Abstract
Inflammasomes are multiprotein complexes, which assembly results in caspase-1 activation and subsequent IL-1β and IL-18 activation and secretion. In a cell-free system, based on cytosols of normally growing cells, the disruption of the cell membrane spontaneously activates the inflammasome. Studying the activation of the inflammasome in cytosolic extracts provides multiple advantages, as it is synchronized, rapid, strong, and mostly plasma membrane-free. This protocol covers the methods required to prepare cell lysates and study inflammasome activation using different read-outs. General considerations are provided that may help in the design of modified methods. This assay can be useful to study potential inflammasome interactors and the signaling pathways involved in its activation.
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Acknowledgments
F.M. is supported by a grant from the European Research Council (starting grant 281996), a Human Frontier Science Program career development award (CDA00059/2011), and a grant from the Swiss National Science Foundation (31003A-130476).
Y.J. is supported by a grant from the Foundation for the Development of Internal Medicine in Europe (FDIME), a grant from the Société Nationale Française de Médecine Interne (SNFMI)/Genzyme, a grant from Groupama foundation, and a “poste d’accueil” at INSERM.
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Jamilloux, Y., Martinon, F. (2016). Cell-Free Assay for Inflammasome Activation. In: Di Virgilio, F., Pelegrín, P. (eds) NLR Proteins. Methods in Molecular Biology, vol 1417. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3566-6_14
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DOI: https://doi.org/10.1007/978-1-4939-3566-6_14
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