Journal article
A FRET biosensor for necroptosis uncovers two different modes of the release of DAMPs
S Murai, Y Yamaguchi, Y Shirasaki, M Yamagishi, R Shindo, JM Hildebrand, R Miura, O Nakabayashi, M Totsuka, T Tomida, S Adachi-Akahane, S Uemura, J Silke, H Yagita, M Miura, H Nakano
Nature Communications | NATURE PUBLISHING GROUP | Published : 2018
Abstract
Necroptosis is a regulated form of necrosis that depends on receptor-interacting protein kinase (RIPK)3 and mixed lineage kinase domain-like (MLKL). While danger-associated molecular pattern (DAMP)s are involved in various pathological conditions and released from dead cells, the underlying mechanisms are not fully understood. Here we develop a fluorescence resonance energy transfer (FRET) biosensor, termed SMART (a sensor for MLKL activation by RIPK3 based on FRET). SMART is composed of a fragment of MLKL and monitors necroptosis, but not apoptosis or necrosis. Mechanistically, SMART monitors plasma membrane translocation of oligomerized MLKL, which is induced by RIPK3 or mutational activat..
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Awarded by Takeda Science Foundation
Funding Acknowledgements
We thank M. Pasparakis for Mlkl-/- mice; J. Murphy for pF-TRE3G-PGK-puro-WT MLKL, MLKL L280P, MLKL Q343A, and anti-MLKL antibody; H. Niwa for pPB-hCMV*1-IRES-mCherry and pCAGGS-PBase; H. Miyoshi for pCAG-HIVgp and pCMV-VSV-G-RSV-Rev; K. Kabashima for HaCaT cells; K. Kawakami for transposon-based expression vector system; and M. Matsuda for a backbone vector (3536NES) for a FRET biosensor, S. Sugano for pEF321-T, respectively. We also thank K. Ogata and S. Yamazaki, and S. Komazawa-Sakon for helpful discussion and technical assistance, respectively. This work was supported in part by Grants-in-Aid for Scientific Research (B) 17H04069 (to H.N.), Scientific Research (C) 16K01378 (to S. M.) and 16K07304 (to T.T.), Scientific Research (S) 16H06385 (to M.M.), and PRESTO JP17940748 (to Y.S.), and Challenging Exploratory Research 17K19533 (to H.N.) from Japan Society for the Promotion of Science (JSPS), Scientific Research on Innovative Areas 26110003 (to H.N.), 26110005 (to Y.Y.), 15H01366 (to Y.S.), 17H05496 (to Y.S.), 17H06017 (to T.T.), and the Japan Agency for Medical Research and Development (AMED) through AMED-CREST with grant number JP17gm0610004 (to M.M.), partly through AMED-Project for Elucidating and Controlling Mechanisms of Aging and Longevity with grant number JP17gm5010001 (to M.M.), and Private University Research Branding project (to H.N.) from the MEXT (Ministry of Education, Culture, Sports, Science and Technology), Japan, research grants from Development Research, NAKATANI Foundation (to H.N.), the Naito Science Foundation (to H.N.), the Uehara Science Foundation (to H.N.), and the Takeda Science Foundation (to H.N.). J.M.H. and J.S. are supported by Australian NHMRC grants GNT1142669, GNT1107149, and GNT1105023.