Reverse Genetic Characterization of Cytosolic Acetyl-CoA Generation by ATP-Citrate Lyase in Arabidopsis

A feature of heart failure (HF) is excessive extracellular matrix deposition and cardiac remodeling by a differentiated fibroblast population known as myofibroblasts. Identifying mechanisms of myofibroblast differentiation in cardiac fibrosis could yield novel therapeutic targets to delay or reverse HF. Recent evidence suggests that myofibroblast differentiation requires metabolic reprogramming for transcriptional activation of the myofibroblast gene program by chromatin-dependent mechanisms. We previously reported that inhibition of histone demethylation blocks myofibroblast formation, however, whether histone acetylation (e.g., H3K27ac, a prominent mark associated with gene transcription) is involved in fibroblast reprogramming remains unclear. ATP-citrate lyase (ACLY) synthesizes acetyl-CoA and therein supplies acetyl-CoA to the nucleus, where it is used as a substrate by histone acetyltransferases (HATs). To define the role of acetyl-CoA metabolism in myofibroblast differentiation, we stimulated differentiation in mouse embryonic fibroblasts (MEFs) and adult mouse cardiac fibroblasts (ACFs) with the pro-fibrotic agonist transforming growth factor β (TGFβ) and treated cells with a pharmacological inhibitor of ACLY. ACLY inhibition decreased myofibroblast gene expression in ACF and MEFs in TGFβ-stimulated myofibroblast differentiation, in addition to decreasing the population of αSMA positive MEFs. Genetic deletion of ACLY in MEFs recapitulated the results observed with pharmacological inhibition. Encouragingly, the ACLY inhibitor was sufficient to revert fully differentiated myofibroblasts under continuous TGFβ stimulation to a quiescent, non-fibrotic phenotype. Altogether, our data indicate that ACLY activity is necessary for myofibroblast differentiation and persistence. We hypothesize that ACLY-dependent acetyl-CoA synthesis is necessary for histone acetylation and transcriptional activation of the myofibroblast gene program. Currently, we are examining mechanisms of ACLY-dependent chromatin remodeling in fibroblasts and the in vivo relevance of this mechanism in mutant mice. In summary, ACLY is a potential target to reverse cardiac fibrosis and lessen HF.

Download Full-text

RNA-seq reveals novel mechanistic targets of withaferin A in prostate cancer cells

Carcinogenesis ◽

10.1093/carcin/bgaa009 ◽

2020 ◽

Vol 41 (6) ◽

pp. 778-789 ◽

Cited By ~ 5

Author(s):

Su-Hyeong Kim ◽

Eun-Ryeong Hahm ◽

Krishna B Singh ◽

Sruti Shiva ◽

Jacob Stewart-Ornstein ◽

...

Keyword(s):

Prostate Cancer ◽

Fatty Acid ◽

Fatty Acid Synthase ◽

Acid Synthesis ◽

Acetyl Coa Carboxylase ◽

Rna Seq ◽

Atp Citrate Lyase ◽

Acetyl Coa ◽

Citrate Lyase

Abstract Withaferin A (WA) is a promising phytochemical exhibiting in vitro and in vivo anticancer activities against prostate and other cancers, but the mechanism of its action is not fully understood. In this study, we performed RNA-seq analysis using 22Rv1 human prostate cancer cell line to identify mechanistic targets of WA. Kyoto Encyclopedia of Genes and Genomes pathway analysis of the differentially expressed genes showed most significant enrichment of genes associated with metabolism. These results were validated using LNCaP and 22Rv1 human prostate cancer cells and Hi-Myc transgenic mice as models. The intracellular levels of acetyl-CoA, total free fatty acids and neutral lipids were decreased significantly following WA treatment in both cells, which was accompanied by downregulation of mRNA (confirmed by quantitative reverse transcription-polymerase chain reaction) and protein levels of key fatty acid synthesis enzymes, including ATP citrate lyase, acetyl-CoA carboxylase 1, fatty acid synthase and carnitine palmitoyltransferase 1A. Ectopic expression of c-Myc, but not constitutively active Akt, conferred a marked protection against WA-mediated suppression of acetyl-CoA carboxylase 1 and fatty acid synthase protein expression, and clonogenic cell survival. WA was a superior inhibitor of cell proliferation and fatty acid synthesis in comparison with known modulators of fatty acid metabolism including cerulenin and etomoxir. Intraperitoneal WA administration to Hi-Myc transgenic mice (0.1 mg/mouse, three times/week for 5 weeks) also resulted in a significant decrease in circulating levels of total free fatty acids and phospholipids, and expression of ATP citrate lyase, acetyl-CoA carboxylase 1, fatty acid synthase and carnitine palmitoyltransferase 1A proteins in the prostate in vivo.

Download Full-text

Enhancement of docosahexaenoic acid production by overexpression of ATP-citrate lyase and acetyl-CoA carboxylase in Schizochytrium sp.

