A missense mutation in the MLKL brace region promotes lethal neonatal inflammation and hematopoietic dysfunction

Hildebrand, Joanne M.; Kauppi, Maria; Majewski, Ian J.; Liu, Zikou; Cox, Allison; Miyake, Shingo; Petrie, Emma J.; Silk, Michael; Li, Zhixiu; Tanzer, Maria C.; Brumatti, Gabriela; Young, Samuel N.; Hall, Cathrine; Garnish, Sarah E; Corbin, Jason; Stutz, Michael D.; Rago, Ladina Di; Gangatirkar, Pradnya; Josefsson, Emma C.; Rigbye, Kristin; Anderton, Holly; Rickard, James A; Tripaydonis, Anne; Sheridan, Julie; Scerri, Thomas; Jackson, Victoria E.; Czabotar, P.E.; Zhang, Jianguo; Varghese, Leila N.; Allison, Cody; Pellegrini, Marc; Tannahill, Gillian M.; Hatchell, Esme C.; Willson, Tracy A.; Stockwell, Dina; Graaf, Carolyn A. de; Collinge, J; Hilton, Adrienne; Silke, Natasha; Spall, Sukhdeep K; Chau, Diep; Athanasopoulos, Vicki; Metcalf, Donald; Laxer, Ronald M.; Bassuk, Alexander G.; Darbro, Benjamin W.; Singh, Maria A. Fiatarone; Vlahovich, Nicole; Hughes, David; Kozlovskaia, Maria; Ascher, David B.; Warnatz, Klaus; Venhoff, Nils; Thiel, Jens; Biben, Christine; Blum, Stefan; Reveille, John D.; Hildebrand, Michael S.; Vinuesa, Carola G.; McCombe, Pamela A.; Brown, Matthew A.; Kile, Benjamin T.; McLean, Catriona; Bahlo, Melanie; Masters, Seth L.; Nakano, Hiroyasu; Ferguson, Polly J.; Murphy, James M.; Alexander, Warren S.; Silke, John

doi:10.1038/s41467-020-16819-z

Cited by 82 publications

(105 citation statements)

References 83 publications

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“…These single amino acid changes can readily disrupt the intricate network of intramolecular interactions, affecting how a protein folds, its stability, dynamics, and ultimately protein function. Beyond phenotypic outcomes, 1–22 it also has direct implications for their experimental study, protein engineering, 23,24 drug design, 25–30 and use in industrial processes 31 …”

Section: Introductionmentioning

confidence: 99%

DynaMut2: Assessing changes in stability and flexibility upon single and multiple point missense mutations

2020

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Predicting the effect of missense variations on protein stability and dynamics is important for understanding their role in diseases, and the link between protein structure and function. Approaches to estimate these changes have been proposed, but most only consider single‐point missense variants and a static state of the protein, with those that incorporate dynamics are computationally expensive. Here we present DynaMut2, a web server that combines Normal Mode Analysis (NMA) methods to capture protein motion and our graph‐based signatures to represent the wildtype environment to investigate the effects of single and multiple point mutations on protein stability and dynamics. DynaMut2 was able to accurately predict the effects of missense mutations on protein stability, achieving Pearson's correlation of up to 0.72 (RMSE: 1.02 kcal/mol) on a single point and 0.64 (RMSE: 1.80 kcal/mol) on multiple‐point missense mutations across 10‐fold cross‐validation and independent blind tests. For single‐point mutations, DynaMut2 achieved comparable performance with other methods when predicting variations in Gibbs Free Energy (ΔΔG) and in melting temperature (ΔTm). We anticipate our tool to be a valuable suite for the study of protein flexibility analysis and the study of the role of variants in disease. DynaMut2 is freely available as a web server and API at http://biosig.unimelb.edu.au/dynamut2.

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Section: Introductionmentioning

confidence: 99%

DynaMut2: Assessing changes in stability and flexibility upon single and multiple point missense mutations

2020

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“…Phosphorylated MLKL forms oligomers that translocate to intracellular membranes and the plasma membrane, inducing membrane rupture, and the release of DAMPs 29 . Necroptosis has been implicated in mediating various diseases, including systemic inflammation, ischemic reperfusion injury and neurodegeneration 20 , 30 , 31 .…”

Section: Introductionmentioning

confidence: 99%

The necroptotic cell death pathway operates in megakaryocytes, but not in platelet synthesis

et al. 2021

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Necroptosis is a pro-inflammatory cell death program executed by the terminal effector, mixed lineage kinase domain-like (MLKL). Previous studies suggested a role for the necroptotic machinery in platelets, where loss of MLKL or its upstream regulator, RIPK3 kinase, impacted thrombosis and haemostasis. However, it remains unknown whether necroptosis operates within megakaryocytes, the progenitors of platelets, and whether necroptotic cell death might contribute to or diminish platelet production. Here, we demonstrate that megakaryocytes possess a functional necroptosis signalling cascade. Necroptosis activation leads to phosphorylation of MLKL, loss of viability and cell swelling. Analyses at steady state and post antibody-mediated thrombocytopenia revealed that platelet production was normal in the absence of MLKL, however, platelet activation and haemostasis were impaired with prolonged tail re-bleeding times. We conclude that MLKL plays a role in regulating platelet function and haemostasis and that necroptosis signalling in megakaryocytes is dispensable for platelet production.

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“…ecroptosis is a caspase-independent, lytic form of programmed cell death [1][2][3][4][5][6][7] , which ancestrally has been implicated in host defense 3,[8][9][10][11] . Necroptosis is also thought to participate in multiple human pathologies including ischemia-reperfusion injuries 12,13 , degenerative diseases 14 and inflammatory diseases 12,[15][16][17][18][19][20] . Necroptosis is known to be induced downstream of death, Toll-like and interferon receptor activation by their cognate ligands.…”

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

et al. 2020

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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-actindependent 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.

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A missense mutation in the MLKL brace region promotes lethal neonatal inflammation and hematopoietic dysfunction

Cited by 82 publications

References 83 publications

DynaMut2: Assessing changes in stability and flexibility upon single and multiple point missense mutations

DynaMut2: Assessing changes in stability and flexibility upon single and multiple point missense mutations

The necroptotic cell death pathway operates in megakaryocytes, but not in platelet synthesis

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

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