NEK7 is an essential mediator of NLRP3 activation downstream of potassium efflux

Yuan He; Melody Y. Zeng; Dahai Yang; Benny Motro; Gabriel Núñez

doi:10.1038/nature16959

NEK7 is an essential mediator of NLRP3 activation downstream of potassium efflux

Yuan He, Melody Y. Zeng, Dahai Yang, Benny Motro, Gabriel Núñez

The Mina and Everard Goodman Faculty of Life Sciences

Research output: Contribution to journal › Article › peer-review

953 Scopus citations

Abstract

Inflammasomes are intracellular protein complexes that drive the activation of inflammatory caspases1. So far, four inflammasomes involving NLRP1, NLRP3, NLRC4 and AIM2 have been described that recruit the common adaptor protein ASC to activate caspase-1, leading to the secretion of mature IL-1â and IL-18 proteins2,3. The NLRP3 inflammasome has been implicated in the pathogenesis of several acquired inflammatory diseases4,5 as well as cryopyrinassociated periodic fever syndromes (CAPS) caused by inherited NLRP3 mutations6,7. Potassium efflux is a common step that is essential for NLRP3 inflammasome activation induced by many stimuli8,9. Despite extensive investigation, the molecular mechanism leading to NLRP3 activation in response to potassium efflux remains unknown. Here we report the identification of NEK7, a member of the family of mammalian NIMA-related kinases (NEK proteins)10, as an NLRP3-binding protein that acts downstream of potassium efflux to regulate NLRP3 oligomerization and activation. In the absence of NEK7, caspase-1 activation and IL-1â release were abrogated in response to signals that activate NLRP3, but not NLRC4 or AIM2 inflammasomes. NLRP3-activating stimuli promoted the NLRP3-NEK7 interaction in a process that was dependent on potassium efflux. NLRP3 associated with the catalytic domain of NEK7, but the catalytic activity of NEK7 was shown to be dispensable for activation of the NLRP3 inflammasome. Activated macrophages formed a high-molecular mass NLRP3-NEK7 complex, which, along with ASC oligomerization and ASC speck formation, was abrogated in the absence of NEK7. NEK7 was required for macrophages containing the CAPS-associated NLRP3(R258W) activating mutation to activate caspase-1. Mouse chimaeras reconstituted with wild-type, Nek7-/-or Nlrp3-/-haematopoietic cells showed that NEK7 was required for NLRP3 inflammasome activation in vivo. These studies demonstrate that NEK7 is an essential protein that acts downstream of potassium efflux to mediate NLRP3 inflammasome assembly and activation.

Original language	English
Pages (from-to)	354-357
Number of pages	4
Journal	Nature
Volume	530
Issue number	7590
DOIs	https://doi.org/10.1038/nature16959
State	Published - 18 Feb 2016

Bibliographical note

Publisher Copyright:
© 2016 Macmillan Publishers Limited.

Funding

Acknowledgements We thank R. Muñoz-Planillo for potassium efflux assays, S. Varadarajan for plasmid construction, L. Burmeister for animal husbandry, J. Whitfield for ELISA assays, and V. Basrur for mass spectrometry analysis. Y.H. was supported by NIH training grant T32HL007517. M.Y.Z. was supported by NIH training grants T32HL007517 and T32DK094775. D.Y. was supported by the State Scholarship Fund from China Scholarship Council (no. 201306740018). This work was supported by NIH grants R01AI063331 and R01DK091191 to G.N., research funds by the Israel Science Foundation (grant no. 768/11) to B.M., and funds to the Michigan Comprehensive Cancer Center Immunology Monitoring Core from the University of Michigan’s Cancer Center Support Grant. We thank R. Muñoz-Planillo for potassium efflux assays, S. Varadarajan for plasmid construction, L. Burmeister for animal husbandry, J. Whitfield for ELISA assays, and V. Basrur for mass spectrometry analysis. Y.H. was supported by NIH training grant T32HL007517. M.Y.Z. was supported by NIH training grants T32HL007517 and T32DK094775. D.Y. was supported by the State Scholarship Fund from China Scholarship Council (no. 201306740018). This work as supported by NIH grants R01AI063331 and R01DK091191 to G.N., research funds by the Israel Science Foundation (grant no. 768/11) to B.M., and funds to the Michigan Comprehensive Cancer Center Immunology Monitoring Core from the University of Michigan''s Cancer Center Support Grant.

Funders	Funder number
National Institutes of Health	T32DK094775, T32HL007517
National Institute of Diabetes and Digestive and Kidney Diseases	P30DK034933
University of Michigan
Israel Science Foundation	768/11
China Scholarship Council	201306740018, R01AI063331, R01DK091191

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title = "NEK7 is an essential mediator of NLRP3 activation downstream of potassium efflux",

