Thiolase Research Articles

A rare case of β-ketothiolase deficiency presenting as mimicker of diabetic ketoacidosis

β-ketothiolase (3-oxothiolase, BKT), also called mitochondrial acetoacetyl-coenzyme-A thiolase (T2), is a mitochondrial enzyme involved in isoleucine catabolism and ketone metabolism. BKT or T2 deficiency is inherited as an autosomal recessive trait and results in a rare inborn error of metabolism called alpha-acetoacetic aciduria. Patients with this disorder usually present with intermittent attacks of ketoacidosis. Here, we report a case of BKT deficiency that mimicked diabetic ketoacidosis (DKA) at presentation and discuss the clinical profile and interpretation of laboratory findings. An 8-month-old male child presented with seizures, coma, hyperglycemia, shock, and high anion gap metabolic acidosis with normal serum lactate and negative urinary ketone requiring correction with sodium bicarbonate, management of shock and sepsis, and mechanical ventilation. During his stay in the hospital, he developed ketonuria and episodes of hypoglycemia on a glucose insulin infusion with persistent metabolic acidosis which prompted us to look for an inborn error of metabolism, as hemoglobin A1C (HbA1C) and C-peptide levels were normal. Both tandem mass spectrometry (TMS) and urinary gas chromatography mass spectrometry (GCMS) were confirmatory. Diagnosis of BKT deficiency was further confirmed with genetic testing which revealed a novel homozygous mutation in the acetyl-CoA acetyltransferase 1 (ACAT1) gene. In infants manifesting with clinical features of diabetic ketoacidosis (DKA), cautious interpretation of laboratory findings and consideration of inborn errors of metabolism including the rare BKT deficiency are needed for a favorable outcome and prediction of prognosis.

ACAT1 deficiency in myeloid cells promotes glioblastoma progression by enhancing the accumulation of myeloid-derived suppressor cells

Glioblastoma (GBM) is a highly aggressive and lethal brain tumor with an immunosuppressive tumor microenvironment (TME). In this environment, myeloid cells, such as myeloid-derived suppressor cells (MDSCs), play a pivotal role in suppressing antitumor immunity. Lipometabolism is closely related to the function of myeloid cells. Here, our study reports that acetyl-CoA acetyltransferase 1 (ACAT1), the key enzyme of fatty acid oxidation (FAO) and ketogenesis, is significantly downregulated in the MDSCs infiltrated in GBM patients. To investigate the effects of ACAT1 on myeloid cells, we generated mice with myeloid-specific (LyzM-cre) depletion of ACAT1. The results show that these mice exhibited a remarkable accumulation of MDSCs and increased tumor progression both ectopically and orthotopically. The mechanism behind this effect is elevated secretion of C–X–C motif ligand 1 (CXCL1) of macrophages (Mφ). Overall, our findings demonstrate that ACAT1 could serve as a promising drug target for GBM by regulating the function of MDSCs in the TME.

Adaptive responses and metabolic strategies of Novosphingobium sp. ES2-1–17β-estradiol analyzed through integration of genomic and proteomic approaches

Environmental 17β-estradiol (E2) can cause potential harm to ecological balance and human health. Novosphingobium sp. ES2–1 is an E2-degrading bacterium previously obtained, which converts E2 to estrone (E1) and then to 4-hydroxyestrone (4-OH-E1) followed by oxidation to form metabolites with long-chain structure during upstream degradation. Herein, we found that intracellular enzymes were the major contributors to E2 biodegradation by strain ES2–1. A total of 243 proteins were dys-expressed under E2 condition, 123 were up-regulated and 120 were down-regulated thereinto. The up-regulated members of ABC transport systems, aromatics degradation, and fatty acid degradation indicated a reinforced transfer and utilization of E2. Cytochrome P450 monooxygenase (EstP1), 2-keto-4-pentenoate hydratase, pyruvate dehydrogenase, acetyl-CoA acetyltransferase, TonB-dependent receptor were involved in E2 catabolism. During downstream degradation, the metabolites with long-chain structure were decomposed adopting β-oxidation pattern and ultimately entered the TCA cycle; 2-keto-4-pentenoic acid might be an emblematic product of such process. Furthermore, E2 converting to E1 was catalyzed by 17β-dehydrogenase probably encoded by IM701_16645 or IM701_16910; 4-OH-E1 meta-cleavage was catalyzed by a dioxygenase encoded by IM701_20340 or IM701_21000 or IM701_09625. Our study provided an in-depth insight into the adaptive responses and metabolic strategies of Novosphingobium to E2.

