21 results on '"Paul A. Grimsrud"'
Search Results
2. HGFAC is a ChREBP-regulated hepatokine that enhances glucose and lipid homeostasis
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Ashot Sargsyan, Ludivine Doridot, Sarah A. Hannou, Wenxin Tong, Harini Srinivasan, Rachael Ivison, Ruby Monn, Henry H. Kou, Jonathan M. Haldeman, Michelle Arlotto, Phillip J. White, Paul A. Grimsrud, Inna Astapova, Linus T. Tsai, and Mark A. Herman
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Metabolism ,Medicine - Abstract
Carbohydrate response element–binding protein (ChREBP) is a carbohydrate-sensing transcription factor that regulates both adaptive and maladaptive genomic responses in coordination of systemic fuel homeostasis. Genetic variants in the ChREBP locus associate with diverse metabolic traits in humans, including circulating lipids. To identify novel ChREBP-regulated hepatokines that contribute to its systemic metabolic effects, we integrated ChREBP ChIP-Seq analysis in mouse liver with human genetic and genomic data for lipid traits and identified hepatocyte growth factor activator (HGFAC) as a promising ChREBP-regulated candidate in mice and humans. HGFAC is a protease that activates the pleiotropic hormone hepatocyte growth factor. We demonstrate that HGFAC-KO mice had phenotypes concordant with putative loss-of-function variants in human HGFAC. Moreover, in gain- and loss-of-function genetic mouse models, we demonstrate that HGFAC enhanced lipid and glucose homeostasis, which may be mediated in part through actions to activate hepatic PPARγ activity. Together, our studies show that ChREBP mediated an adaptive response to overnutrition via activation of HGFAC in the liver to preserve glucose and lipid homeostasis.
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- 2023
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3. Proteomics and phosphoproteomics datasets of a muscle-specific STIM1 loss-of-function mouse model
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Scott P. Lyons, Rebecca J. Wilson, Deborah M. Muoio, and Paul A. Grimsrud
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Mass spectrometry ,Protein phosphorylation ,Protein abundance ,Isobaric tags ,R script ,Calcium homeostasis ,Computer applications to medicine. Medical informatics ,R858-859.7 ,Science (General) ,Q1-390 - Abstract
STIM1 is an ER/SR transmembrane protein that interacts with ORAI1 to activate store operated Ca2+ entry (SOCE) upon ER/SR depletion of calcium. Normally highly expressed in skeletal muscle, STIM1 deficiency causes significant changes to mitochondrial ultrastructure that do not occur with loss of ORAI1 or other components of SOCE. The datasets in this article are from large-scale proteomics and phosphoproteomics experiments in an inducible mouse model of skeletal muscle-specific STIM1 knock out (KO). These data reveal statistically significant changes in the relative abundance of specific proteins and sites of protein phosphorylation in STIM1 KO gastrocnemius. Protein samples from five biological replicates of each condition (+/- STIM1) were enzymatically digested, the resulting peptides labeled with tandem mass tag (TMT) reagents, mixed, and fractionated. Phosphopeptides were enriched and a small amount of each input retained for protein abundance analysis. All phosphopeptide and input fractions were analyzed by nano LC-MS/MS on a Q Exactive Plus Orbitrap mass spectrometer, searched with Proteome Discoverer software, and processed with in-house R-scripts for data normalization and statistical analysis. Article published in Molecular Metabolism [1].
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- 2022
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4. Disruption of STIM1-mediated Ca2+ sensing and energy metabolism in adult skeletal muscle compromises exercise tolerance, proteostasis, and lean mass
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Rebecca J. Wilson, Scott P. Lyons, Timothy R. Koves, Victoria G. Bryson, Hengtao Zhang, TianYu Li, Scott B. Crown, Jin-Dong Ding, Paul A. Grimsrud, Paul B. Rosenberg, and Deborah M. Muoio
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Skeletal muscle ,STIM1 ,Proteostasis ,Energy metabolism ,Exercise tolerance ,Mitochondria ,Internal medicine ,RC31-1245 - Abstract
Objective: Stromal interaction molecule 1 (STIM1) is a single-pass transmembrane endoplasmic/sarcoplasmic reticulum (E/SR) protein recognized for its role in a store operated Ca2+ entry (SOCE), an ancient and ubiquitous signaling pathway. Whereas STIM1 is known to be indispensable during development, its biological and metabolic functions in mature muscles remain unclear. Methods: Conditional and tamoxifen inducible muscle STIM1 knock-out mouse models were coupled with multi-omics tools and comprehensive physiology to understand the role of STIM1 in regulating SOCE, mitochondrial quality and bioenergetics, and whole-body energy homeostasis. Results: This study shows that STIM1 is abundant in adult skeletal muscle, upregulated by exercise, and is present at SR-mitochondria interfaces. Inducible tissue-specific deletion of STIM1 (iSTIM1 KO) in adult muscle led to diminished lean mass, reduced exercise capacity, and perturbed fuel selection in the settings of energetic stress, without affecting whole-body glucose tolerance. Proteomics and phospho-proteomics analyses of iSTIM1 KO muscles revealed molecular signatures of low-grade E/SR stress and broad activation of processes and signaling networks involved in proteostasis. Conclusion: These results show that STIM1 regulates cellular and mitochondrial Ca2+ dynamics, energy metabolism and proteostasis in adult skeletal muscles. Furthermore, these findings provide insight into the pathophysiology of muscle diseases linked to disturbances in STIM1-dependent Ca2+ handling.
