Transcriptional control of a metabolic switch regulating cellular methylation reactions is part of a common response to stress in divergent bee species.

Recent global declines in bee health have elevated the need for a more complete understanding of the cellular stress mechanisms employed by diverse bee species. We recently uncovered the biomarker lethal (2) essential for life [l(2)efl] genes as part of a shared transcriptional program in response to a number of cell stressors in the western honey bee (Apis mellifera). Here, we describe another shared stress-responsive gene, glycine N-methyltransferase (Gnmt), which is known as a key metabolic switch controlling cellular methylation reactions. We observed Gnmt induction by both abiotic and biotic stressors. We also found increased levels of the GNMT reaction product sarcosine in the midgut after stress, linking metabolic changes with the observed changes in gene regulation. Prior to this study, Gnmt upregulation had not been associated with cellular stress responses in other organisms. To determine whether this novel stress-responsive gene would behave similarly in other bee species, we first characterized the cellular response to endoplasmic reticulum (ER) stress in lab-reared adults of the solitary alfalfa leafcutting bee (Megachile rotundata) and compared this with age-matched honey bees. The novel stress gene Gnmt was induced in addition to a number of canonical gene targets induced in both bee species upon unfolded protein response (UPR) activation, suggesting that stress-induced regulation of cellular methylation reactions is a common feature of bees. Therefore, this study suggests that the honey bee can serve as an important model for bee biology more broadly, although studies on diverse bee species will be required to fully understand global declines in bee populations.

The Glycine N-Methyltransferase Case Study: Another Challenge for QM-Cluster Models?

The methyl transfer reaction between SAM and glycine catalyzed by glycine N-methyltransferase (GNMT) was examined using QM-cluster models generated by Residue Interaction Network ResidUe Selector (RINRUS). RINRUS is a Python-based tool that can build QM-cluster models with rules-based processing of the active site residue interaction network. This way of enzyme model-building allows quantitative analysis of residue and fragment contributions to kinetic and thermodynamic properties of the enzyme. Many residue fragments are important for the GNMT catalytic reaction, such as Gly137, Asn138, and Arg175, which interact with the glycine substrate, and Trp30, Asp85, and Tyr242, which interact with the SAM cofactor. Our study shows that active site fragments that interact with the glycine substrate and the SAM cofactor must both be included in the QM-cluster models. Even though the proposed mechanism is a simple one-step reaction, GNMT may be a rather challenging case study for QM-cluster models because convergence in energetics requires models with >350 atoms. "Maximal" QM-cluster models built with either qualitative contact count ranking or quantitative interaction energies from functional group symmetry adapted perturbation theory provide acceptable results. Hence, important residue fragments that contribute to the energetics of the methyl-transfer reaction in GNMT are correctly identified in the RIN. Observations from this work suggest new directions to better establish an effective approach for constructing atomic-level enzyme models.

Folic Acid Ameliorates Renal Injury in Experimental Obstructive Nephropathy: Role of Glycine N-Methyltransferase.

Folic acid exerts both anti-inflammatory and antifibrotic effects. Glycine N-methyltransferase (GNMT), the major folic acid-binding protein in the liver, is a crucial enzyme that regulates the cellular methylation process by maintaining S-adenosylmethionine levels. However, as yet neither the therapeutic effects of folic acid in renal fibrosis nor whether GNMT is involved in these folic acid-associated mechanisms has been investigated. First, the expression of GNMT was examined in human kidneys with or without obstructive nephropathy. Later, wild-type and GNMT knockout (GNMT-/-) mice were subjected to unilateral ureteral obstruction (UUO) and then treated with either folic acid or vehicle for 14 days. Renal tubular injury, inflammation, fibrosis, and autophagy were evaluated by histological analysis and Western blotting. We observed increased expression of GNMT in humans with obstructive nephropathy. Furthermore, UUO significantly increased the expression of GNMT in mice; in addition, it caused renal injury as well as the development of both hydronephrosis and tubular injury. These were all alleviated by folic acid treatment. In contrast, GNMT-/- mice exhibited exacerbated UUO-induced renal injury, but the protective effect of folic acid was not observed in GNMT-/- mice. We propose a novel role for folic acid in the treatment of renal fibrosis, which indicates that GNMT may be a therapeutic target.