Role of acetyl-CoA acetyltransferase 1 expression in the molecular mechanism of adenomyosis.

Adenomyosis is a benign uterine illness characterized by endometrial gland and stromal invasion into the myometrium. Acetyl-CoA acetyltransferase 1 (ACAT1) is an enzyme localized in mitochondria that is involved in ketogenesis and ketolysis processes by reversibly catalyzing the formation of acetoacetyl-CoA from two acetyl-CoA molecules. The current study investigated the expression of the ACAT1 molecule in tissue samples of patients diagnosed with adenomyosis and healthy endometrial tissues. It is aimed to determine the differences in ACAT1 gene expression and in this way to discover the first information about the role of ACAT1 in the development and molecular mechanism of adenomyosis. In the current retrospective study, formalin-fixed paraffin-embedded archival tissues were employed. A total of 76 patient samples were included in the study. Of these samples, 28 are adenomyotic tissue (Group I), 30 are eutopic endometrial tissue (Group II), and 18 are the Control Group. In these groups, the expression levels of the ACAT1 gene were determined by the reverse transcription-polymerase chain reaction method. When the expression results of the ACAT1 gene were evaluated, statistically significant differences were found between the groups (p<0.05). There was a difference between Group I-Group II and Group I-Control Group regarding the ACAT1 gene. No statistically significant change was observed between Group II and Control Group. It is a remarkable finding that the expression of ACAT1 in adenomyosis tissue is decreased compared with both eutopic endometrium and control groups tissues. The results suggest that ACAT1 may be associated with the molecular pathogenesis of adenomyosis.

The modulatory effects on enterohepatic cholesterol metabolism of novel cholesterol-lowering peptides from gastrointestinal digestion of Xuanwei ham

The aim of this study was to investigate the effects and mechanism of in vitro protein digestive products of Xuanwei ham with different ripening periods on cholesterol metabolism and hypercholesterolemia. The results showed that compared with other gastrointestinal digestion (GID) groups, the GID group of Xuanwei ham with 3-year ripening period (XWH3-GID) inhibited the expression of Niemann-Pick C1-like 1 (NPC1L1) and acetyl-CoA acetyltransferase 2 (ACAT2) through hepatocyte nuclear factor 1-alpha (HNF-1α), which in turn effectively inhibited cholesterol absorption in Caco-2 cell monolayers. Following absorption by Caco-2 cell monolayers, the XWH3-GID group suppressed the expression and secretion of proprotein convertase subtilisin/kexin type 9 (PCSK9) via HNF-1α, which enhanced the protein expression and fluorescence intensity of low density lipoprotein receptor (LDLR) on the HepG2 cell membrane, and thus promoted the uptake of low density lipoprotein (LDL). Importantly, three novel peptides (LFP, PKF and VPFP) derived from titin were identified after intestinal epithelial transport in the XWH3-GID group, which could exert cholesterol-lowering effects through inhibiting intestinal cholesterol absorption and promoting peripheral hepatic LDL uptake, and effectively ameliorate western diet-induced hypercholesterolemia in ApoE-/- mice. These results suggest that Xuanwei ham with 3-year ripening period can be used as a source of cholesterol-lowering peptides and has potential to intervene in hypercholesterolemia.

Acetyl-coenzyme A acetyltransferase 1 promotes brown adipogenesis by activating the AMPK-PGC1α signaling pathway

Brown adipose tissue (BAT) is thermogenic, expressing high levels of uncoupling protein-1 to convert nutrient energy to heat energy, bypassing ATP synthesis. BAT is a promising therapeutic target for treatment of obesity and type 2 diabetes since it converts fatty acids into heat but mechanisms controlling brown adipogenesis remain unclear. Knockdown of acetyl-Coenzyme A acetyltransferase 1 (ACAT1) in C3H10T1/2 cells suppressed brown adipocyte maturation during the current study and ACAT1 overexpression promoted brown adipocyte maturation. The downstream target of AMP-activated protein kinase (AMPK), peroxisome proliferator-activated receptor gamma coactivator-1-α (PGC1α), was involved in the action of ACAT1 on brown adipocyte maturation. ACAT1 overexpression enhanced AMPK phosphorylation and promoted PGC1α expression. It is suggested that ACAT1 promotes brown adipocyte maturation by activating the AMPK-PGC1α signaling pathway.

