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Molecular basis of caspase-1 polymerization and its inhibition by a new capping mechanism

Nature Structural & Molecular Biology volume 23, pages 416425 (2016) | Download Citation

Abstract

Inflammasomes are cytosolic caspase-1-activation complexes that sense intrinsic and extrinsic danger signals, and trigger inflammatory responses and pyroptotic cell death. Homotypic interactions among Pyrin domains and caspase recruitment domains (CARDs) in inflammasome-complex components mediate oligomerization into filamentous assemblies. Several cytosolic proteins consisting of only interaction domains exert inhibitory effects on inflammasome assembly. In this study, we determined the structure of the human caspase-1 CARD domain (caspase-1CARD) filament by cryo-electron microscopy and investigated the biophysical properties of two caspase-1-like CARD-only proteins: human inhibitor of CARD (INCA or CARD17) and ICEBERG (CARD18). Our results reveal that INCA caps caspase-1 filaments, thereby exerting potent inhibition with low-nanomolar Ki on caspase-1CARD polymerization in vitro and inflammasome activation in cells. Whereas caspase-1CARD uses six complementary surfaces of three types for filament assembly, INCA is defective in two of the six interfaces and thus terminates the caspase-1 filament.

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Acknowledgements

The work was supported by US National Institutes of Health (NIH) grants to H.W. (Pioneer Award DP1-HD-087988), to H.L.P. (Pioneer Award DP1-GM-106409), and to Q.Y. (R00: 4R00AI108793-02). F.I.S. was supported by an Advanced Postdoc.Mobility Fellowship from the Swiss National Science Foundation. The cryo-EM facility was funded through the NIH grant AI100645, Center for HIV/AIDS Vaccine Immunology and Immunogen Design (CHAVI-ID). The experiments were performed in part at the Center for Nanoscale Systems at Harvard University, a member of the National Nanotechnology Infrastructure Network (NNIN), which is supported by the US National Science Foundation (NSF) under award no. ECS-0335765. We thank E. Egelman for generous guidance in methods of helical reconstruction.

Author information

Author notes

    • Qian Yin

    Present address: Department of Biological Science, Florida State University, Tallahassee, Florida, USA.

    • Alvin Lu
    •  & Yang Li

    These authors contributed equally to this work.

Affiliations

  1. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts, USA.

    • Alvin Lu
    • , Yang Li
    • , Qian Yin
    • , Tian-Min Fu
    • , Alexander B Tong
    •  & Hao Wu
  2. Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, Massachusetts, USA.

    • Alvin Lu
    • , Yang Li
    • , Qian Yin
    • , Tian-Min Fu
    • , Alexander B Tong
    •  & Hao Wu
  3. Whitehead Institute for Biomedical Research, Cambridge, Massachusetts, USA.

    • Florian I Schmidt
    •  & Hidde L Ploegh
  4. Center for Quantitative Biology, Peking-Tsinghua Joint Center for Life Sciences, Academy for Advanced Interdisciplinary Studies, State Key Laboratory for Artificial Microstructures and Mesoscopic Physics, School of Physics, Peking University, Beijing, China.

    • Shuobing Chen
    •  & Youdong Mao
  5. Department of Cancer Immunology and Virology, Intel Parallel Computing Center for Structural Biology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.

    • Shuobing Chen
    •  & Youdong Mao
  6. Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.

    • Hidde L Ploegh
  7. Department of Microbiology and Immunobiology, Harvard Medical School, Boston, Massachusetts, USA.

    • Youdong Mao

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Contributions

A.L., Y.L., F.I.S., Q.Y., S.C., T.-M.F., and Y.M. performed the experiments and analyzed the data. A.L., Q.Y., and T.-M.F. purified the recombinant proteins and performed biochemical experiments. S.C. and Y.M. collected the cryo-EM data, and Y.L. processed the data and completed the helical reconstruction. F.I.S. generated stable cell lines and performed cellular assays, and H.L.P. supervised the experiments. A.B.T. performed Rosetta refinement. A.L. and H.W. conceived the study and wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Hao Wu.

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    Supplementary Data Set 1

    Original gel images for Figure 2h

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DOI

https://doi.org/10.1038/nsmb.3199

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