Glycine and aging: Evidence and mechanisms.

The restriction of calories, branched-chain amino acids, and methionine have all been shown to extend lifespan in model organisms. Recently, glycine was found to boost longevity in genetically heterogenous mice. This simple amino acid similarly extends lifespan in rats and improves health in mammalian models of age-related disease. While compelling data indicate that glycine is a pro-longevity molecule, divergent mechanisms may underlie its effects on aging. Glycine is abundant in collagen, a building block for glutathione, a precursor to creatine, and an acceptor for the enzyme glycine N-methyltransferase (GNMT). A review of the literature strongly implicates GNMT, which clears methionine from the body by taking a methyl group from S-adenosyl-L-methionine and methylating glycine to form sarcosine. In flies, Gnmt is required for reduced insulin/insulin-like growth factor 1 signaling and dietary restriction to fully extend lifespan. The geroprotector spermidine requires Gnmt to upregulate autophagy genes and boost longevity. Moreover, the overexpression of Gnmt is sufficient to extend lifespan and reduce methionine levels. Sarcosine, or methylglycine, declines with age in multiple species and is capable of inducing autophagy both in vitro and in vivo. Taken all together, existing evidence suggests that glycine prolongs life by mimicking methionine restriction and activating autophagy.

Reversal of High-Fat Diet-Induced Non-Alcoholic Fatty Liver Disease by Metformin Combined with PGG, an Inducer of Glycine N-Methyltransferase.

Nonalcoholic fatty liver disease (NAFLD) is a major cause of liver-related morbidities and mortality, and no effective drug treatment currently exists. We aimed to develop a novel treatment strategy to induce the expression of glycine N-methyltransferase (GNMT), which is an important enzyme regulating S-adenosylmethionine metabolism whose expression is downregulated in patients with NAFLD. Because 1,2,3,4,6-pentagalloyl glucose (PGG) is a GNMT inducer, and metformin was shown to upregulate liver mitochondrial GNMT protein expression, the effect of PGG and metformin was evaluated. Biochemical analysis, histopathological examination, immunohistochemical staining, reverse transcription-quantitative PCR (RT-qPCR), Western blotting (WB), proteomic analysis and Seahorse XF Cell Mito Stress Test were performed. The high-fat diet (HFD)-induced NAFLD mice were treated with PGG and metformin. Combination of PGG and metformin nearly completely reversed weight gain, elevation of serum aminotransferases, and hepatic steatosis and steatohepatitis. In addition, the downregulated GNMT expression in liver tissues of HFD-induced NAFLD mice was restored. The GNMT expression was further confirmed by RT-qPCR and WB analysis using both in vitro and in vivo systems. In addition, PGG treatment was shown to increase oxygen consumption rate (OCR) maximum capacity in a dose-dependent manner, and was capable of rescuing the suppression of mitochondrial OCR induced by metformin. Proteomic analysis identified increased expression of glutathione S-transferase mu 4 (GSTM4), heat shock protein 72 (HSP72), pyruvate carboxylase (PYC) and 40S ribosomal protein S28 (RS28) in the metformin plus PGG treatment group. Our findings show that GNMT expression plays an important role in the pathogenesis of NAFLD, and combination of an inducer of GNMT and metformin can be of therapeutic potential for patients with NAFLD.

Dysregulation of S-adenosylmethionine Metabolism in Nonalcoholic Steatohepatitis Leads to Polyamine Flux and Oxidative Stress.