Insights into the therapeutic outcomes of trimetazidine/doxorubicin combination in Ehrlich solid-phase carcinoma mouse tumor model

One of the key features of cancer is metabolic reprogramming that can be exploited to sensitize cancer cells to chemotherapy. Trimetazidine (TMZ) is a metabolic anti-ischemic drug that blocks the activity of long-chain 3-ketoacyl CoA thiolase leading to the inhibition of fatty acid oxidation. AimsThe objective of the current investigation was to evaluate the idea that TMZ could synergize the antitumor activity of doxorubicin (DOX). Main methodsThe hypothesis was examined in vitro using the human breast cancer cell lines MCF-7 and MDA-MB231. In addition, the in vivo experiments were conducted using the Ehrlich solid phase carcinoma model. Key findingsIn vitro cytotoxicity experiments demonstrated that TMZ improved the potency of DOX in MCF-7 cell lines in a synergistic manner. In vivo testing confirmed that DOX/TMZ combination exhibits synergistic effect at both DOX/TMZ 1:10 and 1:5 ratios, where DOX was administered at one tenth and one fifth of its original dose, respectively. The co-treatment (1:5 ratio) significantly reduced tumor Nicotinamide adenine dinucleotide (NAD)+/NADH ratio (6.1-fold) and Adenosine triphosphate (ATP) levels (61 %) with concurrent activation of AMP-activated protein kinase (AMPK) (2.2-fold) and peroxisome proliferator-activated receptor-gamma coactivator (PGC)1-α (5.5-fold) protein expression versus control. The same treatment decreased the nuclear levels of NF-κB (p65) (57.5 %) and induced tumor apoptosis as evidenced by elevated Bax/Bcl-2 ratio (6.8-fold) along with active caspase-3 levels (6.6-fold) against control. SignificanceThe current investigation constitutes a proof-of-concept study that provided preclinical evidence for the anticancer activity of DOX/TMZ combination and warrants further investigation for repurposing TMZ in DOX protocols.

Acetyl-CoA Acetyltransferase 2 Confers Radioresistance by Inhibiting Ferroptosis in Esophageal Squamous Cell Carcinoma

The overall survival of patients with esophageal squamous cell carcinoma (ESCC) is not high due to the lack of markers to evaluate concurrent chemoradiation therapy (CCRT) resistance. The aim of this study is to use proteomics to identify a protein related to radiation therapy resistance and explore its molecular mechanisms. Proteomic data for pretreatment biopsy tissues from 18 patients with ESCC who underwent CCRT (complete response [CR] group, n=8; incomplete response [<CR] group, n=10) were collected and combined with ESCC proteomic data from iProx (n=124) to identify candidate proteins that confer resistance to CCRT. Subsequently, 125 paraffin-embedded biopsies were used for immunohistochemical validation. Colony formation assays, after ionizing radiation (IR), of acetyl-CoA acetyltransferase 2 (ACAT2)-overexpressing, -knockdown or knockout ESCC cells were used to determine the effects of ACAT2 on radioresistance. reactive oxygen species, C11-BODIPY, and Western blotting were employed to reveal the potential mechanism of ACAT2-mediated radioresistance after IR. Enrichment analysis of differentially expressed proteins (<CR vs CR) showed that the pathways of molecules conferring CCRT resistance in ESCC were related to lipid metabolism, whereas molecules conferring CCRT sensitivity were mainly related to immunity pathways. ACAT2 was selected from proteomics and validated by immunohistochemistry as a risk factor for reduced overall survival and CCRT or radiation therapy resistance among ESCC patients. ACAT2 overexpression conferred resistance to IR treatment, whereas ACAT2 knockdown or knockout conferred IR sensitivity. ACAT2 knockout cells were prone to have elevated reactive oxygen species production, enhanced lipid peroxidation, and reduced levels of glutathione peroxidase 4 after IR compared with irradiated wild-type cells. ACAT2 knockout cells could be rescued from IR-mediated toxicity by ferrostatin-1 and liproxstatin. ACAT2 overexpression confers radioresistance by inhibiting ferroptosis in ESCC, suggesting ACAT2 could be a potential biomarker of poor radiotherapeutic response and a therapeutic target for enhancing the radiosensitivity of ESCC.

