Ribosome stalling is a signal for metabolic regulation by the ribotoxic stress response

Snieckute, Goda; Genzor, Aitana Victoria; Vind, Anna; Ryder, Laura; Stoneley, Mark; Chamois, Sébastien; Dreos, René; Nordgaard, Cathrine; Sass, Frederike; Blasius, Melanie; López, Aida Rodríguez; Brynjólfsdóttir, Sólveig Hlín; Andersen, Kasper Langebjerg; Willis, Anne E.; Frankel, Lisa B.; Poulsen, Steen Seier; Gatfield, David; Clemmensen, Christoffer; Bekker‐Jensen, Simon

doi:10.1016/j.cmet.2022.10.011

Cited by 45 publications

(36 citation statements)

References 46 publications

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“…This double mutant was also defective for the characteristic anisomycin-induced gel mobility shift ( Figure 1 b), which is indicative of ZAKα autophosphorylation [ 20 ]. We recently showed that cells incubated in the starvation medium Earle’s Balanced Salt Solution (EBSS) also activate p38 and JNK in a ZAKα-dependent fashion [ 18 ]. Similar to the anisomycin treatment above, this response was also dependent on the integrity of the S domain peptide motifs within ZAKα ( Figure 1 c).…”

Section: Resultsmentioning

confidence: 99%

“…Furthermore, some chemotherapeutics (e.g., doxorubicin) and UV-irradiation are recognized as ribotoxic agents [ 13 , 16 ], the latter due to crosslinking of nucleotides in mRNA templates [ 17 ]. Recently, translational down-regulation following nutrient depletion and amino acid starvation were also demonstrated to be potent ZAKα-activating stimuli [ 18 ]. ZAKα homologues are absent from unicellular organisms, such as the yeast Saccharomyces cerevisiae , but clearly present in the genomes of vertebrate species, including fish and mammals.…”

Section: Introductionmentioning

confidence: 99%

“…Some knowledge about the mechanism(s) underlying ZAKα activation has been gleaned in recent years, such as the ability of the kinase to bind directly to ribosomes through dedicated sensor domains [ 20 ] and become activated by collided [ 17 ] as well as stalled ribosomes [ 18 ]. However, the ability of the kinase to discriminate between ribosomal states remains unexplained, in part because stable ribosome-ZAKα complexes have escaped structural elucidation by cryo-EM.…”

Section: Introductionmentioning

confidence: 99%

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Computational and Functional Analysis of Structural Features in the ZAKα Kinase

Johansen

Snieckute

Vind

et al. 2023

Cells

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The kinase ZAKα acts as the proximal sensor of translational impairment and ribotoxic stress, which results in the activation of the MAP kinases p38 and JNK. Despite recent insights into the functions and binding partners of individual protein domains in ZAKα, the mechanisms by which ZAKα binds ribosomes and becomes activated have remained elusive. Here, we highlight a short, thrice-repeated, and positively charged peptide motif as critical for the ribotoxic stress-sensing function of the Sensor (S) domain of ZAKα. We use this insight to demonstrate that the mutation of the SAM domain uncouples ZAKα activity from ribosome binding. Finally, we use 3D structural comparison to identify and functionally characterize an additional folded domain in ZAKα with structural homology to YEATS domains. These insights allow us to formulate a model for ribosome-templated ZAKα activation based on the re-organization of interactions between modular protein domains. In sum, our work both advances our understanding of the protein domains and 3D architecture of the ZAKα kinase and furthers our understanding of how the ribotoxic stress response is activated.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Computational and Functional Analysis of Structural Features in the ZAKα Kinase

Johansen

Snieckute

Vind

et al. 2023

Cells

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“…As antibody-DT conjugates are currently used to treat certain cancer types (Frankel et al, 2002), the contribution of RSR and the NLRP1 inflammasome in their therapeutic effect warrants further investigation. In addition, given that ZAKα-driven RSR is conserved in rodents (Snieckute et al, 2022; Silva et al, 2022), the immunological consequence of DTA as an in vivo cell ablation tool should also be more carefully examined.…”

