Long-chain Fatty acyl-CoA Research Articles

ACSL1 improves pulmonary fibrosis by reducing mitochondrial damage and activating PINK1/Parkin mediated mitophagy

Pulmonary fibrosis is a chronic interstitial lung disease with no curative therapeutic treatment, leading to significant mortality. The aims of this study were to investigate the regulatory mechanisms of mitophagy in the progression of pulmonary fibrosis. Through bioinformatics analysis, we identified the downregulation of long-chain fatty acyl-CoA synthetase 1 (ACSL1) as being associated with the severity of pulmonary fibrosis. A pulmonary fibrosis model was established through bleomycin (BLM) exposure both in vivo and in vitro. Mitoquinone (MitoQ) pretreatment significantly decreased redox damage, stabilized mitochondrial membrane potential (MMP), improved mitochondrial dynamics, and activated PINK1/Parkin-mediated mitophagy, thereby alleviating pulmonary fibrosis. In vitro, overexpression of ACSL1 mitigated mitochondrial damage and restored PINK1/Parkin-mediated mitophagy under BLM exposure. In contrast, ACSL1 inhibition exacerbated pulmonary fibrosis, and these adverse effects could not be reversed by MitoQ treatment. Taken together, our study reveals a novel mechanism underlying the pathogenesis of pulmonary fibrosis and suggests a potential therapeutic target for its treatment.

Activity and phosphatidylcholine transfer protein interactions of skeletal muscle thioesterase Them2 enable hepatic steatosis and insulin resistance

Thioesterase superfamily member 2 (Them2), a long-chain fatty acyl-CoA thioesterase that is highly expressed in oxidative tissues, interacts with phosphatidylcholine transfer protein (PC-TP) to regulate hepatic lipid and glucose metabolism and to suppress insulin signaling. High-fat diet (HFD)-fed mice lacking Them2 globally or specifically in skeletal muscle, but not liver, exhibit reduced hepatic steatosis and insulin resistance. Here, we report that the capacity of Them2 in skeletal muscle to promote hepatic steatosis and insulin resistance depends on both its catalytic activity and interaction with PC-TP. Two residues of Them2 catalytic site were mutated (N50A/D65A) to produce the inactive enzyme while maintaining its homotetrameric structure and interaction with PC-TP. Restoration of skeletal muscle expression in Them2-/- mice using recombinant adeno-associated virus revealed that wild-type (WT), but not N50A/D65A Them2, promoted HFD-induced weight gain and hepatic steatosis. This was accompanied by greater impairment of insulin sensitivity in WT compared with N50A/D65A Them2. Pharmacological inhibition or genetic ablation of PC-TP attenuated these effects. In reductionist experiments, conditioned medium collected from WT primary cultured myotubes promoted excess lipid accumulation in oleic acid-treated primary cultured hepatocytes relative to Them2-/- myotubes, which was attributable to secreted extracellular vesicles (EV). Reconstitution of Them2 expression in Them2-/- myotubes affirmed the requirements for catalytic activity and PC-TP interactions for EV to promote lipid accumulation in hepatocytes. These studies provide valuable mechanistic insights whereby Them2 in skeletal muscle promotes hepatic steatosis and establish both Them2 and PC-TP as represent attractive targets for managing metabolic dysfunction-associated steatotic liver disease.

Role of AMPK in Atrial Metabolic Homeostasis and Substrate Preference.

