ATP Citrate Lyase is a General Tumour Biomarker and Contributes to the Development of Cutaneous Squamous Cell Carcinoma.

ATP citrate lyase, the first rate-limiting enzyme in de novo lipogenesis, plays a crucial role in tumour progression. This study explores ATP citrate lyase's potential as a tumour biomarker and its role in cutaneous squamous cell carcinoma. ATP citrate lyase expression patterns were analysed using TCGA and TIMER databases, and patient skin specimens were collected for immunohistochemistry to determine ATP citrate lyase levels. Cell proliferation, cell cycle, apoptosis, and c-Myc expression were assessed in A431 and SCL-1 cells. Stable cell lines with reduced ATP citrate lyase expression were obtained and subcutaneously implanted into nude mice to evaluate in vivo tumour growth. Ki67, c-Myc expression and TUNEL staining were analysed in subcutaneous tumours. ATP citrate lyase exhibited upregulation in various tumours, and showed significant associations with prognosis and immune infiltrate. Moreover, ATP citrate lyase was highly expressed in cutaneous squamous cell carcinoma. After ATP citrate lyase silencing, cutaneous squamous cell carcinoma cell growth decelerated, the cell cycle halted, cell apoptosis increased, and c-Myc expression decreased. Animal experiments revealed that, following ATP citrate lyase knockdown, tumour tissue growth slowed down, and there was a reduction in Ki-67 and c-Myc expression, accompanied by enhanced TUNEL staining. In conclusion, ATP citrate lyase may serve as a tumour biomarker. It is highly expressed in cutaneous squamous cell carcinoma and may serve as a therapeutic target.

Oxidized Low-Density Lipoprotein Accumulation in Macrophages Impairs Lipopolysaccharide-Induced Activation of AKT2, ATP Citrate Lyase, Acetyl-Coenzyme A Production, and Inflammatory Gene H3K27 Acetylation.

The accumulation of lipid and the formation of macrophage foam cells is a hallmark of atherosclerosis, a chronic inflammatory disease. To better understand the role of macrophage lipid accumulation in inflammation during atherogenesis, we studied early molecular events that follow the accumulation of oxidized low-density lipoprotein (oxLDL) in cultured mouse macrophages. We previously showed that oxLDL accumulation downregulates the inflammatory response in conjunction with downregulation of late-phase glycolysis. In this study, we show that within hours after LPS stimulation, macrophages with accumulated oxLDL maintain early-phase glycolysis but selectively downregulate activation of AKT2, one of three AKT isoforms. The inhibition of AKT2 activation reduced LPS-induced ATP citrate lyase activation, acetyl-CoA production, and acetylation of histone 3 lysine 27 (H3K27ac) in certain inflammatory gene promoters. In contrast to oxLDL, multiple early LPS-induced signaling pathways were inhibited in macrophages with accumulated cholesterol, including TBK1, AKT1, AKT2, MAPK, and NF-κB, and early-phase glycolysis. The selective inhibition of LPS-induced AKT2 activation was dependent on the generation of mitochondrial oxygen radicals during the accumulation of oxLDL in macrophages prior to LPS stimulation. This is consistent with increased oxidative phosphorylation, fatty acid synthesis, and oxidation pathways found by comparative transcriptomic analyses of oxLDL-loaded versus control macrophages. Our study shows a functional connection between oxLDL accumulation, inactivation of AKT2, and the inhibition of certain inflammatory genes through epigenetic changes that occur soon after LPS stimulation, independent of early-phase glycolysis.

[Expression and Clinical Significance of ATP Citrate Lyase in Hepatocellular Carcinoma].