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- 2022
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5. Nicotinamide riboside supplementation confers marginal metabolic benefits in obese mice without remodeling the muscle acetyl-proteome
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Ashley S. Williams, Timothy R. Koves, Yasminye D. Pettway, James A. Draper, Dorothy H. Slentz, Paul A. Grimsrud, Olga R. Ilkayeva, and Deborah M. Muoio
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Physiology ,Proteomics ,Nutrition ,Science - Abstract
Summary: Nicotinamide riboside supplements (NRS) have been touted as a nutraceutical that promotes cardiometabolic and musculoskeletal health by enhancing nicotinamide adenine dinucleotide (NAD+) biosynthesis, mitochondrial function, and/or the activities of NAD-dependent sirtuin deacetylase enzymes. This investigation examined the impact of NRS on whole body energy homeostasis, skeletal muscle mitochondrial function, and corresponding shifts in the acetyl-lysine proteome, in the context of diet-induced obesity using C57BL/6NJ mice. The study also included a genetically modified mouse model that imposes greater demand on sirtuin flux and associated NAD+ consumption, specifically within muscle tissues. In general, whole body glucose control was marginally improved by NRS when administered at the midpoint of a chronic high-fat diet, but not when given as a preventative therapy upon initiation of the diet. Contrary to anticipated outcomes, the study produced little evidence that NRS increases tissue NAD+ levels, augments mitochondrial function, and/or mitigates diet-induced hyperacetylation of the skeletal muscle proteome.
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- 2022
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6. Lipids Reprogram Metabolism to Become a Major Carbon Source for Histone Acetylation
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Eoin McDonnell, Scott B. Crown, Douglas B. Fox, Betül Kitir, Olga R. Ilkayeva, Christian A. Olsen, Paul A. Grimsrud, and Matthew D. Hirschey
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histone ,acetylation ,epigenetics ,metabolism ,fatty acid ,lipids ,gene expression ,proteomics ,metabolomics ,Biology (General) ,QH301-705.5 - Abstract
Cells integrate nutrient sensing and metabolism to coordinate proper cellular responses to a particular nutrient source. For example, glucose drives a gene expression program characterized by activating genes involved in its metabolism, in part by increasing glucose-derived histone acetylation. Here, we find that lipid-derived acetyl-CoA is a major source of carbon for histone acetylation. Using 13C-carbon tracing combined with acetyl-proteomics, we show that up to 90% of acetylation on certain histone lysines can be derived from fatty acid carbon, even in the presence of excess glucose. By repressing both glucose and glutamine metabolism, fatty acid oxidation reprograms cellular metabolism, leading to increased lipid-derived acetyl-CoA. Gene expression profiling of octanoate-treated hepatocytes shows a pattern of upregulated lipid metabolic genes, demonstrating a specific transcriptional response to lipid. These studies expand the landscape of nutrient sensing and uncover how lipids and metabolism are integrated by epigenetic events that control gene expression.
