scholarly journals ACSS2-mediated acetyl-CoA synthesis from acetate is necessary for human cytomegalovirus infection

2017 ◽  
Vol 114 (8) ◽  
pp. E1528-E1535 ◽  
Author(s):  
Anna Vysochan ◽  
Arjun Sengupta ◽  
Aalim M. Weljie ◽  
James C. Alwine ◽  
Yongjun Yu
Keyword(s):  
Human Cytomegalovirus ◽  
Mammalian Cells ◽  
Human Fibroblasts ◽  
Lipid Synthesis ◽  
Atp Citrate Lyase ◽  
Acetyl Coa ◽  
Glucose Carbon ◽  
Citrate Lyase ◽  
Infected Cells ◽  
Human Cytomegalovirus Infection

Recent studies have shown that human cytomegalovirus (HCMV) can induce a robust increase in lipid synthesis which is critical for the success of infection. In mammalian cells the central precursor for lipid biosynthesis, cytosolic acetyl CoA (Ac-CoA), is produced by ATP-citrate lyase (ACLY) from mitochondria-derived citrate or by acetyl-CoA synthetase short-chain family member 2 (ACSS2) from acetate. It has been reported that ACLY is the primary enzyme involved in making cytosolic Ac-CoA in cells with abundant nutrients. However, using CRISPR/Cas9 technology, we have shown that ACLY is not essential for HCMV growth and virally induced lipogenesis. Instead, we found that in HCMV-infected cells glucose carbon can be used for lipid synthesis by both ACLY and ACSS2 reactions. Further, the ACSS2 reaction can compensate for the loss of ACLY. However, in ACSS2-KO human fibroblasts both HCMV-induced lipogenesis from glucose and viral growth were sharply reduced. This reduction suggests that glucose-derived acetate is being used to synthesize cytosolic Ac-CoA by ACSS2. Previous studies have not established a mechanism for the production of acetate directly from glucose metabolism. Here we show that HCMV-infected cells produce more glucose-derived pyruvate, which can be converted to acetate through a nonenzymatic mechanism.

scholarly journals Lipid biosynthesis in liver slices of the foetal guinea pig

1976 ◽  
Vol 154 (1) ◽  
pp. 149-158 ◽  
Author(s):  
C T Jones ◽  
I K Ashton
Keyword(s):  
Phosphoenolpyruvate Carboxykinase ◽  
Fatty Acid Synthetase ◽  
Lipid Synthesis ◽  
Short Chain Fatty Acids ◽  
Acetate Incorporation ◽  
Atp Citrate Lyase ◽  
Acetyl Coa ◽  
Citrate Lyase ◽  
Effect Of Glucose

Lipid synthesis as measured by the incorporation of acetate or 3H2O into slices of foetal liver, is much higher than in slices of adult liver and shows a peak at about two-thirds of gestation. At this time the synthesis from glucose was low and reached a peak 10 days later. The changes in the activity of ATP citrate lyase, which mirrored acetate incorporation, and the effect of glucose and pyruvate on acetate corporation into lipid suggests that some of the lipid synthesis occurs via intramitochondrial acetyl-CoA production from acetate. Despite this, lipid synthesis was not inhibited by (-)-hydroxycitrate. The low rate of synthesis from glucose at two-thirds of gestation is ascribed to the low activity of pyruvate carboxylase at this time and a role for a phosphoenolpyruvate carboxykinase in providing oxaloacetate for lipogenesis is proposed. The activity of fatty acid synthetase broadly agreed with the changes in lipid synthesis, whereas the activity of acetyl-CoA carboxylase was barely sufficient to account for the rates of lipid synthesis in vivo. Acetate and short-chain fatty acids are likely to be the major precursors for lipid synthesis in vivo.

