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Cell Chem Biol. 2019 Jun 20;26(6):863-877.e7. doi: 10.1016/j.chembiol.2019.03.010. Epub 2019 Apr 25.

Direct Activation of Human MLKL by a Select Repertoire of Inositol Phosphate Metabolites.

Author information

1
Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA; Department of Chemical Biology and Therapeutics, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
2
Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA.
3
Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
4
Department of Immunology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
5
Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
6
Molecular Interaction Analysis Shared Resource, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
7
Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA. Electronic address: carette@stanford.edu.
8
Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA; Department of Chemical Biology and Therapeutics, St. Jude Children's Research Hospital, Memphis, TN 38105, USA. Electronic address: tudor.moldoveanu@stjude.org.

Abstract

Necroptosis is an inflammatory form of programmed cell death executed through plasma membrane rupture by the pseudokinase mixed lineage kinase domain-like (MLKL). We previously showed that MLKL activation requires metabolites of the inositol phosphate (IP) pathway. Here we reveal that I(1,3,4,6)P4, I(1,3,4,5,6)P5, and IP6 promote membrane permeabilization by MLKL through directly binding the N-terminal executioner domain (NED) and dissociating its auto-inhibitory region. We show that IP6 and inositol pentakisphosphate 2-kinase (IPPK) are required for necroptosis as IPPK deletion ablated IP6 production and inhibited necroptosis. The NED auto-inhibitory region is more extensive than originally described and single amino acid substitutions along this region induce spontaneous necroptosis by MLKL. Activating IPs bind three sites with affinity of 100-600 μM to destabilize contacts between the auto-inhibitory region and NED, thereby promoting MLKL activation. We therefore uncover MLKL's activating switch in NED triggered by a select repertoire of IP metabolites.

KEYWORDS:

CRISPR Cas9 gene editing; MLKL; NMR structure determination; cell death; inositol phosphate; inositol phosphate kinase; membrane permeabilization; metabolite; necroptosis; plasma membrane rupture

PMID:
31031142
PMCID:
PMC6588482
[Available on 2020-06-20]
DOI:
10.1016/j.chembiol.2019.03.010

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