MLKL trafficking and accumulation at the plasma membrane control the kinetics and threshold for necroptosis

Andre L Samson1,2, Ying Zhang3,4, Niall D Geoghegan3,4, Xavier J Gavin3, Katherine A Davies3,4, Michael J Mlodzianoski3,4, Lachlan W Whitehead3,4, Daniel Frank3,4, Sarah E Garnish3,4, Cheree Fitzgibbon3, Anne Hempel3, Samuel N Young3, Annette V Jacobsen3,4, Wayne Cawthorne3,4, Emma J Petrie3,4, Maree C Faux3,4, Kristy Shield-Artin3,4, Najoua Lalaoui3,4, Joanne M Hildebrand3,4, John Silke3,4, Kelly L Rogers3,4, Guillaume Lessene3,4,5, Edwin D Hawkins6,7, James M Murphy8,9

  1. The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia. samson.a@wehi.edu.au.
  2. Department of Medical Biology, The University of Melbourne, Parkville, VIC, Australia. samson.a@wehi.edu.au.
  3. The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia.
  4. Department of Medical Biology, The University of Melbourne, Parkville, VIC, Australia.
  5. Department of Pharmacology and Therapeutics, The University of Melbourne, Parkville, VIC, Australia.
  6. The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia. hawkins.e@wehi.edu.au.
  7. Department of Medical Biology, The University of Melbourne, Parkville, VIC, Australia. hawkins.e@wehi.edu.au.
  8. The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia. jamesm@wehi.edu.au.
  9. Department of Medical Biology, The University of Melbourne, Parkville, VIC, Australia. jamesm@wehi.edu.au.

Abstract

Mixed lineage kinase domain-like (MLKL) is the terminal protein in the pro-inflammatory necroptotic cell death program. RIPK3-mediated phosphorylation is thought to initiate MLKL oligomerization, membrane translocation and membrane disruption, although the precise choreography of events is incompletely understood. Here, we use single-cell imaging approaches to map the chronology of endogenous human MLKL activation during necroptosis. During the effector phase of necroptosis, we observe that phosphorylated MLKL assembles into higher order species on presumed cytoplasmic necrosomes. Subsequently, MLKL co-traffics with tight junction proteins to the cell periphery via Golgi-microtubule-actin-dependent mechanisms. MLKL and tight junction proteins then steadily co-accumulate at the plasma membrane as heterogeneous micron-sized hotspots. Our studies identify MLKL trafficking and plasma membrane accumulation as crucial necroptosis checkpoints. Furthermore, the accumulation of phosphorylated MLKL at intercellular junctions accelerates necroptosis between neighbouring cells, which may be relevant to inflammatory bowel disease and other necroptosis-mediated enteropathies.

Presented By Andre Samson

Cell & Developmental Bio, BiochemistryNicholas Liao2020-06 (Jun)