S-lactoyl modification of KEAP1 by a reactive glycolytic metabolite activates NRF2 signaling

Ko, Yeonjin; Hong, Mannkyu; Lee, Seungbeom; Kumar, Manoj; Ibrahim, Lara; Nutsch, Kayla; Stanton, Caroline; Sondermann, Peter; Sandoval, Braddock A.; Bulos, Maya L.; Iaconelli, Jonathan; Chatterjee, Arnab K.; Wiseman, R. Luke; Schultz, Peter G.; Bollong, Michael J.

doi:10.1073/pnas.2300763120

Cited by 16 publications

(11 citation statements)

References 30 publications

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“…Treating HEK293T cells with 1 mM propachlor for 30 min decreased the GAPDH activity by 25% (Figure A). GAPDH inhibition is associated with the accumulation of its substrate glyceraldehyde-3-phosphate, which can convert to reactive species that alkylate KEAP1 to induce the oxidative stress response . Indeed, we see that KEAP1 is significantly destabilized by propachlor exposure by the Hsp40 affinity assay (FC = 1.3, p = 0.019, q = 0.02) (Table S4).…”

Section: Resultsmentioning

confidence: 93%

Cellular Exposure to Chloroacetanilide Herbicides Induces Distinct Protein Destabilization Profiles

et al. 2023

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Herbicides in the widely used chloroacetanilide class harbor a potent electrophilic moiety, which can damage proteins through nucleophilic substitution. In general, damaged proteins are subject to misfolding. Accumulation of misfolded proteins compromises cellular integrity by disrupting cellular proteostasis networks, which can further destabilize the cellular proteome. While direct conjugation targets can be discovered through affinity-based protein profiling, there are few approaches to probe how cellular exposure to toxicants impacts the stability of the proteome. We apply a quantitative proteomics methodology to identify chloroacetanilide-destabilized proteins in HEK293T cells based on their binding to the H31Q mutant of the human Hsp40 chaperone DNAJB8. We find that a brief cellular exposure to the chloroacetanilides acetochlor, alachlor, and propachlor induces misfolding of dozens of cellular proteins. These herbicides feature distinct but overlapping profiles of protein destabilization, highly concentrated in proteins with reactive cysteine residues. Consistent with the recent literature from the pharmacology field, reactivity is driven by neither inherent nucleophilic nor electrophilic reactivity but is idiosyncratic. We discover that propachlor induces a general increase in protein aggregation and selectively targets GAPDH and PARK7, leading to a decrease in their cellular activities. Hsp40 affinity profiling identifies a majority of propachlor targets identified by competitive activity-based protein profiling (ABPP), but ABPP can only identify about 10% of protein targets identified by Hsp40 affinity profiling. GAPDH is primarily modified by the direct conjugation of propachlor at a catalytic cysteine residue, leading to global destabilization of the protein. The Hsp40 affinity strategy is an effective technique to profile cellular proteins that are destabilized by cellular toxin exposure. Raw proteomics data is available through the PRIDE Archive at PXD030635.

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Section: Resultsmentioning

confidence: 93%

Cellular Exposure to Chloroacetanilide Herbicides Induces Distinct Protein Destabilization Profiles

et al. 2023

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“…Methylglyoxal is highly reactive and could non‐enzymatically cross‐link and dimerize Keap1 molecules via a methyl imidazole‐based linkage between cysteine (C151) and arginine (R15 or R135) residues, facilitating Nrf2 signaling 72 . More recently, it has been shown that accumulation of glyceraldehyde 3‐p elicits S‐lactoylation of several cysteines in Keap1 by mechanisms that have yet to be elucidated 73 . Since glyceraldehyde 3‐p readily accumulates during sprint exercise, 13 it could contribute to Nrf2 signaling by impeding the inhibitory action of Keap1 through its S‐lactoylation.…”

Section: Discussionmentioning

confidence: 99%

“…13,15,24,71 Keap1 cysteines may be covalently modified by several endogenous metabolites. 62,63,72,73 For example, by methylglyoxal, an electrophile that results from the elimination of triose phosphate, which accumulates in conditions with high glycolytic flux, as occurs during sprint in normoxia 13,71 and sprint 15 or moderateintensity exercise in hypoxia. 74 Methylglyoxal is highly reactive and could non-enzymatically cross-link and dimerize Keap1 molecules via a methyl imidazole-based linkage between cysteine (C151) and arginine (R15 or R135) residues, facilitating Nrf2 signaling.…”