Human cytomegalovirus infection stimulates Cl−/ HCO 3 − exchanger activity in human fibroblasts

1998 ◽  
Vol 275 (2) ◽  
pp. C515-C526 ◽  
Author(s):  
Lilia M. Maglova ◽  
William E. Crowe ◽  
Aníbal A. Altamirano ◽  
John M. Russell
Keyword(s):  
Human Cytomegalovirus ◽  
Hcmv Infection ◽  
Human Lung ◽  
Human Fibroblasts ◽  
Cell Types ◽  
Recovery Phase ◽  
Lung Fibroblasts ◽  
Continuous Exposure ◽  
Infected Cells ◽  
Human Cytomegalovirus Infection

The effects of human cytomegalovirus (HCMV) infection on Cl−/[Formula: see text]exchanger activity in human lung fibroblasts (MRC-5 cells) were studied using fluorescent, ion-sensitive dyes. The intracellular pH (pHi) of mock- and HCMV-infected cells bathed in a solution containing 5% CO2-25 mM[Formula: see text] were nearly the same. However, replacement of external Cl−with gluconate caused an H2DIDS-inhibitable (100 μM) increase in the pHi of HCMV-infected cells but not in mock-infected cells. Continuous exposure to hyperosmotic external media containing CO2/[Formula: see text]caused the pHi of both cell types to increase. The pHi remained elevated in mock-infected cells. However, in HCMV-infected cells, the pHi peaked and then recovered toward control values. This pHirecovery phase was completely blocked by 100 μM H2DIDS. In the presence of CO2/[Formula: see text], there was an H2DIDS-sensitive component of net Cl− efflux (external Cl− was substituted with gluconate) that was less in mock- than in HCMV-infected cells. When nitrate was substituted for external Cl− (in the nominal absence of CO2/[Formula: see text]), the H2DIDS-sensitive net Cl− efflux was much greater from HCMV- than from mock-infected cells. In mock-infected cells, H2DIDS-sensitive, net Cl− efflux decreased as pHi increased, whereas for HCMV-infected cells, efflux increased as pHi increased. All these results are consistent with an HCMV-induced enhancement of Cl−/[Formula: see text]exchanger activity.

Human cytomegalovirus infection enhances osmotic stimulation of Na+/H+exchange in human fibroblasts

1997 ◽  
Vol 273 (5) ◽  
pp. C1739-C1748 ◽  
Author(s):  
William E. Crowe ◽  
Aníbal A. Altamirano ◽  
John M. Russell
Keyword(s):  
Human Cytomegalovirus ◽  
Hcmv Infection ◽  
Human Fibroblasts ◽  
Maximum Velocity ◽  
Alkaline Shift ◽  
Solute Uptake ◽  
Buffering Power ◽  
Infected Cells ◽  
Human Cytomegalovirus Infection ◽  
Acid Loading

Infection with human cytomegalovirus (HCMV) causes an enlargement (cytomegaly) of human fibroblasts (MRC-5). As a first step toward determining whether solute uptake, mediated in part by Na+/H+exchange, is responsible for the development of cytomegaly, we studied the effects of HCMV infection on intracellular pH (pHi) regulation (nominal CO2/[Formula: see text]concn = 0) by comparing cytomegalic cells with mock-infected cells. Seventy-two hours after HCMV infection of MRC-5 cells we observed the following changes relative to mock-infected cells: resting pHiis 0.1–0.2 pH unit more alkaline; the intrinsic buffering power of the cytoplasm was reduced by ∼40–50%; acid-loading H+-equivalent fluxes were reduced; and there were alterations of Na+/H+exchanger (NHE) properties, including an alkaline shift of the pHidependence of activity, a reduction of the apparent affinity for extracellular Na+, and an increase of the apparent maximum velocity and a large increase in stimulation by a hyperosmotic challenge. These results indicate that HCMV infection exerts a profound effect on functional properties of the NHE, on acid-loading mechanisms, and on intrinsic cellular buffering power. These effects are consistent with a role for the NHE in the development of cytomegaly.

scholarly journals Accumulation of Substrates of the Anaphase-Promoting Complex (APC) during Human Cytomegalovirus Infection Is Associated with the Phosphorylation of Cdh1 and the Dissociation and Relocalization of APC Subunits

2007 ◽  
Vol 82 (1) ◽  
pp. 529-537 ◽  
Author(s):  
Karen Tran ◽  
Jeffrey A. Mahr ◽  
Jiwon Choi ◽  
Jose G. Teodoro ◽  
Michael R. Green ◽  
...  
Keyword(s):  
Human Cytomegalovirus ◽  
Human Fibroblasts ◽  
Productive Infection ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Core Complex ◽  
The Core ◽  
Phosphorylated Form ◽  
Infected Cells ◽  
Human Cytomegalovirus Infection