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- 2016
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7. Respiratory Phenomics across Multiple Models of Protein Hyperacylation in Cardiac Mitochondria Reveals a Marginal Impact on Bioenergetics
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Kelsey H. Fisher-Wellman, James A. Draper, Michael T. Davidson, Ashley S. Williams, Tara M. Narowski, Dorothy H. Slentz, Olga R. Ilkayeva, Robert D. Stevens, Gregory R. Wagner, Rami Najjar, Mathew D. Hirschey, J. Will Thompson, David P. Olson, Daniel P. Kelly, Timothy R. Koves, Paul A. Grimsrud, and Deborah M. Muoio
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Biology (General) ,QH301-705.5 - Abstract
Summary: Acyl CoA metabolites derived from the catabolism of carbon fuels can react with lysine residues of mitochondrial proteins, giving rise to a large family of post-translational modifications (PTMs). Mass spectrometry-based detection of thousands of acyl-PTMs scattered throughout the proteome has established a strong link between mitochondrial hyperacylation and cardiometabolic diseases; however, the functional consequences of these modifications remain uncertain. Here, we use a comprehensive respiratory diagnostics platform to evaluate three disparate models of mitochondrial hyperacylation in the mouse heart caused by genetic deletion of malonyl CoA decarboxylase (MCD), SIRT5 demalonylase and desuccinylase, or SIRT3 deacetylase. In each case, elevated acylation is accompanied by marginal respiratory phenotypes. Of the >60 mitochondrial energy fluxes evaluated, the only outcome consistently observed across models is a ∼15% decrease in ATP synthase activity. In sum, the findings suggest that the vast majority of mitochondrial acyl PTMs occur as stochastic events that minimally affect mitochondrial bioenergetics. : Fisher-Wellman et al. use a recently developed mitochondrial diagnostics platform for deep phenotyping of heart mitochondria derived from three disparate genetic models of protein hyperacylation. Their findings oppose the notion that hyperacylation of the mitochondrial proteome leads to broad-ranging vulnerabilities in respiratory function and bioenergetics. Keywords: mitochondrial diagnostics, lysine acylation, malonylation, ATP synthase
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- 2019
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8. The Acetyl Group Buffering Action of Carnitine Acetyltransferase Offsets Macronutrient-Induced Lysine Acetylation of Mitochondrial Proteins
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Michael N. Davies, Lilja Kjalarsdottir, J. Will Thompson, Laura G. Dubois, Robert D. Stevens, Olga R. Ilkayeva, M. Julia Brosnan, Timothy P. Rolph, Paul A. Grimsrud, and Deborah M. Muoio
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Biology (General) ,QH301-705.5 - Abstract
Lysine acetylation (AcK), a posttranslational modification wherein a two-carbon acetyl group binds covalently to a lysine residue, occurs prominently on mitochondrial proteins and has been linked to metabolic dysfunction. An emergent theory suggests mitochondrial AcK occurs via mass action rather than targeted catalysis. To test this hypothesis, we performed mass spectrometry-based acetylproteomic analyses of quadriceps muscles from mice with skeletal muscle-specific deficiency of carnitine acetyltransferase (CrAT), an enzyme that buffers the mitochondrial acetyl-CoA pool by converting short-chain acyl-CoAs to their membrane permeant acylcarnitine counterparts. CrAT deficiency increased tissue acetyl-CoA levels and susceptibility to diet-induced AcK of broad-ranging mitochondrial proteins, coincident with diminished whole body glucose control. Sub-compartment acetylproteome analyses of muscles from obese mice and humans showed remarkable overrepresentation of mitochondrial matrix proteins. These findings reveal roles for CrAT and L-carnitine in modulating the muscle acetylproteome and provide strong experimental evidence favoring the nonenzymatic carbon pressure model of mitochondrial AcK.
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- 2016
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9. Remodeling of the Acetylproteome by SIRT3 Manipulation Fails to Affect Insulin Secretion or β Cell Metabolism in the Absence of Overnutrition
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Brett S. Peterson, Jonathan E. Campbell, Olga Ilkayeva, Paul A. Grimsrud, Matthew D. Hirschey, and Christopher B. Newgard
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Biology (General) ,QH301-705.5 - Abstract
Summary: SIRT3 is a nicotinamide adenine dinucleotide (NAD+)-dependent mitochondrial protein deacetylase purported to influence metabolism through post-translational modification of metabolic enzymes. Fuel-stimulated insulin secretion, which involves mitochondrial metabolism, could be susceptible to SIRT3-mediated effects. We used CRISPR/Cas9 technology to manipulate SIRT3 expression in β cells, resulting in widespread SIRT3-dependent changes in acetylation of key metabolic enzymes but no appreciable changes in glucose- or pyruvate-stimulated insulin secretion or metabolomic profile during glucose stimulation. Moreover, these broad changes in the SIRT3-targeted acetylproteome did not affect responses to nutritional or ER stress. We also studied mice with global SIRT3 knockout fed either standard chow (STD) or high-fat and high-sucrose (HFHS) diets. Only when chronically fed HFHS diet do SIRT3 KO animals exhibit a modest reduction in insulin secretion. We conclude that broad changes in mitochondrial protein acetylation in response to manipulation of SIRT3 are not sufficient to cause changes in islet function or metabolism. : Peterson et al. report that ablation of SIRT3 in 832/13 β cells dramatically alters the mitochondrial acetylproteome but does not affect insulin secretion, metabolomic profile, or β cell survival. Moreover, SIRT3 knockout causes a modest reduction in insulin secretion in mice fed a high-fat and high-sucrose but not a standard chow diet. Keywords: acetylation, metabolism, insulin secretion, pancreatic islets, mitochondria, post-translational modifications, diabetes
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- 2018
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10. Medicago PhosphoProtein Database: a repository for Medicago truncatula phosphoprotein data
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Christopher M. Rose, Muthusubramanian eVenkateshwaran, Paul A. Grimsrud, Michael S. Westphall, Michael R. Sussman, Joshua J. Coon, and Jean-Michel eAné
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Medicago truncatula ,Proteomics ,CAD ,phosphoproteome ,ETD ,Plant culture ,SB1-1110 - Abstract
The ability of legume crops to fix atmospheric nitrogen via a symbiotic association with soil rhizobia makes them an essential component of many agricultural systems. Initiation of this symbiosis requires protein phosphorylation-mediated signaling in response to rhizobial signals named Nod factors. Medicago truncatula (Medicago) is the model system for studying legume biology, making the study of its phosphoproteome essential. Here, we describe the Medicago Phosphoprotein Database (http://phospho.medicago.wisc.edu), a repository built to house phosphoprotein, phosphopeptide, and phosphosite data specific to Medicago. Currently, the Medicago Phosphoprotein Database holds 3,457 unique phosphopeptides that contain 3,404 non-redundant sites of phosphorylation on 829 proteins. Through the web-based interface, users are allowed to browse identified proteins or search for proteins of interest. Furthermore, we allow users to conduct BLAST searches of the database using both peptide sequences and phosphorylation motifs as queries. The data contained within the database are available for download to be investigated at the user’s discretion. The Medicago Phosphoprotein Database will be updated continually with novel phosphoprotein and phosphopeptide identifications, with the intent of constructing an unparalleled compendium of large-scale Medicago phosphorylation data.
