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Metabolic dysfunction-associated fatty liver disease (MAFLD) has a global prevalence of about 25% and no approved therapy. Using metabolomic and proteomic analyses, we identified high expression of hepatic transketolase (TKT), a metabolic enzyme of the pentose phosphate pathway, in human and mouse MAFLD. Hyperinsulinemia promoted TKT expression through the insulin receptor-CCAAT/enhancer-binding protein alpha axis. Utilizing liver-specific TKT overexpression and knockout mouse models, we demonstrated that TKT was sufficient and required for MAFLD progression. Further metabolic flux analysis revealed that Tkt deletion increased hepatic inosine levels to activate the protein kinase A-cAMP response element binding protein cascade, promote phosphatidylcholine synthesis, and improve mitochondrial function. Moreover, insulin induced hepatic TKT to limit inosine-dependent mitochondrial activity. Importantly, N-acetylgalactosamine (GalNAc)-siRNA conjugates targeting hepatic TKT showed promising therapeutic effects on mouse MAFLD. Our study uncovers how hyperinsulinemia regulates TKT-orchestrated inosine metabolism and mitochondrial function and provides a novel therapeutic strategy for MAFLD prevention and treatment., Competing Interests: Declaration of interests B.L. is a co-founder of Biotheus Inc and chairman of its scientific advisory board. J.L. is a co-founder of Shanghai Synvida Biotechnology Co. Ltd. X.T. has a patent related to transketolase therapy for metabolic disorders., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Regulatory T (T reg ) cells are critical for maintaining immune homeostasis and preventing autoimmunity. Here, we show that the non-oxidative pentose phosphate pathway (PPP) regulates T reg function to prevent autoimmunity. Deletion of transketolase (TKT), an indispensable enzyme of non-oxidative PPP, in T reg cells causes a fatal autoimmune disease in mice, with impaired T reg suppressive capability despite regular T reg numbers and normal Foxp3 expression levels. Mechanistically, reduced glycolysis and enhanced oxidative stress induced by TKT deficiency triggers excessive fatty acid and amino acid catabolism, resulting in uncontrolled oxidative phosphorylation and impaired mitochondrial fitness. Reduced α-KG levels as a result of reductive TCA cycle activity leads to DNA hypermethylation, thereby limiting functional gene expression and suppressive activity of TKT-deficient T reg cells. We also find that TKT levels are frequently downregulated in T reg cells of people with autoimmune disorders. Our study identifies the non-oxidative PPP as an integrator of metabolic and epigenetic processes that control T reg function., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Transketolase (Tkt), an enzyme in pentose phosphate pathway, has been reported to regulate genome instability and cell survival in cancers. Yet, the role of Tkt after myocardial ischemic injury remains to be elucidated. Label-free proteomics revealed dramatic elevation of Tkt in murine hearts after myocardial infarction (MI). Lentivirus-mediated Tkt knockdown ameliorated cardiomyocyte apoptosis and preserved the systolic function after myocardial ischemic injury. In contrast, Tkt overexpression led to the opposite effects. Inducible conditional cardiomyocyte Tkt-knockout mice were generated, and cardiomyocyte-expressed Tkt was found to play an intrinsic role in the ischemic heart failure of these model mice. Furthermore, through luciferase assay and chromatin immunoprecipitation, Tkt was shown to be a direct target of transcription factor Krüppel-like factor 5 (Klf5). In cardiomyocytes under ischemic stress, Tkt redistributed into the nucleus. By binding with the full-length poly(ADP-ribose) polymerase 1 (Parp1), facilitating its cleavage, and activating apoptosis inducible factor (Aif) subsequently, nuclear Tkt demonstrated its non-metabolic functions. Overall, our study confirmed that elevated nuclear Tkt plays a noncanonical role in promoting cardiomyocyte apoptosis via the cleaved Parp1/Aif pathway, leading to the deterioration of cardiac dysfunction., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany.)

Inflammatory bowel disease (IBD) has a close association with transketolase (TKT) that links glycolysis and the pentose phosphate pathway (PPP). However, how TKT functions in the intestinal epithelium remains to be elucidated. To address this question, we specifically delete TKT in intestinal epithelial cells (IECs). IEC TKT-deficient mice are growth retarded and suffer from spontaneous colitis. TKT ablation brings about striking alterations of the intestine, including extensive mucosal erosion, aberrant tight junctions, impaired barrier function, and increased inflammatory cell infiltration. Mechanistically, TKT deficiency significantly accumulates PPP metabolites and decreases glycolytic metabolites, thereby reducing ATP production, which results in excessive apoptosis and defective intestinal barrier. Therefore, our data demonstrate that TKT serves as an essential guardian of intestinal integrity and barrier function as well as a potential therapeutic target for intestinal disorders., (© 2021. The Author(s).)

Obesity has recently become a prevalent health threat worldwide. Although emerging evidence has suggested a strong link between the pentose phosphate pathway (PPP) and obesity, the role of transketolase (TKT), an enzyme in the nonoxidative branch of the PPP that connects PPP and glycolysis, remains obscure in adipose tissues. In this study, we specifically deleted TKT in mouse adipocytes and found no obvious phenotype upon normal diet feeding. However, adipocyte TKT abrogation attenuated high-fat diet-induced obesity, reduced hepatic steatosis, improved glucose tolerance, alleviated insulin resistance, and increased energy expenditure. Mechanistically, TKT deficiency accumulated nonoxidative PPP metabolites and decreased glycolysis and pyruvate input into the mitochondria, leading to increased lipolytic enzyme gene expression and enhanced lipolysis, fatty acid oxidation, and mitochondrial respiration. Therefore, our data not only identify a novel role of TKT in regulating lipolysis and obesity but also suggest that limiting glucose-derived carbon into the mitochondria induces lipid catabolism and energy expenditure., (© 2020 by the American Diabetes Association.)

De novo nucleotide biosynthesis is essential for maintaining cellular nucleotide pools, the suppression of which leads to genome instability. The metabolic enzyme transketolase (TKT) in the nonoxidative branch of the pentose phosphate pathway (PPP) regulates ribose 5-phosphate (R5P) levels and de novo nucleotide biosynthesis. TKT is required for maintaining cell proliferation in human liver cancer cell lines, yet the role of TKT in liver injury and cancer initiation remains to be elucidated. In this study, we generated a liver-specific TKT knockout mouse strain by crossing TKT flox/flox mice with albumin-Cre mice. Loss of TKT in hepatocytes protected the liver from diethylnitrosamine (DEN)-induced DNA damage without altering DEN metabolism. DEN treatment of TKT-null liver increased levels of R5P and promoted de novo nucleotide synthesis. More importantly, supplementation of dNTPs in primary hepatocytes alleviated DEN-induced DNA damage, cell death, inflammatory response, and cell proliferation. Furthermore, DEN and high-fat diet (HFD)-induced liver carcinogenesis was reduced in TKT flox/flox Alb-Cre mice compared with control littermates. Mechanistically, loss of TKT in the liver increased apoptosis, reduced cell proliferation, decreased TNFα, IL6, and STAT3 levels, and alleviated DEN/HFD-induced hepatic steatosis and fibrosis. Together, our data identify a key role for TKT in promoting genome instability during liver injury and tumor initiation. SIGNIFICANCE: These findings identify transketolase as a novel metabolic target to maintain genome stability and reduce liver carcinogenesis., (©2019 American Association for Cancer Research.)