Nonalcoholic fatty liver disease (NAFLD) is the number one cause of chronic liver disease worldwide, with 25% of these patients developing nonalcoholic steatohepatitis (NASH). NASH significantly increases the risk of cirrhosis and decompensated liver failure. Past studies in rodent models have shown that glycine-N-methyltransferase (GNMT) knockout results in rapid steatosis, fibrosis, and hepatocellular carcinoma progression. However, the attenuation of GNMT in subjects with NASH and the molecular basis for its impact on the disease process is still unclear. To address this knowledge gap, we show the reduction of GNMT protein levels in the liver of NASH subjects compared to healthy controls. To gain insight into the impact of decreased GNMT in the disease process, we performed global label-free proteome studies on the livers from a murine modified amylin diet-based model of NASH. Histological and molecular characterization of the animal model demonstrate a high resemblance to human disease. We found that a reduction of GNMT leads to a significant increase in S-adenosylmethionine (AdoMet), an essential metabolite for transmethylation reactions and a substrate for polyamine synthesis. Further targeted proteomic and metabolomic studies demonstrated a decrease in GNMT transmethylation, increased flux through the polyamine pathway, and increased oxidative stress production contributing to NASH pathogenesis.

Disrupted liver oxidative metabolism in glycine N-methyltransferase-deficient mice is mitigated by dietary methionine restriction.

OBJECTIVE: One-carbon metabolism is routinely dysregulated in nonalcoholic fatty liver disease. This includes decreased glycine N-methyltransferase (GNMT), a critical regulator of s-adenosylmethionine (SAM). Deletion of GNMT in mice increases SAM and promotes liver steatosis. Lower liver oxidative metabolism, as indicated by a decline in gluconeogenesis, citric acid cycle flux, and oxidative phosphorylation contributes to liver steatosis in GNMT-null mice; however, the extent to which higher SAM mediates this phenotype remains unclear. Here, we determined the SAM-dependent impairment in liver oxidative metabolism by loss of GNMT. METHODS: GNMT knockout (KO) mice were fed a methionine-restricted diet to prevent increased SAM. 2H/13C metabolic flux analysis was performed in conscious, unrestrained mice to quantify liver nutrient fluxes. Metabolomics and high-resolution respirometry were used to quantify liver nutrient pool sizes and mitochondrial oxidative phosphorylation, respectively. Folic acid-supplemented and serine/glycine-deficient diets were used independently to further define the metabolic implications of perturbed one-carbon donor availability. RESULTS: Dietary methionine restriction prevented a 75-fold increase in SAM and a 53% rise in triacylglycerides in livers of KO mice. Dietary methionine restriction increased gluconeogenesis, independent of genotype, and restored cytochrome c oxidase respiratory function in KO mice. Citric acid cycle fluxes remained lower in KO mice irrespective of diet. Restricting dietary methionine abrogated markers of increased lipogenesis and folate cycle dysfunction in KO mice. CONCLUSIONS: The impaired liver oxidative metabolism following loss of GNMT is both dependent and independent of greater SAM availability. Lower in vivo citric acid cycle flux is independent of increased SAM. In contrast, gluconeogenesis and oxidative phosphorylation are negatively regulated by excess SAM. Lipid accumulation in livers of mice lacking GNMT is also linked to higher SAM.

Downregulation of Methionine Cycle Genes MAT1A and GNMT Enriches Protein-Associated Translation Process and Worsens Hepatocellular Carcinoma Prognosis.