abstract = "Inflammasomes are intracellular protein complexes that drive the activation of inflammatory caspases1. So far, four inflammasomes involving NLRP1, NLRP3, NLRC4 and AIM2 have been described that recruit the common adaptor protein ASC to activate caspase-1, leading to the secretion of mature IL-1{\^a} and IL-18 proteins2,3. The NLRP3 inflammasome has been implicated in the pathogenesis of several acquired inflammatory diseases4,5 as well as cryopyrinassociated periodic fever syndromes (CAPS) caused by inherited NLRP3 mutations6,7. Potassium efflux is a common step that is essential for NLRP3 inflammasome activation induced by many stimuli8,9. Despite extensive investigation, the molecular mechanism leading to NLRP3 activation in response to potassium efflux remains unknown. Here we report the identification of NEK7, a member of the family of mammalian NIMA-related kinases (NEK proteins)10, as an NLRP3-binding protein that acts downstream of potassium efflux to regulate NLRP3 oligomerization and activation. In the absence of NEK7, caspase-1 activation and IL-1{\^a} release were abrogated in response to signals that activate NLRP3, but not NLRC4 or AIM2 inflammasomes. NLRP3-activating stimuli promoted the NLRP3-NEK7 interaction in a process that was dependent on potassium efflux. NLRP3 associated with the catalytic domain of NEK7, but the catalytic activity of NEK7 was shown to be dispensable for activation of the NLRP3 inflammasome. Activated macrophages formed a high-molecular mass NLRP3-NEK7 complex, which, along with ASC oligomerization and ASC speck formation, was abrogated in the absence of NEK7. NEK7 was required for macrophages containing the CAPS-associated NLRP3(R258W) activating mutation to activate caspase-1. Mouse chimaeras reconstituted with wild-type, Nek7-/-or Nlrp3-/-haematopoietic cells showed that NEK7 was required for NLRP3 inflammasome activation in vivo. These studies demonstrate that NEK7 is an essential protein that acts downstream of potassium efflux to mediate NLRP3 inflammasome assembly and activation.",

author = "Yuan He and Zeng, {Melody Y.} and Dahai Yang and Benny Motro and Gabriel N{\'u}{\~n}ez",

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AU - He, Yuan

AU - Zeng, Melody Y.

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AU - Motro, Benny

AU - Núñez, Gabriel

PY - 2016/2/18

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N2 - Inflammasomes are intracellular protein complexes that drive the activation of inflammatory caspases1. So far, four inflammasomes involving NLRP1, NLRP3, NLRC4 and AIM2 have been described that recruit the common adaptor protein ASC to activate caspase-1, leading to the secretion of mature IL-1â and IL-18 proteins2,3. The NLRP3 inflammasome has been implicated in the pathogenesis of several acquired inflammatory diseases4,5 as well as cryopyrinassociated periodic fever syndromes (CAPS) caused by inherited NLRP3 mutations6,7. Potassium efflux is a common step that is essential for NLRP3 inflammasome activation induced by many stimuli8,9. Despite extensive investigation, the molecular mechanism leading to NLRP3 activation in response to potassium efflux remains unknown. Here we report the identification of NEK7, a member of the family of mammalian NIMA-related kinases (NEK proteins)10, as an NLRP3-binding protein that acts downstream of potassium efflux to regulate NLRP3 oligomerization and activation. In the absence of NEK7, caspase-1 activation and IL-1â release were abrogated in response to signals that activate NLRP3, but not NLRC4 or AIM2 inflammasomes. NLRP3-activating stimuli promoted the NLRP3-NEK7 interaction in a process that was dependent on potassium efflux. NLRP3 associated with the catalytic domain of NEK7, but the catalytic activity of NEK7 was shown to be dispensable for activation of the NLRP3 inflammasome. Activated macrophages formed a high-molecular mass NLRP3-NEK7 complex, which, along with ASC oligomerization and ASC speck formation, was abrogated in the absence of NEK7. NEK7 was required for macrophages containing the CAPS-associated NLRP3(R258W) activating mutation to activate caspase-1. Mouse chimaeras reconstituted with wild-type, Nek7-/-or Nlrp3-/-haematopoietic cells showed that NEK7 was required for NLRP3 inflammasome activation in vivo. These studies demonstrate that NEK7 is an essential protein that acts downstream of potassium efflux to mediate NLRP3 inflammasome assembly and activation.

AB - Inflammasomes are intracellular protein complexes that drive the activation of inflammatory caspases1. So far, four inflammasomes involving NLRP1, NLRP3, NLRC4 and AIM2 have been described that recruit the common adaptor protein ASC to activate caspase-1, leading to the secretion of mature IL-1â and IL-18 proteins2,3. The NLRP3 inflammasome has been implicated in the pathogenesis of several acquired inflammatory diseases4,5 as well as cryopyrinassociated periodic fever syndromes (CAPS) caused by inherited NLRP3 mutations6,7. Potassium efflux is a common step that is essential for NLRP3 inflammasome activation induced by many stimuli8,9. Despite extensive investigation, the molecular mechanism leading to NLRP3 activation in response to potassium efflux remains unknown. Here we report the identification of NEK7, a member of the family of mammalian NIMA-related kinases (NEK proteins)10, as an NLRP3-binding protein that acts downstream of potassium efflux to regulate NLRP3 oligomerization and activation. In the absence of NEK7, caspase-1 activation and IL-1â release were abrogated in response to signals that activate NLRP3, but not NLRC4 or AIM2 inflammasomes. NLRP3-activating stimuli promoted the NLRP3-NEK7 interaction in a process that was dependent on potassium efflux. NLRP3 associated with the catalytic domain of NEK7, but the catalytic activity of NEK7 was shown to be dispensable for activation of the NLRP3 inflammasome. Activated macrophages formed a high-molecular mass NLRP3-NEK7 complex, which, along with ASC oligomerization and ASC speck formation, was abrogated in the absence of NEK7. NEK7 was required for macrophages containing the CAPS-associated NLRP3(R258W) activating mutation to activate caspase-1. Mouse chimaeras reconstituted with wild-type, Nek7-/-or Nlrp3-/-haematopoietic cells showed that NEK7 was required for NLRP3 inflammasome activation in vivo. These studies demonstrate that NEK7 is an essential protein that acts downstream of potassium efflux to mediate NLRP3 inflammasome assembly and activation.

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NEK7 is an essential mediator of NLRP3 activation downstream of potassium efflux

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