Objective To investigate the role of ATP citrate lyase(ACLY)in the development of hepatocellular carcinoma(HCC)and the impact of this enzyme on the immune microenvironment of HCC.Methods We utilized the University of Alabama at Birmingham Cancer Data Analysis Portal and the Gene Expression Profiling Interactive Analysis to identify the changes in ACLY expression and prognosis across different tumor types from The Cancer Genome Atlas.With HCC as the disease model,we analyzed the ACLY expression in HCC samples from the gene expression database.Furthermore,we collected the clinical specimens from HCC patients to verify the mRNA and protein levels of ACLY.In addition,we conducted transcriptome sequencing after knocking down the expression of ACLY to analyze the differentially expressed genes and investigated the impact of ACLY expression interference on cell proliferation and other functions.Finally,we explored the correlations of ACLY with immune cells and immune infiltration in the tumor microenvironment,new antigens,and immune checkpoint genes.Results ACLY expression was significantly up-regulated in solid tumors including HCC(all P<0.05),and high ACLY expression was associated with overall survival rate in HCC(P=0.005).Furthermore,high ACLY expression affected the presence of immune cells(e.g.,tumor-associated fibroblasts)and the expression of genes involved in lipid metabolism(all P<0.05).Conclusions ACLY is closely related to the occurrence and development of HCC and lipid metabolism abnormalities.Moreover,it has a specific impact on the immune microenvironment of HCC.

Novel insights into the role of acetyl-CoA producing enzymes in epigenetic regulation.

Inflammation-dependent changes in gene expression programs in innate immune cells, such as macrophages, involve extensive reprogramming of metabolism. This reprogramming is essential for the production of metabolites required for chromatin modifications, such as acetyl-CoA, and regulate their usage and availability impacting the macrophage epigenome. One of the most transcriptionally induced proinflammatory mediator is nitric oxide (NO), which has been shown to inhibit key metabolic enzymes involved in the production of these metabolites. Recent evidence indicates that NO inhibits mitochondrial enzymes such as pyruvate dehydrogenase (PDH) in macrophages induced by inflammatory stimulus. PDH is involved in the production of acetyl-CoA, which is essential for chromatin modifications in the nucleus, such as histone acetylation. In addition, acetyl-CoA levels in inflamed macrophages are regulated by ATP citrate lyase (ACLY) and citrate transporter SLC25A1. Interestingly, acetyl-CoA producing enzymes, such as PDH and ACLY, have also been reported to be present in the nucleus and to support the local generation of cofactors such as acetyl-CoA. Here, we will discuss the mechanisms involved in the regulation of acetyl-CoA production by metabolic enzymes, their inhibition by prolonged exposure to inflammation stimuli, their involvement in dynamic inflammatory expression changes and how these emerging findings could have significant implications for the design of novel therapeutic approaches.

BCAT2 promotes melanoma progression by activating lipogenesis via the epigenetic regulation of FASN and ACLY expressions.

Melanoma is the most lethal skin cancer originating from the malignant transformation of epidermal melanocyte. The dysregulation of cellular metabolism is a hallmark of cancer, including in melanoma. Aberrant branched-chain amino acids (BCAA) metabolism and related enzymes has been greatly implicated in the progression of multiple types of cancer, whereas remains far from understood in melanoma. Herein, we reported that the critical BCAA metabolism enzyme branched-chain amino acid transaminase 2 (BCAT2) is an oncogenic factor in melanoma by activating lipogenesis via the epigenetic regulation of fatty acid synthase (FASN) and ATP-citrate lyase (ACLY) expressions. Firstly, we found that BCAT2 expression was prominently increased in melanoma, and highly associated with clinical stage. Then, it was proved that the deficiency of BCAT2 led to impaired tumor cell proliferation, invasion and migration in vitro, and tumor growth and metastasis in vivo. Further, RNA sequencing technology and a panel of biochemical assays demonstrated that BCAT2 regulated de novo lipogenesis via the regulation of the expressions of both FASN and ACLY. Mechanistically, the inhibition of BCAT2 suppressed the generation of intracellular acetyl-CoA, mitigating P300-dependent histone acetylation at the promoter of FASN and ACLY, and thereby their transcription. Ultimately, zinc finger E-box binding homeobox 1 (ZEB1) was identified as the upstream transcriptional factor responsible for BCAT2 up-regulation in melanoma. Our results demonstrate that BCAT2 promotes melanoma progression by epigenetically regulating FASN and ACLY expressions via P300-dependent histone acetylation. Targeting BCAT2 could be exploited as a promising strategy to restrain tumor progression in melanoma.