Atrial fibrillation is the most common clinical arrhythmia and may be due in part to metabolic stress. Atrial specific deletion of the master metabolic sensor, AMP-activated protein kinase (AMPK), induces atrial remodeling culminating in atrial fibrillation in mice, implicating AMPK signaling in the maintenance of atrial electrical and structural homeostasis. However, atrial substrate preference for mitochondrial oxidation and the role of AMPK in regulating atrial metabolism are unknown. Here, using LC-MS/MS methodology combined with infusions of [ 13 C 6 ]glucose and [ 13 C 4 ]β-hydroxybutyrate in conscious mice, we demonstrate that conditional deletion of atrial AMPK catalytic subunits shifts mitochondrial atrial metabolism away from fatty acid oxidation and towards pyruvate oxidation. LC-MS/MS-based quantification of acyl-CoAs demonstrated decreased atrial tissue content of long-chain fatty acyl-CoAs. Proteomic analysis revealed a broad downregulation of proteins responsible for fatty acid uptake (LPL, CD36, FABP3), acylation and oxidation. Atrial AMPK deletion reduced expression of atrial PGC1-α and downstream PGC1-α/PPARα/RXR regulated gene transcripts. In contrast, atrial [ 14 C]2-deoxyglucose uptake and GLUT1 expression increased with fasting in mice with AMPK deletion, while the expression of glycolytic enzymes exhibited heterogenous changes. Thus, these results highlight the crucial homeostatic role of AMPK in the atrium, with loss of atrial AMPK leading to downregulation of the PGC1-α/PPARα pathway and broad metabolic reprogramming with a loss of fatty acid oxidation, which may contribute to atrial remodeling and arrhythmia.

A mutated algal species of Isochrysis sp. obtained via atmospheric room temperature plasma mutagenesis promoted lipid accumulation through the abscisic acid pathway

Isochrysis sp. is an economically important algae rich in unsaturated fatty acids, but its susceptibility to strain degeneration poses challenges for practical applications. To address this, we used atmospheric room temperature plasma (ARTP) for mutagenesis, creating a stable mutant strain, Yield 8 (Y8), with rapid growth and high lipid content. Transcriptome analysis showed upregulation of genes in genetic information processes and metabolic pathways, with notable upregulation of long-chain fatty acid CoA ligase and acyl-CoA acyltransferase (ACAT), consistent with the trend of fatty acid accumulation. Interestingly, the expression levels of ζ-carotene desaturase (ZDS), zeaxanthin epoxidase (ZEP), and phosphatase 2C (PP2C) were upregulated in the Y8 mutant, indicating activation of the ABA pathway. Further, the increased endogenous ABA levels in Y8, along with a 1.17-fold increase in DHA and a 1.26-fold increase in ω3 fatty acids after exogenous ABA application, suggest that Y8's improved traits are mediated by the ABA pathway. These findings highlight Y8's potential for practical applications and further research.

Structural insights into the regulation mechanism of Mycobacterium tuberculosis MftR.

Mycobacterium tuberculosis, the pathogen of the deadly disease tuberculosis, depends on the redox cofactor mycofactocin (MFT) to adapt toand survive under hypoxic conditions. MftR is a TetR family transcription regulator that binds upstream of the MFT gene cluster and controls MFT synthesis. To elucidate the structural basis underlying MftR regulation, we determined the crystal structure of Mycobacterium tuberculosis MftR (TB-MftR). The structure revealed an interconnected hydrogen bond network in the α1-α2-α3 helices of helix-turn-helix (HTH) DNA-binding domain that is essential for nucleic acid interactions. The ligand-binding domain contains a hydrophobic cavity enclosing long-chain fatty acyl-CoAs like the key regulatory ligand oleoyl-CoA. Despite variations in ligand-binding modes, comparative analyses suggest regulatory mechanisms are largely conserved across TetR family acyl-CoA sensors. By elucidating the intricate structural mechanisms governing DNA and ligand binding by TB-MftR, our study enhances understanding of the regulatory roles of this transcription factor under hypoxic conditions, providing insights that could inform future research into Mycobacterium tuberculosis pathogenesis.

Proteome Analysis Related to Unsaturated Fatty Acid Synthesis by Interfering with Bovine Adipocyte ACSL1 Gene.