CRISPR/Cas9-Mediated Genome Editing via Homologous Recombination in a Centric Diatom Chaetoceros muelleri

Chaetoceros, the most abundant genus of marine planktonic diatoms, can be used in mariculture. An effective genetic transformation system with a short transformation period was established in Chaetoceros muelleri by electroporation in our previous study. In this study, a sequence-specific clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 vector applicable for C. muelleri was constructed, and the expressions of sgRNA, resistance gene, and Cas9 gene were driven by the endogenous promoters U6, acetyl-CoA acetyltransferase, and fucoxanthin chlorophyll a/c binding protein, respectively, in the vector. Nitrate reductase (NR) and urease (URE) genes were edited in C. muelleri, and the NR knockout and NR/URE double-knockout lines displayed the strict auxotrophic phenotype. In addition, the DNA double-strand break was repaired by homologous recombination when a donor DNA was introduced. CRISPR/Cas9 technology was successfully applied to C. muelleri with an editing efficiency of up to 86%, providing a molecular tool for the study of basic biology in C. muelleri and its synthetic biology applications.

Abstract 2829: Acetyl-CoA Acetyltransferase 2 confers radioresistance by inhibiting ferroptosis and enhancing the phosphorylation of AKT-S473 in esophageal squamous cell carcinoma

Abstract Purpose: The overall efficacy of patients with esophageal squamous cell carcinoma (ESCC) is not high due to the lack of markers to evaluate concurrent chemoradiotherapy (CCRT) resistance, which is dominated by radiotherapy. The aim of this study is to identify a radioresistant protein from proteomics and explore its molecular mechanism. Methods and Materials: The proteomic data of pre-treatment biopsy tissues of 18 ESCC patients with CCRT (complete response CR group, n = 8; incomplete response &amp;lt;CR group, n = 10) was collected, combined with PXD021701 to explore the radioresistant candidate. Subsequently, immunohistochemistry of 126 paraffin embedded biopsies was used for validation. The ESCC cell models with overexpression, knockdown or knockout of Acetyl-CoA Acetyltransferase 2 (ACAT2) were applied for exploring its effects on radioresistance using clonogenic assay underwent ionizing radiation (IR). ROS, C11-BODIPY, Fer-1 (ferrostatin inhibitor) and western blotting analysis were employed to reveal the potential mechanism ACAT2 triggered after IR. Results: Enrichment analysis of differentially expressed proteins (&amp;lt;CR vs. CR) showed that the pathways of CCRT resistance molecules in ESCC were related to lipid metabolism, while the sensitive ones were mainly aggregated to immunity pathways. ACAT2 was selected from proteomics and verified as a risk factor of overall survival and a radioresistant factor among ESCC patients with CCRT or radiotherapy by immunohistochemistry. ACAT2 overexpression conferred ESCC cell resistance to IR treatment, whereas the ACAT2 knockdown or knockout ones were sensitive to IR. ACAT2 knockout cells were prone to produce ROS and enhance lipid peroxidation after IR, which was rescued by Fer-1. ACAT2 knockout also inhibited GPX4 and phosphorylation of AKT-S473 site after IR. Conclusion: ACAT2 is negatively associated with the response of radiotherapy and prognosis of ESCC patients, suggesting ACAT2 as an effective therapy target to enhance radiosensitivity of ESCC. This work was supported by the National Natural Science Foundation of China (82273108 and 82173034), the Innovative Team Grant of Guangdong Department of Education (2021KCXTD005), the Science and Technology Special Fund project of Guangdong Province in 2021 (No. 210729156901797) and the 2020 Li Ka Shing Foundation Cross-Disciplinary Research Grant (2020LKSFG07B) Citation Format: Heng Jinghua, Li Enmin, Xu Liyan. Acetyl-CoA Acetyltransferase 2 confers radioresistance by inhibiting ferroptosis and enhancing the phosphorylation of AKT-S473 in esophageal squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2829.