ACLY as a modulator of liver cell functions and its role in Metabolic Dysfunction-Associated Steatohepatitis.

BACKGROUND: Non-alcoholic Fatty Liver Disease (NAFLD), now better known as Metabolic (Dysfunction)-Associated Fatty Liver Disease (MAFLD) and its progression to Nonalcoholic Steatohepatitis (NASH), more recently referred to as Metabolic (Dysfunction)-Associated Steatohepatitis (MASH) are the most common causes of liver failure and chronic liver damage. The new names emphasize the metabolic involvement both in relation to liver function and pathological features with extrahepatic manifestations. This study aims to explore the role of the immunometabolic enzyme ATP citrate lyase (ACLY), with a critical function in lipogenesis, carbohydrate metabolism, gene expression and inflammation. METHODS: ACLY function was investigated in TNFα-triggered human hepatocytes and in PBMC-derived macrophages from MASH patients. Evaluation of expression levels was carried out by western blotting and/or RT-qPCR. In the presence or absence of ACLY inhibitors, ROS, lipid peroxidation and GSSG oxidative stress biomarkers were quantified. Chromatin immunoprecipitation (ChIP), transient transfections, immunocytochemistry, histone acetylation quantitation were used to investigate ACLY function in gene expression reprogramming. IL-6 and IL-1ß were quantified by Lumit immunoassays. RESULTS: Mechanistically, ACLY inhibition reverted lipid accumulation and oxidative damage while reduced secretion of inflammatory cytokines in TNFα-triggered human hepatocytes. These effects impacted not only on lipid metabolism but also on other crucial features of liver function such as redox status and production of inflammatory mediators. Moreover, ACLY mRNA levels together with those of malic enzyme 1 (ME1) increased in human PBMC-derived macrophages from MASH patients when compared to age-matched healthy controls. Remarkably, a combination of hydroxycitrate (HCA), the natural ACLY inhibitor, with red wine powder (RWP) significantly lowered ACLY and ME1 mRNA amount as well as IL-6 and IL-1ß production in macrophages from subjects with MASH. CONCLUSION: Collectively, our findings for the first time highlight a broad spectrum of ACLY functions in liver as well as in the pathogenesis of MASH and its diagnostic and therapeutic potential value.

ACL and HAT1 form a nuclear module to acetylate histone H4K5 and promote cell proliferation.

Acetyl-CoA utilized by histone acetyltransferases (HAT) for chromatin modification is mainly generated by ATP-citrate lyase (ACL) from glucose sources. How ACL locally establishes acetyl-CoA production for histone acetylation remains unclear. Here we show that ACL subunit A2 (ACLA2) is present in nuclear condensates, is required for nuclear acetyl-CoA accumulation and acetylation of specific histone lysine residues, and interacts with Histone AcetylTransferase1 (HAT1) in rice. The rice HAT1 acetylates histone H4K5 and H4K16 and its activity on H4K5 requires ACLA2. Mutations of rice ACLA2 and HAT1 (HAG704) genes impair cell division in developing endosperm, result in decreases of H4K5 acetylation at largely the same genomic regions, affect the expression of similar sets of genes, and lead to cell cycle S phase stagnation in the endosperm dividing nuclei. These results indicate that the HAT1-ACLA2 module selectively promotes histone lysine acetylation in specific genomic regions and unravel a mechanism of local acetyl-CoA production which couples energy metabolism with cell division.

BCKDHA contributes to melanoma progression by promoting the expressions of lipogenic enzymes FASN and ACLY.