The major biological methyl donor, S-adenosylmethionine (adoMet) synthesis occurs mainly in the liver. Methionine adenosyltransferase 1A (MAT1A) and glycine N-methyltransferase (GNMT) are two key enzymes involved in the functional implications of that variation. We collected 42 RNA-seq data from paired hepatocellular carcinoma (HCC) and its adjacent normal liver tissue from the Cancer Genome Atlas (TCGA). There was no mutation found in MAT1A or GNMT RNA in the 42 HCC patients. The 11,799 genes were annotated in the RNA-Seq data, and their expression levels were used to investigate the phenotypes of low MAT1A and low GNMT by Gene Set Enrichment Analysis (GSEA). The REACTOME_TRANSLATION gene set was enriched and visualized in a heatmap along with corresponding differences in gene expression between low MAT1A versus high MAT1A and low GNMT versus high GNMT. We identified 43 genes of the REACTOME_TRANSLATION gene set that are powerful prognosis factors in HCC. The significantly predicted genes were referred into eukaryotic translation initiation (EIF3B, EIF3K), eukaryotic translation elongation (EEF1D), and ribosomal proteins (RPs). Cell models expressing various MAT1A and GNMT proved that simultaneous restoring the expression of MAT1A and GNMT decreased cell proliferation, invasion, as well as the REACTOME_TRANSLATION gene EEF1D, consistent with a better prognosis in human HCC. We demonstrated new findings that downregulation or defect in MAT1A and GNMT genes can enrich the protein-associated translation process that may account for poor HCC prognosis. This is the first study demonstrated that MAT1A and GNMT, the 2 key enzymes involved in methionine cycle, could attenuate the function of ribosome translation. We propose a potential novel mechanism by which the diminished GNMT and MAT1A expression may confer poor prognosis for HCC.

Discovery of an Orally Efficacious MYC Inhibitor for Liver Cancer Using a GNMT-Based High-Throughput Screening System and Structure-Activity Relationship Analysis.

Glycine-N-methyl transferase (GNMT) downregulation results in spontaneous hepatocellular carcinoma (HCC). Overexpression of GNMT inhibits the proliferation of liver cancer cell lines and prevents carcinogen-induced HCC, suggesting that GNMT induction is a potential approach for anti-HCC therapy. Herein, we used Huh7 GNMT promoter-driven screening to identify a GNMT inducer. Compound K78 was identified and validated for its induction of GNMT and inhibition of Huh7 cell growth. Subsequently, we employed structure-activity relationship analysis and found a potent GNMT inducer, K117. K117 inhibited Huh7 cell growth in vitro and xenograft in vivo. Oral administration of a dosage of K117 at 10 mpk (milligrams per kilogram) can inhibit Huh7 xenograft in a manner equivalent to the effect of sorafenib at a dosage of 25 mpk. A mechanistic study revealed that K117 is an MYC inhibitor. Ectopic expression of MYC using CMV promoter blocked K117-mediated MYC inhibition and GNMT induction. Overall, K117 is a potential lead compound for HCC- and MYC-dependent cancers.

Acute hypermethioninemia impairs redox homeostasis and acetylcholinesterase activity in the hippocampus, striatum, and cerebellum of young rats.

Hypermethioninemia is characterized by high plasma concentrations of methionine (Met) and its metabolites, such as methionine sulfoxide (MetO), and neurological changes, such as cerebral edema and cognitive deficits. The aim of this study was to analyze the redox status and acetylcholinesterase (AChE) activity in the hippocampus, striatum, and cerebellum of young Wistar rats subjected to an acute hypermethioninemia protocol. The animals received, by subcutaneous injection, a single dose of Met (0.4 g/kg), MetO (0.1 g/kg), and Met + MetO, and 1 or 3 hr after administration, the animals were euthanatized for brain structure obtaining. In the hippocampus, an increase in lipid peroxidation and glutathione peroxidase (GPx) activity was observed at 1 hr in the MetO and Met + MetO groups, and a reduction in the superoxide dismutase activity was found in the Met + MetO group. Met and/or MetO induced a decrease in the thiol content and GPx activity and enhanced the lipid peroxidation at 3 hr. In the striatum, a reduction in the thiol content and GPx activity, an increase in lipid peroxidation, and AChE activity were induced by Met and/or MetO at 1 or 3 hr. Additionally, in the cerebellum, an increase in the AChE in the MetO and Met + MetO groups 1 hr after administration was observed. These data help to better understand the pathophysiological mechanisms that underlie the neurological changes found in hypermethioninemia patients.

