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J Cell Biol. 2017 Sep 4;216(9):2891-2909. doi: 10.1083/jcb.201703103. Epub 2017 Jul 12.

Dynamics of in vivo ASC speck formation.

Author information

1
Directors' Research Unit, European Molecular Biology Laboratory, Heidelberg, Germany.
2
Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany.
3
Centre for Organismal Studies, Heidelberg University, Heidelberg, Germany.
4
Electron Microscopy Core Facility, European Molecular Biology Laboratory, Heidelberg, Germany.
5
Directors' Research Unit, European Molecular Biology Laboratory, Heidelberg, Germany mleptin@uni-koeln.de.
6
Institute of Genetics, University of Cologne, Cologne, Germany.
7
European Molecular Biology Organization, Heidelberg, Germany.

Abstract

Activated danger or pathogen sensors trigger assembly of the inflammasome adaptor ASC into specks, large signaling platforms considered hallmarks of inflammasome activation. Because a lack of in vivo tools has prevented the study of endogenous ASC dynamics, we generated a live ASC reporter through CRISPR/Cas9 tagging of the endogenous gene in zebrafish. We see strong ASC expression in the skin and other epithelia that act as barriers to insult. A toxic stimulus triggered speck formation and rapid pyroptosis in keratinocytes in vivo. Macrophages engulfed and digested that speck-containing, pyroptotic debris. A three-dimensional, ultrastructural reconstruction, based on correlative light and electron microscopy of the in vivo assembled specks revealed a compact network of highly intercrossed filaments, whereas pyrin domain (PYD) or caspase activation and recruitment domain alone formed filamentous aggregates. The effector caspase is recruited through PYD, whose overexpression induced pyroptosis but only after substantial delay. Therefore, formation of a single, compact speck and rapid cell-death induction in vivo requires a full-length ASC.

PMID:
28701426
PMCID:
PMC5584180
DOI:
10.1083/jcb.201703103
[Indexed for MEDLINE]
Free PMC Article

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