The dysregulation of branched-chain amino acid (BCAA) metabolism and related enzymes has been greatly implicated in the progression of multiple types of cancer, whereas remains far from understood in melanoma. Here, we explored the role of the BCAA metabolism enzyme BCKDHA in melanoma pathogenesis and elucidated the underlying mechanisms. In vitro cell biology experiments and in vivo pre-clinical mice model experiments were performed to investigate the role of BCKDHA in melanoma progression. RNA sequencing, immunohistochemical/immunofluorescence staining and bioinformatics analysis were used to examine the underlying mechanism. BCKDHA expression was prominently increased in both melanoma tissues and cell lines. The up-regulation of BCKDHA promoted long-term tumour cell proliferation, invasion and migration in vitro and tumour growth in vivo. Through RNA-sequencing technology, it was found that BCKDHA regulated the expressions of lipogenic fatty acid synthase (FASN) and ATP-citrate lyase (ACLY), which was thereafter proved to mediate the oncogenic role of BCKDHA in melanoma. Our results demonstrate that BCKDHA promotes melanoma progression by regulating FASN and ACLY expressions. Targeting BCKDHA could be exploited as a promising strategy to restrain tumour progression in melanoma.

Inter-organelle cross-talk supports acetyl-coenzyme A homeostasis and lipogenesis under metabolic stress.

Proliferating cells rely on acetyl-CoA to support membrane biogenesis and acetylation. Several organelle-specific pathways are available for provision of acetyl-CoA as nutrient availability fluctuates, so understanding how cells maintain acetyl-CoA homeostasis under such stresses is critically important. To this end, we applied 13C isotope tracing cell lines deficient in these mitochondrial [ATP-citrate lyase (ACLY)]-, cytosolic [acetyl-CoA synthetase (ACSS2)]-, and peroxisomal [peroxisomal biogenesis factor 5 (PEX5)]-dependent pathways. ACLY knockout in multiple cell lines reduced fatty acid synthesis and increased reliance on extracellular lipids or acetate. Knockout of both ACLY and ACSS2 (DKO) severely stunted but did not entirely block proliferation, suggesting that alternate pathways can support acetyl-CoA homeostasis. Metabolic tracing and PEX5 knockout studies link peroxisomal oxidation of exogenous lipids as a major source of acetyl-CoA for lipogenesis and histone acetylation in cells lacking ACLY, highlighting a role for inter-organelle cross-talk in supporting cell survival in response to nutrient fluctuations.

Allosteric role of the citrate synthase homology domain of ATP citrate lyase.

ATP citrate lyase (ACLY) is the predominant nucleocytosolic source of acetyl-CoA and is aberrantly regulated in many diseases making it an attractive therapeutic target. Structural studies of ACLY reveal a central homotetrameric core citrate synthase homology (CSH) module flanked by acyl-CoA synthetase homology (ASH) domains, with ATP and citrate binding the ASH domain and CoA binding the ASH-CSH interface to produce acetyl-CoA and oxaloacetate products. The specific catalytic role of the CSH module and an essential D1026A residue contained within it has been a matter of debate. Here, we report biochemical and structural analysis of an ACLY-D1026A mutant demonstrating that this mutant traps a (3S)-citryl-CoA intermediate in the ASH domain in a configuration that is incompatible with the formation of acetyl-CoA, is able to convert acetyl-CoA and OAA to (3S)-citryl-CoA in the ASH domain, and can load CoA and unload acetyl-CoA in the CSH module. Together, this data support an allosteric role for the CSH module in ACLY catalysis.

Active post-transcriptional regulation and ACLY-mediated acetyl-CoA synthesis as a pivotal target of Shuang-Huang-Sheng-Bai formula for lung adenocarcinoma treatment.