Cellular stress signaling activates type-I IFN response through FOXO3-regulated lamin posttranslational modification.

Neural stem/progenitor cells (NSPCs) persist over the lifespan while encountering constant challenges from age or injury related brain environmental changes like elevated oxidative stress. But how oxidative stress regulates NSPC and its neurogenic differentiation is less clear. Here we report that acutely elevated cellular oxidative stress in NSPCs modulates neurogenic differentiation through induction of Forkhead box protein O3 (FOXO3)-mediated cGAS/STING and type I interferon (IFN-I) responses. We show that oxidative stress activates FOXO3 and its transcriptional target glycine-N-methyltransferase (GNMT) whose upregulation triggers depletion of s-adenosylmethionine (SAM), a key co-substrate involved in methyl group transfer reactions. Mechanistically, we demonstrate that reduced intracellular SAM availability disrupts carboxymethylation and maturation of nuclear lamin, which induce cytosolic release of chromatin fragments and subsequent activation of the cGAS/STING-IFN-I cascade to suppress neurogenic differentiation. Together, our findings suggest the FOXO3-GNMT/SAM-lamin-cGAS/STING-IFN-I signaling cascade as a critical stress response program that regulates long-term regenerative potential.

Carnosine alleviates diabetic nephropathy by targeting GNMT, a key enzyme mediating renal inflammation and fibrosis.

Diabetic nephropathy (DN) is a common microvascular complication of diabetes and the main cause of end-stage nephropathy (ESRD). Inflammation and fibrosis play key roles in the development and progression of diabetic nephropathy. By using in vivo and in vitro DN models, our laboratory has identified the protective role of carnosine (CAR) on renal tubules. Our results showed that carnosine restored the onset and clinical symptoms as well as renal tubular injury in DN. Furthermore, carnosine decreased kidney inflammation and fibrosis in DN mice. These results were consistent with high glucose (HG)-treated mice tubular epithelial cells (MTECs). Using web-prediction algorithms, cellular thermal shift assay (CETSA) and molecular docking, we identified glycine N-methyltransferase (GNMT) as a carnosine target. Importantly, we found that GNMT, a multiple functional protein that regulates the cellular pool of methyl groups by controlling the ratio of S-adenosylmethionine (SAM) to S-adenosylhomocysteine (SAH), was down-regulated significantly in the serum of Type 1 DM patients and renal tissues of DN mice. Moreover, using cultured TECs, we confirmed that the increased GNMT expression by transient transfection mimicked the protective role of carnosine in reducing inflammation and fibrosis. Conversely, the inhibition of GNMT expression abolished the protective effects of carnosine. In conclusion, carnosine might serve as a promising therapeutic agent for DN and GNMT might be a potential therapeutic target for DN.

Ameliorative effect of tannic acid on hypermethioninemia-induced oxidative and nitrosative damage in rats: biochemical-based evidences in liver, kidney, brain, and serum.

We investigated the ability of tannic acid (TA) to prevent oxidative and nitrosative damage in the brain, liver, kidney, and serum of a rat model of acute hypermethioninemia. Young Wistar rats were divided into four groups: I (control), II (TA 30 mg/kg), III (methionine (Met) 0.4 g/kg + methionine sulfoxide (MetO) 0.1 g/kg), and IV (TA/Met + MetO). Rats in groups II and IV received TA orally for seven days, and rats of groups I and III received an equal volume of water. After pretreatment with TA, rats from groups II and IV received a single subcutaneous injection of Met + MetO, and were euthanized 3 h afterwards. In specific brain structures and the kidneys, we observed that Met + MetO led to increased reactive oxygen species (ROS), nitrite, and lipid peroxidation levels, followed by a reduction in thiol content and antioxidant enzyme activity. On the other hand, pretreatment with TA prevented both oxidative and nitrosative damage. In the serum, Met + MetO caused a decrease in the activity of antioxidant enzymes, which was again prevented by TA pretreatment. In contrast, in the liver, there was a reduction in ROS levels and an increase in total thiol content, which was accompanied by a reduction in catalase and superoxide dismutase activities in the Met + MetO group, and pretreatment with TA was able to prevent only the reduction in catalase activity. Conclusively, pretreatment with TA has proven effective in preventing oxidative and nitrosative changes caused by the administration of Met + MetO, and may thus represent an adjunctive therapeutic approach for treatment of hypermethioninemia.