Section: T a B L Ementioning

confidence: 99%

Physiological and molecular predictors of cycling sprint performance

Galvan‐Alvarez,

Gallego‐Selles,

Martinez‐Canton

et al. 2024

Scandinavian Med Sci Sports

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The study aimed to identify novel muscle phenotypic factors that could determine sprint performance using linear regression models including the lean mass of the lower extremities (LLM), myosin heavy chain composition (MHC), and proteins and enzymes implicated in glycolytic and aerobic energy generation (citrate synthase, OXPHOS proteins), oxygen transport and diffusion (myoglobin), ROS sensing (Nrf2/Keap1), antioxidant enzymes, and proteins implicated in calcium handling. For this purpose, body composition (dual‐energy X‐ray absorptiometry) and sprint performance (isokinetic 30‐s Wingate test: peak and mean power output, Wpeak and Wmean) were measured in young physically active adults (51 males and 10 females), from which a resting muscle biopsy was obtained from the musculus vastus lateralis. Although females had a higher percentage of MHC I, SERCA2, pSer16/Thr17‐phospholamban, and Calsequestrin 2 protein expressions (all p < 0.05), and 18.4% lower phosphofructokinase 1 protein expression than males (p < 0.05), both sexes had similar sprint performance when it was normalized to body weight or LLM. Multiple regression analysis showed that Wpeak could be predicted from LLM, SDHB, Keap1, and MHC II % (R 2 = 0.62, p < 0.001), each variable contributing to explain 46.4%, 6.3%, 4.4%, and 4.3% of the variance in Wpeak, respectively. LLM and MHC II % explained 67.5% and 2.1% of the variance in Wmean, respectively (R 2 = 0.70, p < 0.001). The present investigation shows that SDHB and Keap1, in addition to MHC II %, are relevant determinants of peak power output during sprinting.

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“…Interestingly, KEAP1 can also be modified at cysteine residues by fumarate and glyceraldehyde 3-phosphate ( 57 , 58 ), and by succinylation at a critical lysine sensor residue, also leading to increases in Nrf2 protein levels ( 59 ). Hence, it appears that these PTMs of KEAP1 serve as a generic switch to induce a regulatory stress response to different reactive metabolites ( 59 ).…”

Section: Impact Of Itaconate On Cell Signaling and Metabolismmentioning

confidence: 99%

Control of immune cell signaling by the immuno-metabolite itaconate

Lang,

Siddique

2024

Front. Immunol.

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Immune cell activation triggers signaling cascades leading to transcriptional reprogramming, but also strongly impacts on the cell’s metabolic activity to provide energy and biomolecules for inflammatory and proliferative responses. Macrophages activated by microbial pathogen-associated molecular patterns and cytokines upregulate expression of the enzyme ACOD1 that generates the immune-metabolite itaconate by decarboxylation of the TCA cycle metabolite cis-aconitate. Itaconate has anti-microbial as well as immunomodulatory activities, which makes it attractive as endogenous effector metabolite fighting infection and restraining inflammation. Here, we first summarize the pathways and stimuli inducing ACOD1 expression in macrophages. The focus of the review then lies on the mechanisms by which itaconate, and its synthetic derivatives and endogenous isomers, modulate immune cell signaling and metabolic pathways. Multiple targets have been revealed, from inhibition of enzymes to the post-translational modification of many proteins at cysteine or lysine residues. The modulation of signaling proteins like STING, SYK, JAK1, RIPK3 and KEAP1, transcription regulators (e.g. Tet2, TFEB) and inflammasome components (NLRP3, GSDMD) provides a biochemical basis for the immune-regulatory effects of the ACOD1-itaconate pathway. While the field has intensely studied control of macrophages by itaconate in infection and inflammation models, neutrophils have now entered the scene as producers and cellular targets of itaconate. Furthermore, regulation of adaptive immune responses by endogenous itaconate, as well as by exogenously added itaconate and derivatives, can be mediated by direct and indirect effects on T cells and antigen-presenting cells, respectively. Taken together, research in ACOD1-itaconate to date has revealed its relevance in diverse immune cell signaling pathways, which now provides opportunities for potential therapeutic or preventive manipulation of host defense and inflammation.

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S-lactoyl modification of KEAP1 by a reactive glycolytic metabolite activates NRF2 signaling

Cited by 16 publications

References 30 publications

Cellular Exposure to Chloroacetanilide Herbicides Induces Distinct Protein Destabilization Profiles

Cellular Exposure to Chloroacetanilide Herbicides Induces Distinct Protein Destabilization Profiles

Physiological and molecular predictors of cycling sprint performance

Control of immune cell signaling by the immuno-metabolite itaconate

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