Impact of Pneumococcal and Viral Pneumonia on the Respiratory and Intestinal Tract Microbiomes of Mice.

With 2.56 million deaths worldwide annually, pneumonia is one of the leading causes of death. The most frequent causative pathogens are Streptococcus pneumoniae and influenza A virus. Lately, the interaction between the pathogens, the host, and its microbiome have gained more attention. The microbiome is known to promote the immune response toward pathogens; however, our knowledge on how infections affect the microbiome is still scarce. Here, the impact of colonization and infection with S. pneumoniae and influenza A virus on the structure and function of the respiratory and gastrointestinal microbiomes of mice was investigated. Using a meta-omics approach, we identified specific differences between the bacterial and viral infection. Pneumococcal colonization had minor effects on the taxonomic composition of the respiratory microbiome, while acute infections caused decreased microbial complexity. In contrast, richness was unaffected following H1N1 infection. Within the gastrointestinal microbiome, we found exclusive changes in structure and function, depending on the pathogen. While pneumococcal colonization had no effects on taxonomic composition of the gastrointestinal microbiome, increased abundance of Akkermansiaceae and Spirochaetaceae as well as decreased amounts of Clostridiaceae were exclusively found during invasive S. pneumoniae infection. The presence of Staphylococcaceae was specific for viral pneumonia. Investigation of the intestinal microbiomés functional composition revealed reduced expression of flagellin and rubrerythrin and increased levels of ATPase during pneumococcal infection, while increased amounts of acetyl coenzyme A (acetyl-CoA) acetyltransferase and enoyl-CoA transferase were unique after H1N1 infection. In conclusion, identification of specific taxonomic and functional profiles of the respiratory and gastrointestinal microbiome allowed the discrimination between bacterial and viral pneumonia. IMPORTANCE Pneumonia is one of the leading causes of death worldwide. Here, we compared the impact of bacterial- and viral-induced pneumonia on the respiratory and gastrointestinal microbiome. Using a meta-omics approach, we identified specific profiles that allow discrimination between bacterial and viral causative.

Metabolic engineering of Escherichia coli for biological production of 1, 3-Butanediol

The production of 1,3-butanediol (1,3-BDO) from glucose was investigated using Escherichia coli as the host organism. A pathway was engineered by overexpressing genes phaA (acetyl-CoA acetyltransferase), phaB (acetoacetyl-CoA reductase), bld (CoA-acylating aldehyde dehydrogenase), and yqhD (alcohol dehydrogenase). The expression levels of these genes were optimized to improve 1,3-BDO production and pathways that compete with 1,3-BDO synthesis were disrupted. Culture conditions were also optimized, including the C: N ratio, aeration, induction time, temperature, and supplementation of amino acids, resulting in a strain that could produce 1,3-BDO at 257 mM in 36 h, with a yield of 0.51 mol/mol in a fed-batch bioreactor experiment. To the best of our knowledge, this is the highest titer of 1,3-BDO production ever reported using biological methods, and our findings provide a promising strategy for the development of microbial cell factories for the sustainable synthesis of other acetyl-CoA-derived chemicals.

Effects of UBE3A on Cell and Liver Metabolism through the Ubiquitination of PDHA1 and ACAT1.