Metabolism of Sulfur-Containing Amino Acids: How the Body Copes with Excess Methionine, Cysteine, and Sulfide.

Metabolism of excess methionine (Met) to homocysteine (Hcy) by transmethylation is facilitated by the expression of methionine adenosyltransferase (MAT) I/III and glycine N-methyltransferase (GNMT) in liver, and a lack of either enzyme results in hypermethioninemia despite normal concentrations of MATII and methyltransferases other than GNMT. The further metabolism of Hcy by the transsulfuration pathway is facilitated by activation of cystathionine ß-synthase (CBS) by S-adenosylmethionine (SAM) as well as the relatively high KM of CBS for Hcy. Transmethylation plus transsulfuration effects catabolism of the Met molecule along with transfer of the sulfur atom of Met to serine to synthesize cysteine (Cys). Oxidation and excretion of Met sulfur depend upon Cys catabolism and sulfur oxidation pathways. Excess Cys is oxidized by cysteine dioxygenase 1 (CDO1) and further metabolized to taurine or sulfate. Some Cys is normally metabolized by desulfhydration pathways, and the hydrogen sulfide (H2S) produced is further oxidized to sulfate. If Cys or Hcy concentrations are elevated, Cys or Hcy desulfhydration can result in excess H2S and thiosulfate production. Excess Cys or Met may also promote their limited metabolism by transamination pathways.

Targeted and untargeted metabolomics provide insight into the consequences of glycine-N-methyltransferase deficiency including the novel finding of defective immune function.

Fatty liver disease is increasing along with the prevalence of obesity and type-2 diabetes. Hepatic fibrosis is a major health complication for which there are no efficacious treatment options available. A better understanding of the fundamental mechanisms that contribute to the accumulation of fibrosis is needed. Glycine-N-methyltransferase (GNMT) is a critical enzyme in one-carbon metabolism that serves to regulate methylation and remethylation reactions. GNMT knockout (GNMT-/- ) mice display spontaneous hepatic fibrosis and later develop hepatocellular carcinoma. Previous literature supports the idea that hypermethylation as a consequence of GNMT deletion contributes to the hepatic phenotype observed. However, limited metabolomic information is available and the underlying mechanisms that contribute to hepatic fibrogenesis in GNMT-/- mice are still incomplete. Therefore, our goals were to use dietary intervention to determine whether increased lipid load exacerbates steatosis and hepatic fibrosis in this model and to employ both targeted and untargeted metabolomics to further understand the metabolic consequences of GNMT deletion. We find that GNMT mice fed high-fat diet do not accumulate more lipid or fibrosis in the liver and are in fact resistant to weight gain. Metabolomics analysis confirmed that pan-hypermethylation occurs in GNMT mice resulting in a depletion of nicotinamide intermediate metabolites. Further, there is a disruption in tryptophan catabolism that prevents adequate immune cell activation in the liver. The chronic cellular damage cannot be appropriately cleared due to a lack of immune checkpoint activation. This mouse model is an excellent example of how a disruption in small molecule metabolism can significantly impact immune function.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) dysregulates hepatic one carbon metabolism during the progression of steatosis to steatohepatitis with fibrosis in mice.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), a persistent environmental contaminant, induces steatosis that can progress to steatohepatitis with fibrosis, pathologies that parallel stages in the development of non-alcoholic fatty liver disease (NAFLD). Coincidently, one carbon metabolism (OCM) gene expression and metabolites are often altered during NAFLD progression. In this study, the time- and dose-dependent effects of TCDD were examined on hepatic OCM in mice. Despite AhR ChIP-seq enrichment at 2 h, OCM gene expression was not changed within 72 h following a bolus dose of TCDD. Dose-dependent repression of methionine adenosyltransferase 1A (Mat1a), adenosylhomocysteinase (Achy) and betaine-homocysteine S-methyltransferase (Bhmt) mRNA and protein levels following repeated treatments were greater at 28 days compared to 8 days. Accordingly, levels of methionine, betaine, and homocysteic acid were dose-dependently increased, while S-adenosylmethionine, S-adenosylhomocysteine, and cystathionine exhibited non-monotonic dose-dependent responses consistent with regulation by OCM intermediates and repression of glycine N-methyltransferase (Gnmt). However, the dose-dependent effects on SAM-dependent metabolism of polyamines and creatine could not be directly attributed to alterations in SAM levels. Collectively, these results demonstrate persistent AhR activation disrupts hepatic OCM metabolism at the transcript, protein and metabolite levels within context of TCDD-elicited progression of steatosis to steatohepatitis with fibrosis.