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- 2012
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11. Leveraging proteomics to understand plant-microbe interactions
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Dhileepkumar eJayaraman, Kari L. Forshey, Paul A. Grimsrud, and Jean-Michel eAné
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Plants ,Proteomics ,Symbiosis ,signaling ,Defense ,Plant culture ,SB1-1110 - Abstract
Understanding the interactions of plants with beneficial and pathogenic microbes is a promising avenue to improve crop productivity and agriculture sustainability. Proteomic techniques provide a unique angle to describe these intricate interactions and test hypotheses. The various approaches for proteome analysis generally include protein/peptide separation and identification, but can also provide quantification and the characterization of post-translational modifications. In this review, we discuss how these techniques have been applied to the study to plant-microbe interactions. We also present some areas where this field of study would benefit from the utilization of newly developed methods that overcome previous limitations. Finally, we reinforce the need for expanding, integrating, and curating protein databases, as well as the benefits of combining protein-level datasets with genetics and other high-throughput large-scale approaches for a systems-level understanding of plant-microbe interactions.
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- 2012
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12. Nicotinamide riboside supplementation confers marginal metabolic benefits in obese mice without remodeling the muscle acetyl-proteome
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Ashley S. Williams, Timothy R. Koves, Yasminye D. Pettway, James A. Draper, Dorothy H. Slentz, Paul A. Grimsrud, Olga R. Ilkayeva, and Deborah M. Muoio
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Proteomics ,Multidisciplinary ,Physiology ,Science ,Article ,Nutrition - Abstract
Summary Nicotinamide riboside supplements (NRS) have been touted as a nutraceutical that promotes cardiometabolic and musculoskeletal health by enhancing nicotinamide adenine dinucleotide (NAD+) biosynthesis, mitochondrial function, and/or the activities of NAD-dependent sirtuin deacetylase enzymes. This investigation examined the impact of NRS on whole body energy homeostasis, skeletal muscle mitochondrial function, and corresponding shifts in the acetyl-lysine proteome, in the context of diet-induced obesity using C57BL/6NJ mice. The study also included a genetically modified mouse model that imposes greater demand on sirtuin flux and associated NAD+ consumption, specifically within muscle tissues. In general, whole body glucose control was marginally improved by NRS when administered at the midpoint of a chronic high-fat diet, but not when given as a preventative therapy upon initiation of the diet. Contrary to anticipated outcomes, the study produced little evidence that NRS increases tissue NAD+ levels, augments mitochondrial function, and/or mitigates diet-induced hyperacetylation of the skeletal muscle proteome., Graphical abstract, Highlights • Dietary NR supplementation (NRS) raises plasma and muscle NAM levels in obese mice • NRS given post obesity slightly improved glucose control and mitochondrial function • NRS did not oppose obesity-induced remodeling of the muscle acetyl-proteome • NRS during weight gain did not protect glucose control and mitochondrial function, Physiology; Proteomics; Nutrition
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- 2021
13. Lipids Reprogram Metabolism to Become a Major Carbon Source for Histone Acetylation
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Douglas B. Fox, Matthew D. Hirschey, Paul A. Grimsrud, Olga Ilkayeva, Scott B. Crown, Christian A. Olsen, Eoin McDonnell, and Betül Kitir