Unsaturated fatty acids (UFAs) in beef play a vital role in promoting human health. Long-chain fatty acyl-CoA synthase 1 (ACSL1) is a crucial gene for UFA synthesis in bovine adipocytes. To investigate the protein expression profile during UFA synthesis, we performed a proteomic analysis of bovine adipocytes by RNA interference and non-interference with ACSL1 using label-free techniques. A total of 3558 proteins were identified in both the NC and si-treated groups, of which 1428 were differentially expressed proteins (DEPs; fold change ≥ 1.2 or ≤ 0.83 and p-value < 0.05). The enrichment analysis of the DEPs revealed signaling pathways related to UFA synthesis or metabolism, including cAMP, oxytocin, fatty acid degradation, glycerol metabolism, insulin, and the regulation of lipolysis in adipocytes (p-value < 0.05). Furthermore, based on the enrichment analysis of the DEPs, we screened 50 DEPs that potentially influence the synthesis of UFAs and constructed an interaction network. Moreover, by integrating our previously published transcriptome data, this study established a regulatory network involving differentially expressed long non-coding RNAs (DELs), highlighting 21 DEPs and 13 DELs as key genes involved in UFA synthesis. These findings present potential candidate genes for further investigation into the molecular mechanisms underlying UFA synthesis in bovines, thereby offering insights to enhance the quality of beef and contribute to consumer health in future studies.

Lipid peroxidation-induced ferroptosis as a therapeutic target for mitigating neuronal injury and inflammation in sepsis-associated encephalopathy: insights into the hippocampal PEBP-1/15-LOX/GPX4 pathway

BackgroundSepsis-associated encephalopathy (SAE) refers to the widespread impairment of brain function caused by noncentral nervous system infection mediated by sepsis. Lipid peroxidation-induced ferroptosis contributes to the occurrence and course of SAE. This study aimed to investigate the relationship between neuronal injury and lipid peroxidation-induced ferroptosis in SAE.MethodsBaseline data were collected from pediatric patients upon admission, and the expression levels of various markers related to lipid peroxidation and ferroptosis were monitored in the serum and peripheral blood mononuclear cells (PBMCs) of patients with SAE as well as SAE model mice. The hippocampal phosphatidylethanolamine-binding protein (PEBP)-1/15-lysine oxidase (LOX)/ glutathione peroxidase 4 (GPX4) pathway was assessed for its role on the inhibitory effect of ferroptosis in SAE treatment.ResultsThe results showed elevated levels of S100 calcium-binding protein beta (S-100β), glial fibrillary acidic protein, and malondialdehyde in the serum of SAE patients, while superoxide dismutase levels were reduced. Furthermore, analysis of PBMCs revealed increased transcription levels of PEBP1, LOX, and long-chain fatty acyl-CoA synthetase family member 4 (ACSL4) in SAE patients, while the transcription levels of GPX4 and cystine/glutamate transporter xCT (SLC7A11) were decreased. In comparison to the control group, the SAE mice exhibited increased expression of S-100β and neuron-specific enolase (NSE) in the hippocampus, whereas the expression of S-100β and NSE were reduced in deferoxamine (DFO) mice. Additionally, iron accumulation was observed in the hippocampus of SAE mice, while the iron ion levels were reduced in the DFO mice. Inhibition of ferroptosis alleviated the mitochondrial damage (as assessed by transmission electron microscopy, hippocampal mitochondrial ATP detection, and the JC-1 polymer-to-monomer ratio in the hippocampus) and the oxidative stress response induced by SAE as well as attenuated neuroinflammatory reactions. Further investigations revealed that the mechanism underlying the inhibitory effect of ferroptosis in SAE treatment is associated with the hippocampal PEBP-1/15-LOX/GPX4 pathway.ConclusionThese results offer potential therapeutic targets for the management of neuronal injury in SAE and valuable insights into the potential mechanisms of ferroptosis in neurological disorders.

ACSL4 promotes malignant progression of Hepatocellular carcinoma by targeting PAK2 transcription

Long-chain fatty acyl-Coa ligase 4 (ACSL4) is an important enzyme that converts fatty acids to fatty acyl-Coa esters, there is increasing evidence for its role in carcinogenesis. However, the precise role of ACLS4 in hepatocellular carcinoma (HCC) is not clearly understood. In the present study, we provide evidence that ACSL4 expression was specifically elevated in HCC and is associated with poor clinical outcomes. ACSL4 significantly promotes the growth and metastasis of HCC both in vitro and in vivo. RNA sequencing and functional experiments showed that the effect of ACSL4 on HCC development was heavily dependent on PAK2. ACSL4 expression is well correlated with PAK2 in HCC, and ACSL4 even transcriptionally increased PAK2 gene expression mediated by Sp1. In addition, emodin, a naturally occurring anthraquinone derivative, inhibited HCC cell growth and tumor progression by targeting ACSL4. In summary, ACSL4 plays a novel oncogene in HCC development by regulating PAK2 transcription. Targeting ACSL4 could be useful in drug development and therapy for HCC.