BACKGROUND: New therapeutic approaches are required to improve the outcomes of lung cancer (LC), a leading cause of cancer-related deaths worldwide. Chinese herbal medicine formulae widely used in China provide a unique opportunity for improving LC treatment, and the Shuang-Huang-Sheng-Bai (SHSB) formula is a typical example. However, the underlying mechanisms of action remains unclear. PURPOSE: This study aimed to confirm the efficacy of SHSB against lung adenocarcinoma (LUAD), which is a major histological type of LC, unveil the downstream targets of this formula, and assess the clinical relevance and biological roles of the newly identified target. METHODS: An experimental metastasis mouse model and a subcutaneous xenograft mouse model were used to evaluate the anti-cancer activity of SHSB. Multi-omics profiling of subcutaneous tumors and metabolomic profiling of sera were performed to identify downstream targets, especially the metabolic targets of SHSB. A clinical trial was conducted to verify the newly identified metabolic targets in patients. Next, the metabolites and enzymes engaged in the metabolic pathway targeted by SHSB were measured in clinical samples. Finally, routine molecular experiments were performed to decipher the biological functions of the metabolic pathways targeted by SHSB. RESULTS: Oral SHSB administration showed overt anti-LUAD efficacy as revealed by the extended overall survival of the metastasis model and impaired growth of implanted tumors in the subcutaneous xenograft model. Mechanistically, SHSB administration altered protein expression in the post-transcriptional layer and modified the metabolome of LUAD xenografts. Integrative analysis demonstrated that SHSB markedly inhibited acetyl-CoA synthesis in tumors by post-transcriptionally downregulating ATP-citrate lyase (ACLY). Consistently, our clinical trial showed that oral SHSB administration declined serum acetyl-CoA levels of patients with LC. Moreover, acetyl-CoA synthesis and ACLY expression were both augmented in clinical LUAD tissues of patients, and high intratumoral ACLY expression predicted a detrimental prognosis. Finally, we showed that ACLY-mediated acetyl-CoA synthesis is essential for LUAD cell growth by promoting G1/S transition and DNA replication. CONCLUSION: Limited downstream targets of SHSB for LC treatment have been reported in previous hypothesis-driven studies. In this study, we conducted a comprehensive multi-omics investigation and demonstrated that SHSB exerted its anti-LUAD efficacy by actively and post-transcriptionally modulating protein expression and particularly restraining ACLY-mediated acetyl-CoA synthesis.

Cytoskeletal association of ATP citrate lyase controls the mechanodynamics of macropinocytosis.

Macropinocytosis is an actin-dependent mode of nonselective endocytosis that mediates the uptake of extracellular fluid-phase cargoes. It is now well recognized that tumor cells exploit macropinocytosis to internalize macromolecules that can be catabolized and used to support cell growth and proliferation under nutrient-limiting conditions. Therefore, the identification of molecular mechanisms that control macropinocytosis is fundamental to the understanding of the metabolic adaptive landscape of tumor cells. Here, we report that the acetyl-CoA-producing enzyme, ATP citrate lyase (ACLY), is a key regulator of macropinocytosis and describes a heretofore-unappreciated association of ACLY with the actin cytoskeleton. The cytoskeletal tethering of ACLY is required for the spatially defined acetylation of heterodimeric actin capping protein, which we identify as an essential mediator of the actin remodeling events that drive membrane ruffling and macropinocytosis. Furthermore, we identify a requirement for mitochondrial-derived citrate, an ACLY substrate, for macropinocytosis, and show that mitochondria traffic to cell periphery regions juxtaposed to plasma membrane ruffles. Collectively, these findings establish a mode of metabolite compartmentalization that supports the spatiotemporal modulation of membrane-cytoskeletal interactions required for macropinocytosis by coupling regional acetyl-CoA availability with dynamic protein acetylation.

Acly Deficiency Enhances Myelopoiesis through Acetyl Coenzyme A and Metabolic-Epigenetic Cross-Talk.