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0301 basic medicine ,Nutrient sensing ,histone ,General Biochemistry, Genetics and Molecular Biology ,Article ,Histones ,lipids ,03 medical and health sciences ,Mice ,proteomics ,Acetyl Coenzyme A ,Cell Line, Tumor ,Animals ,Humans ,Epigenetics ,Amino Acid Sequence ,Beta oxidation ,lcsh:QH301-705.5 ,acetylation ,Regulation of gene expression ,biology ,epigenetics ,Gene Expression Profiling ,Lipid metabolism ,Lipid Metabolism ,metabolomics ,Carbon ,Histone Deacetylase Inhibitors ,030104 developmental biology ,Histone ,Biochemistry ,Gene Expression Regulation ,lcsh:Biology (General) ,Acetylation ,Histone methyltransferase ,biology.protein ,gene expression ,lipids (amino acids, peptides, and proteins) ,fatty acid ,Caprylates ,Oxidation-Reduction ,metabolism - Abstract
SummaryCells integrate nutrient sensing and metabolism to coordinate proper cellular responses to a particular nutrient source. For example, glucose drives a gene expression program characterized by activating genes involved in its metabolism, in part by increasing glucose-derived histone acetylation. Here, we find that lipid-derived acetyl-CoA is a major source of carbon for histone acetylation. Using 13C-carbon tracing combined with acetyl-proteomics, we show that up to 90% of acetylation on certain histone lysines can be derived from fatty acid carbon, even in the presence of excess glucose. By repressing both glucose and glutamine metabolism, fatty acid oxidation reprograms cellular metabolism, leading to increased lipid-derived acetyl-CoA. Gene expression profiling of octanoate-treated hepatocytes shows a pattern of upregulated lipid metabolic genes, demonstrating a specific transcriptional response to lipid. These studies expand the landscape of nutrient sensing and uncover how lipids and metabolism are integrated by epigenetic events that control gene expression.
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- 2016
14. Nicotinamide mononucleotide requires SIRT3 to improve cardiac function and bioenergetics in a Friedreich’s ataxia cardiomyopathy model
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Kathleen A. Hershberger, Lan Mao, R. Mark Payne, Paul A. Grimsrud, Angelical Martin, Xiaojing Liu, Huaxia Cui, Dhaval P. Bhatt, Jason W. Locasale, Juan Liu, Matthew D. Hirschey, Michael J. Muehlbauer, and Dennis M. Abraham
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0301 basic medicine ,Cardiac function curve ,Ataxia ,SIRT3 ,biology ,Chemistry ,Cardiomyopathy ,General Medicine ,Pharmacology ,medicine.disease ,03 medical and health sciences ,030104 developmental biology ,Biochemistry ,Sirtuin ,biology.protein ,medicine ,Protein deacetylase ,NAD+ kinase ,medicine.symptom ,Nicotinamide mononucleotide ,Research Article - Abstract
Increasing NAD+ levels by supplementing with the precursor nicotinamide mononucleotide (NMN) improves cardiac function in multiple mouse models of disease. While NMN influences several aspects of mitochondrial metabolism, the molecular mechanisms by which increased NAD+ enhances cardiac function are poorly understood. A putative mechanism of NAD+ therapeutic action exists via activation of the mitochondrial NAD+-dependent protein deacetylase sirtuin 3 (SIRT3). We assessed the therapeutic efficacy of NMN and the role of SIRT3 in the Friedreich's ataxia cardiomyopathy mouse model (FXN-KO). At baseline, the FXN-KO heart has mitochondrial protein hyperacetylation, reduced Sirt3 mRNA expression, and evidence of increased NAD+ salvage. Remarkably, NMN administered to FXN-KO mice restores cardiac function to near-normal levels. To determine whether SIRT3 is required for NMN therapeutic efficacy, we generated SIRT3-KO and SIRT3-KO/FXN-KO (double KO [dKO]) models. The improvement in cardiac function upon NMN treatment in the FXN-KO is lost in the dKO model, demonstrating that the effects of NMN are dependent upon cardiac SIRT3. Coupled with cardio-protection, SIRT3 mediates NMN-induced improvements in both cardiac and extracardiac metabolic function and energy metabolism. Taken together, these results serve as important preclinical data for NMN supplementation or SIRT3 activator therapy in Friedreich's ataxia patients.