Blocking sphingosine 1-phosphate receptor 2 accelerates hepatocellular carcinoma progression in a mouse model of NASH.

The role of sphingosine 1-phosphate (S1P) and its sphingosine-1-phosphate receptors (S1PRs) in non-alcoholic steatohepatitis (NASH) is unclear. We aimed to analyze the role of S1P/S1PRs in a Melanocortin-4 receptor (Mc4r)-deficient NASH murine model using FTY720, the functional antagonist of S1PR1, S1PR3, S1PR4, and S1PR5, and JTE-013, the antagonist of S1PR2. We observed that, compared to that in the control, the mRNA of S1pr1 tended to decrease, whereas those of S1pr2 and S1pr3 significantly increased in Mc4r-knockout (KO) mice subjected to a Western diet (WD). While the fat area did not differ, fibrosis progression differed significantly between control mice and mice in which liver S1PRs were blocked. Lipidomic and metabolomic analysis of liver tissues showed that JTE-013-administered mice showed elevation of S-adenosyl-l-methionine level, which can induce aberrant methylation due to reduction in glycine N-methyltransferase (GNMT) and elevation in diacylglycerol (DG) and triacylglycerol (TG) levels, leading to increased susceptibility to hepatocellular carcinoma (HCC). These phenotypes are similar to those of Gnmt-KO mice, suggesting that blocking the S1P/S1PR2 axis triggers aberrant methylation, which may increase DG and TG, and hepatocarcinogenesis. Our observations that the S1P/S1PR2 axis averts HCC occurrence may assist in HCC prevention in NASH.

Hypermethioninemia induces memory deficits and morphological changes in hippocampus of young rats: implications on pathogenesis.

The aim of this study was to investigate the effect of the chronic administration of methionine (Met) and/or its metabolite, methionine sulfoxide (MetO), on the behavior and neurochemical parameters of young rats. Rats were treated with saline (control), Met (0.2-0.4 g/kg), MetO (0.05-0.1 g/kg), and/or a combination of Met + MetO, subcutaneously twice a day from postnatal day 6 (P6) to P28. The results showed that Met, MetO, and Met + MetO impaired short-term and spatial memories (P < 0.05), reduced rearing and grooming (P < 0.05), but did not alter locomotor activity (P > 0.05). Acetylcholinesterase activity was increased in the cerebral cortex, hippocampus, and striatum following Met and/or MetO (P < 0.05) treatment, while Na+, K+-ATPase activity was reduced in the hippocampus (P < 0.05). There was an increase in the level of thiobarbituric acid reactive substances (TBARS) in the cerebral cortex in Met-, MetO-, and Met + MetO-treated rats (P < 0.05). Met and/or MetO treatment reduced superoxide dismutase, catalase, and glutathione peroxidase activity, total thiol content, and nitrite levels, and increased reactive oxygen species and TBARS levels in the hippocampus and striatum (P < 0.05). Hippocampal brain-derived neurotrophic factor was reduced by MetO and Met + MetO compared with the control group. The number of NeuN-positive cells was decreased in the CA3 in Met + MetO group and in the dentate gyrus in the Met, MetO, and Met + MetO groups compared to control group (P < 0.05). Taken together, these findings further increase our understanding of changes in the brain in hypermethioninemia by elucidating behavioral alterations, biological mechanisms, and the vulnerability of brain function to high concentrations of Met and MetO.