Abstract 4283: NAMPT-enriched small extracellular vesicle promotes liver cancer via activation of SLC27A4-mediated glycolysis

Abstract Introduction: Tumor-derived small extracellular vesicles (sEV) act as a major mediator of the tumor microenvironment (TME) and are reported to regulate various metabolic pathways. Methods: Small extracellular vesicles (sEV) were collected from normal human hepatocyte and metastatic HCC cell lines using differential ultracentrifugation. Mass Spectrometry proteomic analysis was employed to identify NAMPT to be enriched in metastatic HCC-sEV. The glycolytic effect of NAMPT-sEV was analyzed by seahorse glycolytic assay. Result: Mass spectrometry protein analysis revealed nicotinamide phosphoribosyl transferase (NAMPT) to be upregulated in metastatic HCC-sEV. NAMPT is a rate-limiting enzyme essential for maintaining cellular level of nicotinamide dinucleotide (NAD+). However, the underlying pathway of sEV-NAMPT-induced metabolic consequence in HCC remains largely unknown. This study elucidated sEV-NAMPT’s carcinogenic role by establishing NAMPT-knockdown (NAMPT-KD) stable clone in metastatic HCC cell lines, which significantly hampered the promoting effect of HCC-sEV in glycolysis. sEV-mediated in vitro and in vivo tumor growth was also inhibited. Through sEV treatment, we found that sEV-NAMPT significantly elevate SLC27A4 in both transcription and protein level of recipient cells. Mechanistically, it is demonstrated that SLC27A4 expression was enhanced by NF-κB transcription factor, which is activated upon binding of sEV-NAMPT to toll-like receptor 4 (TLR4). SLC27A4 mainly function as a long-chain fatty acid transporter or acyl-CoA synthetase. Lipidomic and metabolomic analysis unveiled SLC27A4 to be positively correlated with intracellular triacylglycerol (TG) and dihydroxyacetone phosphate (DHAP) level. Elevation of TG aggravates lipolysis by hepatic lipase and promotes conversion of glycerol-3-P to DHAP, which is an intermediate metabolite that transit between lipid metabolism and glycolysis. This study uncovers a regulatory axis of sEV-NAMPT-mediated SLC27A4 in glycolysis, apart from conventional fatty acid-related metabolisms. Clinically, we explored the therapeutic potential of targeting sEV-NAMPT by inhibitor FK866. Treatment of NAMPT-inhibitor significantly hampered tumor growth in various HCC in vivo models, which suggested an alternative therapeutic strategy for HCC. Conclusion: NAMPT is enriched in metastatic HCC sEV. sEV-NAMPT regulates the promoting effect of HCC-sEV in glycolysis and oncogenesis. SLC27A4 function as the downstream target that promotes HCC progression via glycolysis. NAMPT inhibitor FK866 are proven to reduce in vitro and in vivo tumorigenic ability of metastatic HCC-sEV, which may serve as potential HCC treatment option. Citation Format: Lot Sum Cherlie Yeung, Tu Him ng, Judy Wai Ping Yam. NAMPT-enriched small extracellular vesicle promotes liver cancer via activation of SLC27A4-mediated glycolysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4283.

(-)-Epigallocatechin-3-O-Gallate Regulates Muscle Growth, Antioxidant Status, and Nutritional Composition of Juvenile Common Carp (Cyprinus carpio L.)