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- 2017
15. A Class of Reactive Acyl-CoA Species Reveals the Non-Enzymatic Origins of Protein Acylation
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Gregory R. Wagner, Donald S. Backos, Robert Stevens, Hao Yang, Dhaval P. Bhatt, Matthew D. Hirschey, Grant A. Mitchell, Paul A. Grimsrud, J. Will Thompson, Thomas M. O’Connell, Olga Ilkayeva, and Laura G. Dubois
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0301 basic medicine ,Physiology ,Acylation ,Lysine ,Quantitative proteomics ,Reactive intermediate ,Chemical biology ,Cell Biology ,Biology ,Proteomics ,Malate dehydrogenase ,Article ,Citric acid cycle ,03 medical and health sciences ,Protein acylation ,030104 developmental biology ,0302 clinical medicine ,Biochemistry ,lipids (amino acids, peptides, and proteins) ,Molecular Biology ,Protein Processing, Post-Translational ,030217 neurology & neurosurgery - Abstract
The mechanisms underlying the formation of acyl protein modifications remain poorly understood. By investigating the reactivity of endogenous acyl-CoA metabolites, we found a class of acyl-CoAs that undergoes intramolecular catalysis to form reactive intermediates which non-enzymatically modify proteins. Based on this mechanism, we predicted, validated, and characterized a protein modification: 3-hydroxy-3-methylglutaryl(HMG)-lysine. In a model of altered HMG-CoA metabolism, we found evidence of two additional protein modifications: 3-methylglutaconyl(MGc)-lysine and 3-methylglutaryl(MG)-lysine. Using quantitative proteomics, we compared the ‘acylomes’ of two reactive acyl-CoA species, namely HMG-CoA and glutaryl-CoA, which are generated in different pathways. We found proteins that are uniquely modified by each reactive metabolite, as well as common proteins and pathways. We identified the tricarboxylic acid cycle as a pathway commonly regulated by acylation, and validated malate dehydrogenase as a key target. These data uncover a fundamental relationship between reactive acyl-CoA species and proteins, and define a new regulatory paradigm in metabolism.
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- 2017
16. Downregulation of Adipose Glutathione S-Transferase A4 Leads to Increased Protein Carbonylation, Oxidative Stress, and Mitochondrial Dysfunction
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Olga Ilkayeva, Xin Xu, Wendy Wright, Deborah E. Muoio, Brian M. Wiczer, Paul A. Grimsrud, David A. Bernlohr, David W. Graham, Edgar A. Arriaga, Katherine Cianflone, Jonathan R. Brestoff, Jessica Curtis, and Rocio Foncea
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Mitochondrial ROS ,medicine.medical_specialty ,Endocrinology, Diabetes and Metabolism ,Protein Carbonylation ,Adipose tissue ,Down-Regulation ,Mitochondrion ,Biology ,03 medical and health sciences ,chemistry.chemical_compound ,Mice ,0302 clinical medicine ,Insulin resistance ,Downregulation and upregulation ,Adipocyte ,Internal medicine ,3T3-L1 Cells ,Internal Medicine ,medicine ,Animals ,Humans ,Obesity ,030304 developmental biology ,Glutathione Transferase ,Oligonucleotide Array Sequence Analysis ,chemistry.chemical_classification ,Mice, Knockout ,0303 health sciences ,Reactive oxygen species ,medicine.disease ,Mitochondria ,Mice, Inbred C57BL ,Oxidative Stress ,Endocrinology ,Metabolism ,chemistry ,Original Article ,Insulin Resistance ,030217 neurology & neurosurgery - Abstract
OBJECTIVE Peripheral insulin resistance is linked to an increase in reactive oxygen species (ROS), leading in part to the production of reactive lipid aldehydes that modify the side chains of protein amino acids in a reaction termed protein carbonylation. The primary enzymatic method for lipid aldehyde detoxification is via glutathione S-transferase A4 (GSTA4) dependent glutathionylation. The objective of this study was to evaluate the expression of GSTA4 and the role(s) of protein carbonylation in adipocyte function. RESEARCH DESIGN AND METHODS GSTA4-silenced 3T3-L1 adipocytes and GSTA4-null mice were evaluated for metabolic processes, mitochondrial function, and reactive oxygen species production. GSTA4 expression in human obesity was evaluated using microarray analysis. RESULTS GSTA4 expression is selectively downregulated in adipose tissue of obese insulin-resistant C57BL/6J mice and in human obesity-linked insulin resistance. Tumor necrosis factor-α treatment of 3T3-L1 adipocytes decreased GSTA4 expression, and silencing GSTA4 mRNA in cultured adipocytes resulted in increased protein carbonylation, increased mitochondrial ROS, dysfunctional state 3 respiration, and altered glucose transport and lipolysis. Mitochondrial function in adipocytes of lean or obese GSTA4-null mice was significantly compromised compared with wild-type controls and was accompanied by an increase in superoxide anion. CONCLUSIONS These results indicate that downregulation of GSTA4 in adipose tissue leads to increased protein carbonylation, ROS production, and mitochondrial dysfunction and may contribute to the development of insulin resistance and type 2 diabetes.