Longevity in response to lowered insulin signaling requires glycine N-methyltransferase-dependent spermidine production.

Reduced insulin/IGF signaling (IIS) extends lifespan in multiple organisms. Different processes in different tissues mediate this lifespan extension, with a set of interplays that remain unclear. We here show that, in Drosophila, reduced IIS activity modulates methionine metabolism, through tissue-specific regulation of glycine N-methyltransferase (Gnmt), and that this regulation is required for full IIS-mediated longevity. Furthermore, fat body-specific expression of Gnmt was sufficient to extend lifespan. Targeted metabolomics showed that reducing IIS activity led to a Gnmt-dependent increase in spermidine levels. We also show that both spermidine treatment and reduced IIS activity are sufficient to extend the lifespan of Drosophila, but only in the presence of Gnmt. This extension of lifespan was associated with increased levels of autophagy. Finally, we found that increased expression of Gnmt occurs in the liver of liver-specific IRS1 KO mice and is thus an evolutionarily conserved response to reduced IIS. The discovery of Gnmt and spermidine as tissue-specific modulators of IIS-mediated longevity may aid in developing future therapeutic treatments to ameliorate aging and prevent disease.

miR-873-5p targets mitochondrial GNMT-Complex II interface contributing to non-alcoholic fatty liver disease.

OBJECTIVE: Non-alcoholic fatty liver disease (NAFLD) is a complex pathology in which several dysfunctions, including alterations in metabolic pathways, mitochondrial functionality and unbalanced lipid import/export, lead to lipid accumulation and progression to inflammation and fibrosis. The enzyme glycine N-methyltransferase (GNMT), the most important enzyme implicated in S-adenosylmethionine catabolism in the liver, is downregulated during NAFLD progression. We have studied the mechanism involved in GNMT downregulation by its repressor microRNA miR-873-5p and the metabolic pathways affected in NAFLD as well as the benefit of recovery GNMT expression. METHODS: miR-873-5p and GNMT expression were evaluated in liver biopsies of NAFLD/NASH patients. Different in vitro and in vivo NAFLD murine models were used to assess miR-873-5p/GNMT involvement in fatty liver progression through targeting of the miR-873-5p as NAFLD therapy. RESULTS: We describe a new function of GNMT as an essential regulator of Complex II activity in the electron transport chain in the mitochondria. In NAFLD, GNMT expression is controlled by miR-873-5p in the hepatocytes, leading to disruptions in mitochondrial functionality in a preclinical murine non-alcoholic steatohepatitis (NASH) model. Upregulation of miR-873-5p is shown in the liver of NAFLD/NASH patients, correlating with hepatic GNMT depletion. Importantly, NASH therapies based on anti-miR-873-5p resolve lipid accumulation, inflammation and fibrosis by enhancing fatty acid ß-oxidation in the mitochondria. Therefore, miR-873-5p inhibitor emerges as a potential tool for NASH treatment. CONCLUSION: GNMT participates in the regulation of metabolic pathways and mitochondrial functionality through the regulation of Complex II activity in the electron transport chain. In NAFLD, GNMT is repressed by miR-873-5p and its targeting arises as a valuable therapeutic option for treatment.