Among the polyphenolic compounds commonly found in green tea, (-)-epigallocatechin-3-gallate (EGCG) is known to have various functions, including the promotion of antioxidant and myofiber growth in mammals. However, the effect of EGCG on common carp (Cyprinus carpio L.) remains poorly understood. To evaluate the role of EGCG on juvenile common carp, 255 fish (initial weight 17.33 ± 0.34 g) were fed with five experimental diets containing 0, 0.05, 0.25, 0.5, and 1 g/kg EGCG. The results showed that diet supplementation with 0.05–0.5 g/kg EGCG supplementation significantly enhanced the specific growth rate (SGR) and reduced the feed conversion ratio (FCR). Weight gain rate (WGR) was significantly enhanced in the 0.25 and 0.5 g/kg EGCG groups. As for antioxidants, 0.25–1 g/kg EGCG significantly reduced protein carbonyl (PC) content and upregulated superoxide dismutase (sod) gene expression in the muscle. As for muscle nutritional composition, 0.05–0.5 g/kg EGCG increased total amino acid (TAA) and flavor amino acid (FAA) contents, likely via the rapamycin (mTOR) signaling pathway. Muscle n-3 polyunsaturated fatty acids (n-3 PUFA) content was increased with 0.5 g/kg EGCG, presumably owing to the upregulation of fatty acyl elongase 5 (elovl5), long-chain fatty acyl-CoA synthetase 6 (acsl6), and peroxisome proliferator-actiated receptor α (pparα). Dietary EGCG (0.05–1 g/kg) significantly increased muscle hardness and chewiness, accompanied by an increase in myofiber density. EGCG supplementation (0.25–1 g/kg) increased the pH value and reduced lactate contents in the muscle. However, muscle crude lipid and hydroxyproline contents significantly decreased with 1 g/kg EGCG. Overall, quadratic regression analysis of WGR, SGR, TAA, and FAA showed that optimal EGCG (0.46–0.52 g/kg) dietary supplementation improved the growth and nutritional composition of juvenile common carp.

Phenome-wide association study of population-differentiating genetic variants around gene ACSL1.

Demographic dynamics and natural selection during human evolution shaped the present-day patterns of genetic variations, and geographically varying genetic factors contribute to different disease prevalences across human populations. This study aims to evaluate the presence of positive selection on the gene encoding long-chain fatty acyl-CoA synthetase 1 (ACSL1) and the phenotypic impacts of population-differentiating genetic variants around this gene. Three types of statistical tests for positive selection, based on site frequency spectrum, extended haplotype homozygosity and population differentiation, were applied to the whole-genome sequencing data from the 1000 Genomes Project. A phenome-wide association study of ACSL1 was performed with published genome-wide association studies (GWAS) and transcriptome-wide association studies, including phenome-wide studies in biobanks. Genetic variants associated with ACSL1 expression in various tissues exhibit geographically varying allele frequencies. Three types of statistical tests consistently supported the presence of positive selection on the coding and regulatory regions of ACSL1 in African, European, South Asian and East Asian populations. A phenome-wide association study of ACSL1 revealed associations with type 2 diabetes, blood glucose, age at menopause, mean platelet volume and mean reticulocyte volume. The top allele associated with lower diabetes risk has the highest frequency in European populations, whereas the top allele associated with later menopause has the highest frequency in African populations. Positive selection on ACSL1 resulted in geographically varying genetic variants, which may contribute to differential phenotypes across human populations, including type 2 diabetes and age at menopause.

An optimized fluorescent biosensor for monitoring long-chain fatty acyl-CoAs metabolism in vivo

Long-chain fatty acyl-CoAs (LCACoAs) are intermediates in lipid metabolism that exert a wide range of cellular functions. However, our knowledge about the subcellular distribution and regulatory impacts of LCACoAs is limited by a lack of methods for detecting LCACoAs in living cells and tissues. Here, we report our development of LACSerHR, a genetically encoded fluorescent biosensor that enables precise measurement of subtle fluctuations in the levels of endogenous LCACoAs in vivo. LACSerHR significantly improve the fluorescent brightness and analyte affinity, in vitro and in vivo testing showcased LACSerHR's large dynamic range. We demonstrate LACSerHR's capacity for real-time evaluation of LCACoA levels in specific subcellular compartments, for example in response to disruption of ACSL enzyme function in HEK293T cells. Moreover, we show the application of LACSerHR for sensitive measurement of elevated LCACoA levels in the livers of mouse models for two common metabolic diseases (NAFLD and type 2 diabetes). Thus, our LACSerHR sensor is a powerful, broadly applicable tool for studying LCACoAs metabolism and disease.