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- 2010
17. Quantification of Mitochondrial Acetylation Dynamics Highlights Prominent Sites of Metabolic Regulation*
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Joshua J. Coon, Alan D. Attie, Brendan J. Floyd, Drew R. Gunderson, John M. Denu, David J. Pagliarini, Kristin E. Dittenhafer-Reed, Alexander S. Hebert, Amelia Still, Brendan K. Dolan, Mark P. Keller, Paul A. Grimsrud, Joshua J. Carson, Michael S. Westphall, Craig A. Bingman, and Donald S. Stapleton
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SIRT3 ,Proteome ,Mice, Obese ,Mitochondria, Liver ,Biology ,Proteomics ,Biochemistry ,complex mixtures ,Protein acylation ,Mice ,Acetyl Coenzyme A ,Sirtuin 3 ,Animals ,Acetyl-CoA C-Acetyltransferase ,skin and connective tissue diseases ,Molecular Biology ,ACAT1 ,Acetylation ,Cell Biology ,Metabolic pathway ,Metabolism ,Acetyltransferase ,bacteria ,sense organs - Abstract
Lysine acetylation is rapidly becoming established as a key post-translational modification for regulating mitochondrial metabolism. Nonetheless, distinguishing regulatory sites from among the thousands identified by mass spectrometry and elucidating how these modifications alter enzyme function remain primary challenges. Here, we performed multiplexed quantitative mass spectrometry to measure changes in the mouse liver mitochondrial acetylproteome in response to acute and chronic alterations in nutritional status, and integrated these data sets with our compendium of predicted Sirt3 targets. These analyses highlight a subset of mitochondrial proteins with dynamic acetylation sites, including acetyl-CoA acetyltransferase 1 (Acat1), an enzyme central to multiple metabolic pathways. We performed in vitro biochemistry and molecular modeling to demonstrate that acetylation of Acat1 decreases its activity by disrupting the binding of coenzyme A. Collectively, our data reveal an important new target of regulatory acetylation and provide a foundation for investigating the role of select mitochondrial protein acetylation sites in mediating acute and chronic metabolic transitions.
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- 2013
18. A Proteogenomic Survey of the Medicago truncatula Genome*
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Paul A. Grimsrud, Michael R. Sussman, Maegen Howes-Podoll, Derek J. Bailey, Jean-Michel Ané, Joshua J. Coon, Jeremy D. Volkening, Michael S. Westphall, Muthusubramanian Venkateshwaran, and Christopher M. Rose
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Proteomics ,Proteome ,In silico ,Molecular Sequence Data ,Computational biology ,Biology ,Biochemistry ,Genome ,Deep sequencing ,Mass Spectrometry ,Analytical Chemistry ,Medicago truncatula ,Amino Acid Sequence ,Databases, Protein ,Molecular Biology ,Gene ,Plant Proteins ,Genetics ,Information Dissemination ,Research ,Sequence Analysis, DNA ,Proteogenomics ,biology.organism_classification ,Peptides ,Algorithms ,Genome, Plant - Abstract
Peptide sequencing by computational assignment of tandem mass spectra to a database of putative protein sequences provides an independent approach to confirming or refuting protein predictions based on large-scale DNA and RNA sequencing efforts. This use of mass spectrometrically-derived sequence data for testing and refining predicted gene models has been termed proteogenomics. We report herein the application of proteogenomic methodology to a database of 10.9 million tandem mass spectra collected over a period of two years from proteolytically generated peptides isolated from the model legume Medicago truncatula. These spectra were searched against a database of predicted M. truncatula protein sequences generated from public databases, in silico gene model predictions, and a whole-genome six-frame translation. This search identified 78,647 distinct peptide sequences, and a comparison with the publicly available proteome from the recently published M. truncatula genome supported translation of 9,843 existing gene models and identified 1,568 novel peptides suggesting corrections or additions to the current annotations. Each supporting and novel peptide was independently validated using mRNA-derived deep sequencing coverage and an overall correlation of 93% between the two data types was observed. We have additionally highlighted examples of several aspects of structural annotation for which tandem MS provides unique evidence not easily obtainable through typical DNA or RNA sequencing. Proteogenomic analysis is a valuable and unique source of information for the structural annotation of genomes and should be included in such efforts to ensure that the genome models used by biologists mirror as accurately as possible what is present in the cell.
- Published
- 2012
19. X-ray crystallographic analysis of adipocyte fatty acid binding protein (aP2) modified with 4-hydroxy-2-nonenal
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Douglas H. Ohlendorf, Andrew C. Kruse, David A. Bernlohr, Leonard J. Banaszak, Paul A. Grimsrud, and Kristina Hellberg
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Models, Molecular ,Stereochemistry ,Protein Conformation ,Molecular Sequence Data ,Peptide ,Cysteine Proteinase Inhibitors ,Crystallography, X-Ray ,Fatty Acid-Binding Proteins ,Ligands ,Biochemistry ,Fatty acid-binding protein ,Article ,chemistry.chemical_compound ,Mice ,Side chain ,Adipocytes ,Non-covalent interactions ,Animals ,Carboxylate ,Molecular Biology ,chemistry.chemical_classification ,Aldehydes ,Fatty acid ,Oxidative Stress ,chemistry ,Covalent bond ,lipids (amino acids, peptides, and proteins) ,Target protein ,Dimerization - Abstract
Fatty acid binding proteins (FABP) have been characterized as facilitating the intracellular solubilization and transport of long-chain fatty acyl carboxylates via noncovalent interactions. More recent work has shown that the adipocyte FABP is also covalently modified in vivo on Cys117 with 4-hydroxy-2-nonenal (4-HNE), a bioactive aldehyde linked to oxidative stress and inflammation. To evaluate 4-HNE binding and modification, the crystal structures of adipocyte FABP covalently and noncovalently bound to 4-HNE have been solved to 1.9 A and 2.3 A resolution, respectively. While the 4-HNE in the noncovalently modified protein is coordinated similarly to a carboxylate of a fatty acid, the covalent form show a novel coordination through a water molecule at the polar end of the lipid. Other defining features between the two structures with 4-HNE and previously solved structures of the protein include a peptide flip between residues Ala36 and Lys37 and the rotation of the side chain of Phe57 into its closed conformation. Representing the first structure of an endogenous target protein covalently modified by 4-HNE, these results define a new class of in vivo ligands for FABPs and extend their physiological substrates to include bioactive aldehydes.