Nonalcoholic fatty liver disease (NAFLD) is substantiated by the reprogramming of liver metabolic pathways that disrupts the homeostasis of lipid and glucose metabolism and thus promotes the progression of the disease. The metabolic pathways associated with NAFLD are regulated at different levels from gene transcription to various post-translational modifications including ubiquitination. Here, we used a novel orthogonal ubiquitin transfer platform to identify pyruvate dehydrogenase A1 (PDHA1) and acetyl-CoA acetyltransferase 1 (ACAT1), two important enzymes that regulate glycolysis and ketogenesis, as substrates of E3 ubiquitin ligase UBE3A/E6AP. We found that overexpression of UBE3A accelerated the degradation of PDHA1 and promoted glycolytic activities in HEK293 cells. Furthermore, a high-fat diet suppressed the expression of UBE3A in the mouse liver, which was associated with increased ACAT1 protein levels, while forced expression of UBE3A in the mouse liver resulted in decreased ACAT1 protein contents. As a result, the mice with forced expression of UBE3A in the liver exhibited enhanced accumulation of triglycerides, cholesterol, and ketone bodies. These results reveal the role of UBE3A in NAFLD development by inducing the degradation of ACAT1 in the liver and promoting lipid storage. Overall, our work uncovers an important mechanism underlying the regulation of glycolysis and lipid metabolism through UBE3A-mediated ubiquitination of PDHA1 and ACAT1 to regulate their stabilities and enzymatic activities in the cell.

ACAT1-mediated METTL3 acetylation inhibits cell migration and invasion in triple negative breast cancer.

Triple-negative breast cancer (TNBC) is a heterogeneous and aggressive disease with poor prognosis. Acetylation modifications affect a great number of biological processes of malignant tumors. The current study aims at revealing the role of acetylation-related mechanism in TNBC progression. Methyltransferase like-3 (METTL3) was found to be downregulated in TNBC cells via quantitative polymerase chain reaction (qPCR) and western blot analyses. Co-Immunoprecipitation (Co-IP) and GST pulldown assays revealed the interaction between acetyl-CoA acetyltransferase 1 (ACAT1) and METTL3. Through further immunoprecipitation (IP) assay, we determined that ACAT1 stabilizes METTL3 protein via inhibiting the degradation of ubiquitin-proteasome. Functionally, ACAT1 inhibits TNBC cell migration and invasion. Moreover, nuclear receptor subfamily 2 group F member 6 (NR2F6) regulates ACAT1 expression at transcriptional level. Finally, we demonstrated that NR2F6/ACAT/METTL3 axis suppresses the migration and invasion of TNBC cells via METTL3. In conclusion, NR2F6 transcriptionally activates ACAT1 and promotes the suppressive effects of ACAT1-mediated METTL3 acetylation on TNBC cell migration and invasion.

Screening of candidate genes related to differences in growth and development between Chinese indigenous and Western pig breeds.

Neijiang (NJ) and Yacha (YC) are two indigenous pig breeds in the Sichuan basin of China, displaying higher resistance to diseases, lower lean ratio, and slower growth rate than the commercial Western pig breed Yorkshire (YS). The molecular mechanisms underlying the differences in growth and development between these pig breeds are still unknown. In the present study, five pigs from NJ, YC, and YS breeds were subjected to the whole genome resequencing, and then the differential single-nucleotide polymorphisms (SNPs) were screened using a 10-kb window sliding in 1-kb step using the Fst method. Finally, 48,924, 48,543, and 46,228 nonsynonymous single-nucleotide polymorphism loci (nsSNPs) were identified between NJ and YS, NJ and YC, and YC and YS, which highly or moderately affected 2,490, 800, and 444 genes, respectively. Moreover, three nsSNPs were detected in the genes of acetyl-CoA acetyltransferase 1 (ACAT1) insulin-like growth factor 2 receptor (IGF2R), insulin-like growth factor 2 and mRNA-binding protein 3 (IGF2BP3), which potentially affected the transformation of acetyl-CoA to acetoacetyl-CoA and the normal functions of the insulin signaling pathways. Moreover, serous determinations revealed significantly lower acetyl-CoA content in YC than in YS, supporting that ACAT1 might be a reason explaining the differences in growth and development between YC and YS breeds. Contents of phosphatidylcholine (PC) and phosphatidic acid (PA) significantly differed between the pig breeds, suggesting that glycerophospholipid metabolism might be another reason for the differences between Chinese and Western pig breeds. Overall, these results might contribute basic information to understand the genetic differences determining the phenotypical traits in pigs.