ABSTRACT Cell cycle dysregulation upon human cytomegalovirus (HCMV) infection of human fibroblasts is associated with the inactivation of the anaphase-promoting complex (APC), a multisubunit E3 ubiquitin ligase, and accumulation of its substrates. Here, we have further elucidated the mechanism(s) by which HCMV-induced inactivation of the APC occurs. Our results show that Cdh1 accumulates in a phosphorylated form that may prevent its association with and activation of the APC. The accumulation of Cdh1, but not its phosphorylation, appears to be cyclin-dependent kinase dependent. The lack of an association of exogenously added Cdh1 with the APC from infected cells indicates that the core APC also may be impaired. This is further supported by an examination of the localization and composition of the APC. Coimmunoprecipitation studies show that both Cdh1 and the subunit APC1 become dissociated from the complex. In addition, immunofluorescence analysis demonstrates that as the infection progresses, several subunits redistribute to the cytoplasm, while APC1 remains nuclear. Dissociation of the core complex itself would account for not only the observed inactivity but also its inability to bind to Cdh1. Taken together, these results illustrate that HCMV has adopted multiple mechanisms to inactivate the APC, which underscores its importance for a productive infection.

scholarly journals Glutamine Metabolism Is Essential for Human Cytomegalovirus Infection

2009 ◽  
Vol 84 (4) ◽  
pp. 1867-1873 ◽  
Author(s):  
Jeremy W. Chambers ◽  
Tobi G. Maguire ◽  
James C. Alwine
Keyword(s):  
Human Cytomegalovirus ◽  
Human Fibroblasts ◽  
Tca Cycle ◽  
Glutamine Metabolism ◽  
Glucose Starvation ◽  
Atp Production ◽  
The Tca Cycle ◽  
Infected Cells ◽  
Human Cytomegalovirus Infection ◽  
Glutamine Uptake

ABSTRACT Human fibroblasts infected with human cytomegalovirus (HCMV) were more viable than uninfected cells during glucose starvation, suggesting that an alternate carbon source was used. We have determined that infected cells require glutamine for ATP production, whereas uninfected cells do not. This suggested that during infection, glutamine is used to fill the tricarboxylic acid (TCA) cycle (anaplerosis). In agreement with this, levels of glutamine uptake and ammonia production increased in infected cells, as did the activities of glutaminase and glutamate dehydrogenase, the enzymes needed to convert glutamine to α-ketoglutarate to enter the TCA cycle. Infected cells starved for glutamine beginning 24 h postinfection failed to produce infectious virions. Both ATP and viral production could be rescued in glutamine-starved cells by the TCA intermediates α-ketoglutarate, oxaloacetate, and pyruvate, confirming that in infected cells, a program allowing glutamine to be used anaplerotically is induced. Thus, HCMV infection activates the mechanisms needed to switch the anaplerotic substrate from glucose to glutamine to accommodate the biosynthetic and energetic needs of the viral infection and to allow glucose to be used biosynthetically.

Acetyl-CoA is produced by the citrate synthase homology module of ATP-citrate lyase

2021 ◽  
Author(s):  
Kenneth Verstraete ◽  
Koen H. G. Verschueren ◽  
Ann Dansercoer ◽  
Savvas N. Savvides
Keyword(s):  
Citrate Synthase ◽  
Atp Citrate Lyase ◽  
Acetyl Coa ◽  
Citrate Lyase ◽  
Homology Module

Abstract 15978: Acetyl-coa Catalysis by Atp-citrate Lyase is Essential for Myofibroblast Differentiation and Persistence

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Michael P Lazaropoulos ◽  
Andrew A Gibb ◽  
Anh Huynh ◽  
Kathryn Wellen ◽  
John W Elrod
Keyword(s):  
Histone Acetylation ◽  
Transcriptional Activation ◽  
Cardiac Fibrosis ◽  
Transforming Growth Factor ◽  
Metabolic Reprogramming ◽  
Myofibroblast Differentiation ◽  
Atp Citrate Lyase ◽  
Acetyl Coa ◽  
Matrix Deposition ◽  
Citrate Lyase

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.

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

2020 ◽  
Vol 41 (6) ◽  
pp. 778-789 ◽  
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.

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