Section: Discussionmentioning

confidence: 99%

Corynebacterium diphtheriaecauses keratinocyte-intrinsic ribotoxic stress and NLRP1 inflammasome activation in a model of cutaneous diphtheria

Robinson

Toh

Firdaus

et al. 2023

Preprint

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NLRP1 is an innate immune sensor protein that activates inflammasome-driven pyroptotic cell death. Recent work demonstrates that human NLRP1 has evolved to sense viral infections. Whether and how human NLRP1 responds to other infectious agents is unclear. Here, and in a companion manuscript, we report that human NLRP1, as an integral component of the ribotoxic stress response (RSR), is activated by bacterial exotoxins that target human ribosome elongation factors EEF1 and EEF2, including Diphtheria Toxin (DT) from Corynebacterium diphtheriae, exotoxin A from Pseudomonas aeruginosa and sidI from Legionella pneumophila. In human keratinocytes, DT activates RSR kinases ZAKalpha, p38 and JNKs, upregulates a set of signature RSR transcripts and triggers rapid NLRP1-dependent pyroptosis. Mechanistically, these processes require 1) DtxR-mediated de-repression of DT production in the bacteria, as well as 2) diphthamide synthesis and 3) ZAKalpha/p38-driven NLRP1 phosphorylation in the host. In 3D human skin cultures, Corynebacterium diphtheriae infection disrupts barrier function and induces IL-1 driven inflammation. Pharmacologic inhibition of p38 and ZAKalpha suppresses keratinocyte pyroptosis and rescues barrier integrity of Corynebacterium diphtheriae-treated organotypic skin. In summary, these findings implicate RSR and the NLRP1 inflammasome in antibacterial innate immunity and might explain certain aspects of diphtheria pathogenesis.

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“…One consequence of reduced ribosome speed can be ribosome collisions, which cells can perceive as signalling events that elicit a variety of cellular responses. Ribosome collisions can activate Gcn2, p38 and JNK kinases and reduce global translational activity in the affected cell ( Wu et al, 2020 ; Snieckute et al, 2022 ). At the same time, colliding ribosomes can activate the E3 ubiquitin ligases of the Ribosomal Quality Control pathway ( Juszkiewicz et al, 2020 ) and the ribosomal RNAse activity which is part of the No-Go decay pathway ( Doma and Parker, 2006 ).…”

Section: Nucleotide Modifications and Translational Efficiencymentioning

confidence: 99%

Translation of in vitro-transcribed RNA therapeutics

Haar

Mulroney²,

Hedayioglu³

et al. 2023

Front. Mol. Biosci.

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In vitro transcribed, modified messenger RNAs (IVTmRNAs) have been used to vaccinate billions of individuals against the SARS-CoV-2 virus, and are currently being developed for many additional therapeutic applications. IVTmRNAs must be translated into proteins with therapeutic activity by the same cellular machinery that also translates native endogenous transcripts. However, different genesis pathways and routes of entry into target cells as well as the presence of modified nucleotides mean that the way in which IVTmRNAs engage with the translational machinery, and the efficiency with which they are being translated, differs from native mRNAs. This review summarises our current knowledge of commonalities and differences in translation between IVTmRNAs and cellular mRNAs, which is key for the development of future design strategies that can generate IVTmRNAs with improved activity in therapeutic applications.

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Ribosome stalling is a signal for metabolic regulation by the ribotoxic stress response

Cited by 45 publications

References 46 publications

Computational and Functional Analysis of Structural Features in the ZAKα Kinase

Computational and Functional Analysis of Structural Features in the ZAKα Kinase

Corynebacterium diphtheriaecauses keratinocyte-intrinsic ribotoxic stress and NLRP1 inflammasome activation in a model of cutaneous diphtheria

Translation of in vitro-transcribed RNA therapeutics

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