Dataset Information

Protein-metabolite interactomics of carbohydrate metabolism reveal regulation of lactate dehydrogenase

ABSTRACT:

Metabolic networks are interconnected and influence diverse cellular processes. The protein-metabolite interactions that mediate these networks are frequently low affinity and challenging to systematically discover. We developed mass spectrometry integrated with equilibrium dialysis for the discovery of allostery systematically (MIDAS) to identify such interactions. Analysis of 33 enzymes from human carbohydrate metabolism identified 830 protein-metabolite interactions, including known regulators, substrates and products as well as previously unreported interactions. We functionally validated a subset of interactions, including the isoform-specific inhibition of lactate dehydrogenase by long-chain acyl-coenzyme A. Cell treatment with fatty acids caused a loss of pyruvate-lactate interconversion dependent on lactate dehydrogenase isoform expression. These protein-metabolite interactions may contribute to the dynamic, tissue-specific metabolic flexibility that enables growth and survival in an ever-changing nutrient environment.

INSTRUMENT(S): Flow injection analysis MS - positive, Flow injection analysis MS - negative

SUBMITTER: Jordan Berg

PROVIDER: MTBLS7237 | MetaboLights | 2023-08-07

REPOSITORIES: MetaboLights

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Dataset's files

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			Action	DRS
	MTBLS7237	Other
	FILES	Other
	a_MTBLS7237_FIA-MS_negative__metabolite_profiling.txt	Txt
	a_MTBLS7237_FIA-MS_positive__metabolite_profiling.txt	Txt
	files-all.json	Other

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Publications

Protein-metabolite interactomics of carbohydrate metabolism reveal regulation of lactate dehydrogenase.

Hicks Kevin G KG Cluntun Ahmad A AA Schubert Heidi L HL Hackett Sean R SR Berg Jordan A JA Leonard Paul G PG Ajalla Aleixo Mariana A MA Zhou Youjia Y Bott Alex J AJ Salvatore Sonia R SR Chang Fei F Blevins Aubrie A Barta Paige P Tilley Samantha S Leifer Aaron A Guzman Andrea A Arok Ajak A Fogarty Sarah S Winter Jacob M JM Ahn Hee-Chul HC Allen Karen N KN Block Samuel S Cardoso Iara A IA Ding Jianping J Dreveny Ingrid I Gasper William C WC Ho Quinn Q Matsuura Atsushi A Palladino Michael J MJ Prajapati Sabin S Sun Pengkai P Tittmann Kai K Tolan Dean R DR Unterlass Judith J VanDemark Andrew P AP Vander Heiden Matthew G MG Webb Bradley A BA Yun Cai-Hong CH Zhao Pengkai P Wang Bei B Schopfer Francisco J FJ Hill Christopher P CP Nonato Maria Cristina MC Muller Florian L FL Cox James E JE Rutter Jared J

Science (New York, N.Y.) 20230309 6636

PMID: 36893255

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Project description:Background: In acute kidney injury, macrophages play a major role in regulating inflammation. Classically activated macrophages (M1) undergo drastic metabolic reprogramming during their differentiation and upregulate the aerobic glycolysis pathway to fulfill their pro-inflammatory functions. NAD+ regeneration is crucial for the maintenance of glycolysis and the most direct pathway by which this occurs is via the fermentation of pyruvate to lactate, catalyzed by lactate dehydrogenase A (LDHA). Our previous study determined that LDHA is predominantly expressed in the proximal segments of the nephron in the mouse kidney and increases with hypoxia. This study investigates the potential of LDHA as a therapeutic target for inflammation by exploring its role in macrophage function in vitro. Methods: Bone-marrow-derived macrophages (BMDMs) were isolated from myeloid-specific LDHA knockout mice derived from crossbreeding LysM-Cre transgenic mice and LDHA floxed mice. RNA sequencing and LC-MS/MS metabolomics analyses were used in this study to determine the effect of LDHA deletion on BMDM following stimulation with IFN-γ. Results: LDHA deletion in IFN-γ BMDMs resulted in a significant alteration of the macrophage activation and functional pathways, and change in glycolytic, cytokine, and chemokine gene expression. Metabolite concentrations associated with pro-inflammatory macrophage profiles were diminished while anti-inflammatory-associated ones were increased in LDHA KO BMDMs. Glutamate and amino sugars metabolic pathways were significantly affected by the LDHA deletion. A combined muti-omics analysis highlighted changes in Rap1 signaling, cytokine-cytokine receptor interaction, focal adhesion, and MAPK signaling metabolism pathways. Conclusions: Deletion of LDHA in macrophages results in a notable reduction in the pro-inflammatory profile and concurrent upregulation of anti-inflammatory pathways. These findings suggest that LDHA could serve as a promising therapeutic target for inflammation, a key contributor to the pathogenesis of acute kidney injury.