Cancer Metabolism: Fasting Reset, the Keto-Paradox and Drugs for Undoing

In tumor cells, ketolysis "via" succinyl-CoA: 3-oxoacid-CoAtransferase (SCOT) and acetyl-CoA acetyltransferase 1 (ACAT1) is a major source of mitochondrial acetyl-CoA. Active ACAT1 tetramers stabilize by tyrosine phosphorylation, which facilitates the SCOT reaction and ketolysis. Tyrosine phosphorylation of pyruvate kinase PK M2 has the opposite effect, stabilizing inactive dimers, while pyruvate dehydrogenase (PDH), which is already inhibited by phosphorylation, is acetylated by ACAT1 and is doubly locked. This closes the glycolytic supply of acetyl-CoA. In addition, since tumor cells must synthesize fatty acids to create new membranes, they automatically turn off the degradation of fatty acids into acetyl-CoA ("via" the malonyl-CoA brake for the fatty acid carnityl transporter). Thus, inhibiting SCOT the specific ketolytic enzyme and ACAT1 should hold back tumor progression. However, tumor cells are still able to take up external acetate and convert it into acetyl-CoA in their cytosol "via" an acetyl-CoA synthetase, which feeds the lipogenic pathway; additionally, inhibiting this enzyme would make it difficult for tumor cells to form new lipid membrane and survive.

Transcriptome Analysis of Antrodia cinnamomea Mycelia from Different Wood Substrates

Antrodia cinnamomea, an edible and medicinal fungus with significant economic value and application prospects, is rich in terpenoids, benzenoids, lignans, polysaccharides, and benzoquinone, succinic and maleic derivatives. In this study, the transcriptome of A. cinnamomea cultured on the wood substrates of Cinnamomum glanduliferum (YZM), C. camphora (XZM), and C. kanehirae (NZM) was sequenced using the high-throughput sequencing technology Illumina HiSeq 2000, and the data were assembled by de novo strategy to obtain 78,729 Unigenes with an N50 of 4,463 bp. Compared with public databases, about 11,435, 6,947, and 5,994 Unigenes were annotated to the Non-Redundant (NR), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genome (KEGG), respectively. The comprehensive analysis of the mycelium terpene biosynthesis-related genes in A. cinnamomea revealed that the expression of acetyl-CoA acetyltransferase (AACT), acyl-CoA dehydrogenase (MCAD), 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA), mevalonate pyrophosphate decarboxylase (MVD), and isopentenyl diphosphate isomerase (IDI) was significantly higher on NZM compared to the other two wood substrates. Similarly, the expression of geranylgeranyltransferase (GGT) was significantly higher on YZM compared to NZM and XZM, and the expression of farnesyl transferase (FTase) was significantly higher on XZM. Furthermore, the expressions of 2,3-oxidized squalene cyclase (OCS), squalene synthase (SQS), and squalene epoxidase (SE) were significantly higher on NZM. Overall, this study provides a potential approach to explore the molecular regulation mechanism of terpenoid biosynthesis in A. cinnamomea.

PS-B04-8: YM750, AN ACAT1 INHIBITOR, REDUCES ALDOSTERONE SECRETION BY DECREASING ADRENAL CYP11B2 EXPRESSION

Primary aldosteronism (PA) is a refractory hypertension that accounts for approximately 5%-10% of hypertensive patients. aldosterone-producing adenomas and idiopathic hyperaldosteronism are known causes of PA. In aldosterone-producing adenomas, depolarization due to somatic mutations of ion channels in adrenocortical cells is considered the cause of aldosterone oversecretion. Recently, there have been reports on the expression of various cholesterol-metabolizing enzymes in aldosterone-producing adenomas, suggesting that intracellular cholesterol metabolism is very important for aldosterone secretion. We present here the possibility that an inhibitor of Acetyl-CoA acetyltransferase (ACAT1), an enzyme that converts cholesterol to cholesteryl esters, may inhibit aldosterone secretion by depolarization in adrenal cells. Potassium chloride and YM750, an ACAT1 inhibitor, were added to human adrenocortical carcinoma, H295R cells. Then, we measured aldosterone synthase gene, CYP11B2 mRNA expression level. As a result, the expression level of CYP11B2 decreased. Both NURR1 and NGFIB mRNA expression levels also decreased. Additionally, aldosterone production was also measured and found to be significantly decreased in the YM750 added group. Next, angiotensin II was added and the inhibitory effect of YM750 was examined. The results showed that CYP11B2 expression was not decreased. This indicates that YM750 decreased the induction of CYP11B2 expression in a potassium chloride-induced stimulus-specific manner. The experimental results suggest that YM750 specifically inhibits CYP11B2 gene expression by cell depolarization and suppresses aldosterone secretion. However, there are few reports of cholesterol metabolism affecting CYP11B2 gene expression. In this study, we found that intracellular cholesterol metabolism may affect CYP11B2 gene expression. We plan to investigate the relationship between ACAT inhibition and CYP11B2 gene expression in detail mechanisms in the future.