Biotechnology for Biofuels ◽

10.1186/s13068-020-01767-z ◽

2020 ◽

Vol 13 (1) ◽

Cited By ~ 1

Author(s):

Xiao Han ◽

Zhunan Zhao ◽

Ying Wen ◽

Zhi Chen

Keyword(s):

Docosahexaenoic Acid ◽

Acid Production ◽

Acetyl Coa Carboxylase ◽

Atp Citrate Lyase ◽

Acetyl Coa ◽

Citrate Lyase

Download Full-text

ATP-citrate lyase is essential for high glucose-induced histone hyperacetylation and fibrogenic gene upregulation in mesangial cells

AJP Renal Physiology ◽

10.1152/ajprenal.00029.2017 ◽

2017 ◽

Vol 313 (2) ◽

pp. F423-F429 ◽

Cited By ~ 13

Author(s):

Dilip K. Deb ◽

Yinyin Chen ◽

Jian Sun ◽

Youli Wang ◽

Yan Chun Li

Keyword(s):

Histone Acetylation ◽

High Glucose ◽

Mesangial Cells ◽

Nuclear Accumulation ◽

Atp Citrate Lyase ◽

Acetyl Coa ◽

Ecm Proteins ◽

Histone Hyperacetylation ◽

Citrate Lyase ◽

Tgf Β1

The goal of this study was to address the role of ATP-citrate lyase (ACL), an enzyme that converts citrate to acetyl-CoA, in high glucose (HG)-induced histone acetylation and profibrotic gene expression. Our recent ChIP-Seq studies have demonstrated that HG induces genome-wide histone hyperacetylation in mesangial cells (MCs). Here, we showed that exposure of MCs to HG markedly increased histone acetylation at the H3K9/14 and H3K18 marks and induced the expression of potent profibrotic factors TGF-β1, TGF-β3, and connective tissue growth factor (CTGF). The induction of these profibrotic factors was further enhanced by histone deacetylase inhibitor but suppressed by histone acetyl-transferase inhibitor, confirming the importance of histone acetylation in this regulation. Interestingly, HG not only upregulated ACL expression but also promoted ACL nuclear translocation, evidenced by increased ACL concentration and activity in the nuclear extracts. Consistent with this observation, transfection of MCs with a plasmid-carrying green fluorescent protein (GFP)-ACL fusion protein led to GFP nuclear accumulation when cultured in HG condition. Silencing ACL with siRNAs alleviated HG-induced histone hyperacetylation, as well as upregulation of TGF-β1, TGF-β3, CTGF, and extracellular matrix (ECM) proteins fibronectin and collagen type IV, whereas ACL overexpression further enhanced HG induction of histone acetylation, as well as these profibrotic factors and ECM proteins. Collectively, these observations demonstrate that HG promotes ACL expression and translocation into the nucleus, where ACL converts citrate to acetyl-CoA to provide the substrate for histone acetylation, leading to upregulation of fibrogenic genes. Therefore, ACL plays a critical role in epigenetic regulation of diabetic renal fibrosis.

Download Full-text

Evaluation of the rate-limiting steps in the pathway of glucose metabolism in kidney cortex of normal, diabetic, cortisone-treated and growth hormone-treated rats

Biochemical Journal ◽

10.1042/bj1281293 ◽

1972 ◽

Vol 128 (5) ◽

pp. 1293-1301 ◽

Cited By ~ 13

Author(s):

P. K. Joseph ◽

K. Subrahmanyam

Keyword(s):

Amino Acids ◽

Growth Hormone ◽

Aspartate Aminotransferase ◽

Glycogen Content ◽

Diabetic Rats ◽

Kidney Cortex ◽

Atp Citrate Lyase ◽

Acetyl Coa ◽

Ammonia Content ◽

Citrate Lyase

1. The activities of gluconeogenic and glycolytic enzymes and the concentrations of citrate, ammonia, amino acids, glycogen, glucose 6-phosphate, acetyl-CoA, lactate and pyruvate were measured in kidney cortex of normal, diabetic, cortisone-treated and growth hormone-treated rats. 2. In kidney cortex of diabetic, cortisone-treated and growth hormone-treated rats the activities of glucose 6-phosphatase (EC 3.1.3.9), fructose 1,6-diphosphatase (EC 3.1.3.11) and phosphopyruvate carboxylase (EC 4.1.1.32) were increased. 3. The activities of glutamate dehydrogenase (EC 1.4.1.3), alanine aminotransferase (EC 2.6.1.2), aspartate aminotransferase (EC 2.6.1.10) and pyruvate carboxylase (EC 6.4.1.1) were increased in diabetic and cortisone-treated rats. In growth hormone-treated rats the activity of aspartate aminotransferase was depressed but those of the other three enzymes were unchanged. 4. The activity of hexokinase (EC 2.7.1.1) was not altered in any of these conditions. Phosphofructokinase (EC 2.7.1.11) activity was depressed only in growth hormone-treated rats. Pyruvate kinase (EC 2.7.1.40) activity was depressed in cortisone-treated and growth hormone-treated rats but unchanged in diabetic rats. 5. Amino acids, acetyl-CoA and glucose 6-phosphate contents were increased in rat kidneys in all these three conditions. Ammonia content was increased in diabetic and cortisone-treated rats but was markedly diminished in growth hormone-treated rats. 6. The [lactate]/[pyruvate] ratio was elevated in diabetic and cortisone-treated rats but unchanged in growth hormone-treated rats. Citrate content was increased in the kidney cortex of diabetic and growth hormone-treated rats but was unchanged in cortisone-treated rats. The activity of ATP citrate lyase (EC 4.1.3.8) was depressed in diabetic and growth hormone-treated rats but was increased in cortisone-treated rats. 7. Glycogen content was moderately elevated in growth hormone-treated rats and markedly elevated in diabetic rats, whereas no change in glycogen content was observed in cortisone-treated rats. Glycogen synthetase (EC 2.4.1.11) activity was unchanged in all these three conditions. Phosphorylase (EC 2.4.1.1) activity was not affected in cortisone-treated rats but was depressed in diabetic and growth hormone-treated rats.

Download Full-text