Hematopoiesis integrates cytokine signaling, metabolism, and epigenetic modifications to regulate blood cell generation. These processes are linked, as metabolites provide essential substrates for epigenetic marks. In this study, we demonstrate that ATP citrate lyase (Acly), which metabolizes citrate to generate cytosolic acetyl-CoA and is of clinical interest, can regulate chromatin accessibility to limit myeloid differentiation. Acly was tested for a role in murine hematopoiesis by small-molecule inhibition or genetic deletion in lineage-depleted, c-Kit-enriched hematopoietic stem and progenitor cells from Mus musculus. Treatments increased the abundance of cell populations that expressed the myeloid integrin CD11b and other markers of myeloid differentiation. When single-cell RNA sequencing was performed, we found that Acly inhibitor-treated hematopoietic stem and progenitor cells exhibited greater gene expression signatures for macrophages and enrichment of these populations. Similarly, the single-cell assay for transposase-accessible chromatin sequencing showed increased chromatin accessibility at genes associated with myeloid differentiation, including CD11b, CD11c, and IRF8. Mechanistically, Acly deficiency altered chromatin accessibility and expression of multiple C/EBP family transcription factors known to regulate myeloid differentiation and cell metabolism, with increased Cebpe and decreased Cebpa and Cebpb. This effect of Acly deficiency was accompanied by altered mitochondrial metabolism with decreased mitochondrial polarization but increased mitochondrial content and production of reactive oxygen species. The bias to myeloid differentiation appeared due to insufficient generation of acetyl-CoA, as exogenous acetate to support alternate compensatory pathways to produce acetyl-CoA reversed this phenotype. Acly inhibition thus can promote myelopoiesis through deprivation of acetyl-CoA and altered histone acetylome to regulate C/EBP transcription factor family activity for myeloid differentiation.

The structure of succinyl-CoA synthetase bound to the succinyl-phosphate intermediate clarifies the catalytic mechanism of ATP-citrate lyase.

Succinyl-CoA synthetase (SCS) catalyzes a three-step reaction in the citric acid cycle with succinyl-phosphate proposed as a catalytic intermediate. However, there are no structural data to show the binding of succinyl-phosphate to SCS. Recently, the catalytic mechanism underlying acetyl-CoA production by ATP-citrate lyase (ACLY) has been debated. The enzyme belongs to the family of acyl-CoA synthetases (nucleoside diphosphate-forming) for which SCS is the prototype. It was postulated that the amino-terminal portion catalyzes the full reaction and the carboxy-terminal portion plays only an allosteric role. This interpretation was based on the partial loss of the catalytic activity of ACLY when Glu599 was mutated to Gln or Ala, and on the interpretation that the phospho-citryl-CoA intermediate was trapped in the 2.85 Šresolution structure from cryogenic electron microscopy (cryo-EM). To better resolve the structure of the intermediate bound to the E599Q mutant, the equivalent mutation, E105αQ, was made in human GTP-specific SCS. The structure of the E105αQ mutant shows succinyl-phosphate bound to the enzyme at 1.58 Šresolution when the mutant, after phosphorylation in solution by Mg2+-ATP, was crystallized in the presence of magnesium ions, succinate and desulfo-CoA. The E105αQ mutant is still active but has a specific activity that is 120-fold less than that of the wild-type enzyme, with apparent Michaelis constants for succinate and CoA that are 50-fold and 11-fold higher, respectively. Based on this high-resolution structure, the cryo-EM maps of the E599Q ACLY complex reported previously should have revealed the binding of citryl-phosphate and CoA and not phospho-citryl-CoA.

Inhibiting glutamine utilization creates a synthetic lethality for suppression of ATP citrate lyase in KRas-driven cancer cells.

Metabolic reprogramming is now considered a hallmark of cancer cells. KRas-driven cancer cells use glutaminolysis to generate the tricarboxylic acid cycle intermediate α-ketoglutarate via a transamination reaction between glutamate and oxaloacetate. We reported previously that exogenously supplied unsaturated fatty acids could be used to synthesize phosphatidic acid-a lipid second messenger that activates both mammalian target of rapamycin (mTOR) complex 1 (mTORC1) and mTOR complex 2 (mTORC2). A key target of mTORC2 is Akt-a kinase that promotes survival and regulates cell metabolism. We report here that mono-unsaturated oleic acid stimulates the phosphorylation of ATP citrate lyase (ACLY) at the Akt phosphorylation site at S455 in an mTORC2 dependent manner. Inhibition of ACLY in KRas-driven cancer cells in the absence of serum resulted in loss of cell viability. We examined the impact of glutamine (Gln) deprivation in combination with inhibition of ACLY on the viability of KRas-driven cancer cells. While Gln deprivation was somewhat toxic to KRas-driven cancer cells by itself, addition of the ACLY inhibitor SB-204990 increased the loss of cell viability. However, the transaminase inhibitor aminooxyacetate was minimally toxic and the combination of SB-204990 and aminooxtacetate led to significant loss of cell viability and strong cleavage of poly-ADP ribose polymerase-indicating apoptotic cell death. This effect was not observed in MCF7 breast cancer cells that do not have a KRas mutation or in BJ-hTERT human fibroblasts which have no oncogenic mutation. These data reveal a synthetic lethality between inhibition of glutamate oxaloacetate transaminase and ACLY inhibition that is specific for KRas-driven cancer cells and the apparent metabolic reprogramming induced by activating mutations to KRas.