Transcriptome analysis reveals that jasmonic acid biosynthesis and signaling is associated with the biosynthesis of asperosaponin VI in Dipsacus asperoides.

Dipsacus asperoides is a perennial herb, the roots of which are abundant in asperosaponin VI, which has important medicinal value. However, the molecular mechanism underlying the biosynthesis of asperosaponin VI in D. asperoides remains unclear. In present study, a comprehensive investigation of asperosaponin VI biosynthesis was conducted at the levels of metabolite and transcript during root development. The content of asperosaponin VI was significantly accumulated in two-leaf stage roots, and the spatial distribution of asperosaponin VI was localized in the xylem. The concentration of asperosaponin VI gradually increased in the root with the development process. Transcriptome analysis revealed 3916 unique differentially expressed genes (DEGs) including 146 transcription factors (TFs) during root development in D. asperoides. In addition, α-linolenic acid metabolism, jasmonic acid (JA) biosynthesis, JA signal transduction, sesquiterpenoid and triterpenoid biosynthesis, and terpenoid backbone biosynthesis were prominently enriched. Furthermore, the concentration of JA gradually increased, and genes involved in α-linolenic acid metabolism, JA biosynthesis, and triterpenoid biosynthesis were up-regulated during root development. Moreover, the concentration of asperosaponin VI was increased following methyl jasmonate (MeJA) treatment by activating the expression of genes in the triterpenoid biosynthesis pathway, including acetyl-CoA acetyltransferase (DaAACT), 3-hydroxy-3-methylglutaryl coenzyme A synthase (DaHMGCS), 3-hydroxy-3-methylglutaryl coenzyme-A reductase (DaHMGCR). We speculate that JA biosynthesis and signaling regulates the expression of triterpenoid biosynthetic genes and facilitate the biosynthesis of asperosaponin VI. The results suggest a regulatory network wherein triterpenoids, JA, and TFs co-modulate the biosynthesis of asperosaponin VI in D. asperoides.

Autophagy inhibition and reactive oxygen species elimination by acetyl-CoA acetyltransferase 1 through fused in sarcoma protein to promote prostate cancer

BackgroundProstate cancer is a major health issue affecting the male population worldwide, and its etiology remains relatively unknown. As presented on the Gene Expression Profiling Interactive Analysis database, acetyl-CoA acetyltransferase 1 (ACAT1) acts as a prostate cancer-promoting factor. ACAT1 expression in prostate cancer tissues is considerably higher than that in normal tissues, leading to a poor prognosis in patients with prostate cancer. Here, we aimed to study the role of the ACAT1-fused in sarcoma (FUS) complex in prostate cancer and identify new targets for the diagnosis and treatment of the disease.MethodsWe conducted immunohistochemical analysis of 57 clinical samples and in vitro and in vivo experiments using a mouse model and plasmid constructs to determine the expression of ACAT1 in prostate cancer.ResultsThe relationship between the expression of ACAT1 and the Gleason score was significant. The expression of ACAT1 was higher in tissues with a Gleason score of > 7 than in tissues with a Gleason score of ≤7 (P = 0.0011). In addition, we revealed that ACAT1 can interact with the FUS protein.ConclusionsIn prostate cancer, ACAT1 promotes the expression of P62 and Nrf2 through FUS and affects reactive oxygen species scavenging. These effects are due to the inhibition of autophagy by ACAT1. That is, ACAT1 promotes prostate cancer by inhibiting autophagy and eliminating active oxygen species. The expression of ACAT1 is related to prostate cancer. Studying the underlying mechanism may provide a new perspective on the treatment of prostate cancer.