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- 2010
20. Oxidative Stress and Covalent Modification of Protein with Bioactive Aldehydes*S⃞
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Paul A. Grimsrud, Hongwei Xie, David A. Bernlohr, and Timothy J. Griffin
- Subjects
Proteomics ,Antioxidant ,Protein Carbonylation ,medicine.medical_treatment ,Lysine ,medicine.disease_cause ,Biochemistry ,Mass Spectrometry ,chemistry.chemical_compound ,medicine ,Animals ,Humans ,Electrophoresis, Gel, Two-Dimensional ,Molecular Biology ,Histidine ,chemistry.chemical_classification ,Reactive oxygen species ,Aldehydes ,Proteins ,Minireviews ,Cell Biology ,Oxidative Stress ,chemistry ,Hydroxyl radical ,Reactive Oxygen Species ,Oxidative stress ,Cysteine - Abstract
The term “oxidative stress” links the production of reactive oxygen species to a variety of metabolic outcomes, including insulin resistance, immune dysfunction, and inflammation. Antioxidant defense systems down-regulated due to disease and/or aging result in oxidatively modified DNA, carbohydrates, proteins, and lipids. Increased production of hydroxyl radical leads to the formation of lipid hydroperoxides that produce a family of α,β-unsaturated aldehydes. Such reactive aldehydes are subject to Michael addition reactions with the side chains of lysine, histidine, and cysteine residues, referred to as “protein carbonylation.” Although not widely appreciated, reactive lipids can accumulate to high levels in cells, resulting in extensive protein modification leading to either loss or gain of function. The use of mass spectrometric methods to identify the site and extent of protein carbonylation on a proteome-wide scale has expanded our view of how oxidative stress can regulate cellular processes.
- Published
- 2008
21. The Acetyl Group Buffering Action of Carnitine Acetyltransferase Offsets Macronutrient-Induced Lysine Acetylation of Mitochondrial Proteins
- Author
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Robert Stevens, Paul A. Grimsrud, J. Will Thompson, Lilja Kjalarsdottir, Laura G. Dubois, Olga Ilkayeva, M. Julia Brosnan, Michael N. Davies, Deborah M. Muoio, and Timothy P. Rolph
- Subjects
0301 basic medicine ,endocrine system ,Glucose control ,Lysine ,Biology ,Article ,General Biochemistry, Genetics and Molecular Biology ,Mitochondrial Proteins ,Mice ,03 medical and health sciences ,Carnitine Acetyltransferase ,Animals ,Humans ,Mitochondrial protein ,lcsh:QH301-705.5 ,chemistry.chemical_classification ,Carnitine O-Acetyltransferase ,Acetylation ,Lysine residue ,030104 developmental biology ,Enzyme ,Biochemistry ,chemistry ,lcsh:Biology (General) ,Mitochondrial matrix - Abstract
SummaryLysine acetylation (AcK), a posttranslational modification wherein a two-carbon acetyl group binds covalently to a lysine residue, occurs prominently on mitochondrial proteins and has been linked to metabolic dysfunction. An emergent theory suggests mitochondrial AcK occurs via mass action rather than targeted catalysis. To test this hypothesis, we performed mass spectrometry-based acetylproteomic analyses of quadriceps muscles from mice with skeletal muscle-specific deficiency of carnitine acetyltransferase (CrAT), an enzyme that buffers the mitochondrial acetyl-CoA pool by converting short-chain acyl-CoAs to their membrane permeant acylcarnitine counterparts. CrAT deficiency increased tissue acetyl-CoA levels and susceptibility to diet-induced AcK of broad-ranging mitochondrial proteins, coincident with diminished whole body glucose control. Sub-compartment acetylproteome analyses of muscles from obese mice and humans showed remarkable overrepresentation of mitochondrial matrix proteins. These findings reveal roles for CrAT and L-carnitine in modulating the muscle acetylproteome and provide strong experimental evidence favoring the nonenzymatic carbon pressure model of mitochondrial AcK.
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