Paradoxical activation of transcription factor SREBP1c and de novo lipogenesis by hepatocyte-selective ATP-citrate lyase depletion in obese mice.

Hepatic steatosis associated with high-fat diet, obesity, and type 2 diabetes is thought to be the major driver of severe liver inflammation, fibrosis, and cirrhosis. Cytosolic acetyl CoA (AcCoA), a central metabolite and substrate for de novo lipogenesis (DNL), is produced from citrate by ATP-citrate lyase (ACLY) and from acetate through AcCoA synthase short chain family member 2 (ACSS2). However, the relative contributions of these two enzymes to hepatic AcCoA pools and DNL rates in response to high-fat feeding are unknown. We report here that hepatocyte-selective depletion of either ACSS2 or ACLY caused similar 50% decreases in liver AcCoA levels in obese mice, showing that both pathways contribute to the generation of this DNL substrate. Unexpectedly however, the hepatocyte ACLY depletion in obese mice paradoxically increased total DNL flux measured by D2O incorporation into palmitate, whereas in contrast, ACSS2 depletion had no effect. The increase in liver DNL upon ACLY depletion was associated with increased expression of nuclear sterol regulatory element-binding protein 1c and of its target DNL enzymes. This upregulated DNL enzyme expression explains the increased rate of palmitate synthesis in ACLY-depleted livers. Furthermore, this increased flux through DNL may also contribute to the observed depletion of AcCoA levels because of its increased conversion to malonyl CoA and palmitate. Together, these data indicate that in fat diet-fed obese mice, hepatic DNL is not limited by its immediate substrates AcCoA or malonyl CoA but rather by activities of DNL enzymes.

Dual role of pseudogene TMEM198B in promoting lipid metabolism and immune escape of glioma cells.

Dysregulation of pseudogenes, enhancement of fatty acid synthesis and formation of immunosuppressive microenvironment are important factors that promote the malignant progression of glioma. It is of great significance to search for the molecular mechanism of interaction between the three and then perform targeted interference for improving the treatment of glioma. In this study, we found that pseudogene transmembrane protein 198B (TMEM198B) was highly expressed in glioma tissues and cell lines, and it could promote malignant progression of glioma by regulating lipid metabolism reprogramming and remodeling immune microenvironment. Applying the experimental methods of gene interference, lipidomics and immunology, we further confirmed that TMEM198B promoted PLAG1 like zinc finger 2 (PLAGL2) expression by mediating tri-methylation of histone H3 on lysine 4 (H3K4me3) of PLAGL2 through binding to SET domain containing 1B (SETD1B). Increased PLAGL2 could transcriptional activate ATP citrate lyase (ACLY) and ELOVL fatty acid elongase 6 (ELOVL6) expression, and then influenced the biological behaviors of glioma cells via enhancing the de novo lipogenesis and fatty acid acyl chain elongation. At the same time, TMEM198B promoted macrophages lipid accumulation and intensification of fatty acid oxidation (FAO) through glioma-derived exosomes (GDEs), further induced macrophages to M2 polarization, which subsequently facilitated immune escape of glioma cells. In conclusion, our present study clarifies that the TMEM198B/PLAGL2/ACLY/ELOVL6 pathway conducts crucial regulatory effects on the malignant progression of glioma, which provides novel targets and new ideas for molecular targeted therapy and immunotherapy of glioma.

