by catalyzing NAD+-dependent arginine ADP-riboxanation of caspase-4/11. Here, we identify CA2+-free Calmodulin (CaM) that binds and stimulates OspC3 ADP-riboxanase activity. Crystal structures of OspC3-CaM and OspC3-caspase-4 binary complexes reveal unique CaM binding to an OspC3 N-terminal domain featuring an ADP-ribosyltransferase-like fold and specific recognition of caspase-4 by an OspC3 ankryin repeat domain, respectively. CaM-OspC3-caspase-4 ternary complex structures show that NAD+ binding reorganizes the catalytic pocket, in which D231 and D177 activate the substrate arginine for initial ADP-ribosylation and ribosyl 2'-OH in the ADP-ribosylated arginine, respectively, for subsequent deamination. We also determine structures of unmodified and OspC3-ADP-riboxanated caspase-4. Mechanisms derived from this series of structures covering the entire process of OspC3 action are supported by biochemical analyses in vitro and functional validation in S. flexneri-infected mice.

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