LPS stimulation stabilizes HIF-1α by enhancing HIF-1α acetylation via the PARP1-SIRT1 and ACLY-Tip60 pathways in macrophages.

Hypoxia and inflammatory mediators stabilize hypoxia-inducible factor (HIF)-1α through posttranslational modifications, such as phosphorylation and succinylation. Here, we identified sirtuin 1 (SIRT1) and 60 kDa Tat-interactive protein (Tip60)-mediated acetylation as another critical posttranslational modification that regulates HIF-1α protein stability under lipopolysaccharide (LPS) stimulation. Mechanistically, DNA damage induced by excessive reactive oxygen species (ROS) activated poly (ADP-ribose) polymerase 1 (PARP1) to consume oxidized nicotinamide adenine dinucleotide (NAD+ ). Correspondingly, SIRT1 activity was decreased with the decline in NAD+ levels, resulting in increased HIF-1α acetylation. LPS also activated the ATP-citrate lyase (ACLY)-Tip60 pathway to further enhance HIF-1α acetylation. Acetylation contributed to HIF-1α stability and exacerbated LPS-induced inflammation. Thus, inhibiting HIF-1α stability by decreasing its acetylation could partly alleviate LPS-induced inflammation. In conclusion, we revealed the mechanism by which LPS stabilized HIF-1α by increasing its acetylation via the PARP1-SIRT1 and ACLY-Tip60 pathways in fish macrophages. This study may provide novel insights for manipulation of HIF-1α acetylation as a therapeutic strategy against inflammation from the perspective of acetylation in vertebrates.

Deletion of ACLY Disrupts Histone Acetylation and IL-10 Secretion in Trophoblasts, Which Inhibits M2 Polarization of Macrophages: A Possible Role in Recurrent Spontaneous Abortion.

Changes to macrophage polarization affect the local microenvironment of the placenta, resulting in pathological pregnancy diseases such as recurrent spontaneous abortion (RSA). Macrophages are in close contact with trophoblasts during placental development, and trophoblast-derived cytokines are important regulators of macrophage polarization and function. Histone acetylation can affect the expression and secretion of cytokines, and ATP citrate lyase (ACLY) is an important factor that regulates histone acetylation. The aim of this study was to investigate the effect of ACLY expression differences in trophoblast on macrophage polarization and its mechanism. Our data demonstrate that ACLY level in placental villi of patients with RSA is decreased, which may lead to the inhibition of histone acetylation in trophoblasts, thereby reducing the secretion of IL-10. Reduced IL-10 secretion activates endoplasmic reticulum stress in macrophages, thus inhibiting their M2 polarization.

A non-canonical tricarboxylic acid cycle underlies cellular identity.

The tricarboxylic acid (TCA) cycle is a central hub of cellular metabolism, oxidizing nutrients to generate reducing equivalents for energy production and critical metabolites for biosynthetic reactions. Despite the importance of the products of the TCA cycle for cell viability and proliferation, mammalian cells display diversity in TCA-cycle activity1,2. How this diversity is achieved, and whether it is critical for establishing cell fate, remains poorly understood. Here we identify a non-canonical TCA cycle that is required for changes in cell state. Genetic co-essentiality mapping revealed a cluster of genes that is sufficient to compose a biochemical alternative to the canonical TCA cycle, wherein mitochondrially derived citrate exported to the cytoplasm is metabolized by ATP citrate lyase, ultimately regenerating mitochondrial oxaloacetate to complete this non-canonical TCA cycle. Manipulating the expression of ATP citrate lyase or the canonical TCA-cycle enzyme aconitase 2 in mouse myoblasts and embryonic stem cells revealed that changes in the configuration of the TCA cycle accompany cell fate transitions. During exit from pluripotency, embryonic stem cells switch from canonical to non-canonical TCA-cycle metabolism. Accordingly, blocking the non-canonical TCA cycle prevents cells from exiting pluripotency. These results establish a context-dependent alternative to the traditional TCA cycle and reveal that appropriate TCA-cycle engagement is required for changes in cell state.