Fatty acids and inflammatory stimuli induce expression of long-chain acyl-CoA synthetase 1 to promote lipid remodeling in diabetic kidney disease

Fatty acid handling and complex lipid synthesis are altered in the kidney cortex of diabetic patients. We recently showed that inhibition of the renin-angiotensin system without changes in glycemia can reverse diabetic kidney disease (DKD) and restore the lipid metabolic network in the kidney cortex of diabetic (db/db) mice, raising the possibility that lipid remodeling may play a central role in DKD. However, the roles of specific enzymes involved in lipid remodeling in DKD have not been elucidated. In the present study, we used this diabetic mouse model and a proximal tubule epithelial cell line (HK2) to investigate the potential relationship between long-chain acyl-CoA synthetase 1 (ACSL1) and lipid metabolism in response to fatty acid exposure and inflammatory signals. We found ACSL1 expression was significantly increased in the kidney cortex of db/db mice, and exposure to palmitate (PA) or tumor necrosis factor (TNF-α) significantly increased Acsl1 mRNA expression in HK-2 cells. In addition, PA treatment significantly increased the levels of long-chain acylcarnitines and fatty acyl CoAs in HK2 cells, and these increases were abolished in HK2 cell lines with specific deletion of Acsl1(Acsl1KO cells), suggesting a key role for ACSL1 in fatty acid β-oxidation. In contrast, TNF-α treatment significantly increased the levels short-chain acylcarnitines and long-chain fatty acyl CoAs in HK2 cells but not in Acsl1KO cells, consistent with fatty acid channeling to complex lipids. Taken together, our data demonstrate a key role for ACSL1 in regulating lipid metabolism, fatty acid partitioning, and inflammation.

Avrupa Levreğinde (Dicentrarchus labrax) Karnitin Palmitoiltransferaz 1A (CPT1A) Geni Tanımlanması ve Karakterizasyonu

Objective: CPT1A intervenes in the access of the long-chain fatty acyl-CoA to the mitochondrial matrix. In this study, we conducted the genetic polymorphisms of the partial sequence of the CPT1A gene in European sea bass (Dicentrarchus labrax) that were cage cultured in the Aegean Sea.&#x0D; Material and Methods: Genomic DNA was isolated from 100 European sea bass individuals. After amplifying a partial region of the CPT1A gene by thermal cycler, PCR products were sequenced via Sanger method.&#x0D; Results: We detected two SNPs in partial sequence of the CPT1A gene in European sea bass. TT, TA and AA genotypes were observed for CPT1A g.2080T&gt;A locus with frequencies of 27.04, 49.92 and 23.04%, respectively. The frequencies of the g.2216A&gt;G locus as AA, AG and GG were found to be 1.2, 19.58 and 79.21%, respectively. The CPT1A g.2080T&gt;A locus was in HWE, whereas the g.2216A&gt;G locus was not in HWE.&#x0D; Conclusion: European sea bass has a high commercial value. These findings suggest that two SNPs in the CPT1A gene could be used for genomic selection programs related to fatty acid composition in European sea bass.&#x0D; Keywords: Fatty acid, SNP, sea bass, variation

Novel Effect of p-Coumaric Acid on Hepatic Lipolysis: Inhibition of Hepatic Lipid-Droplets.

p-coumaric acid (p-CA), a common plant phenolic acid with multiple bioactivities, has a lipid-lowering effect. As a dietary polyphenol, its low toxicity, with the advantages of prophylactic and long-term administration, makes it a potential drug for prophylaxis and the treatment of nonalcoholic fatty liver disease (NAFLD). However, the mechanism by which it regulates lipid metabolism is still unclear. In this study, we studied the effect of p-CA on the down-regulation of accumulated lipids in vivo and in vitro. p-CA increased a number of lipase expressions, including hormone-sensitive lipase (HSL), monoacylglycerol lipase (MGL) and hepatic triglyceride lipase (HTGL), as well as the expression of genes related to fatty acid oxidation, including long-chain fatty acyl-CoA synthetase 1 (ACSL1), carnitine palmitoyltransferase-1 (CPT1), by activating peroxisome proliferator-activated receptor α, and γ (PPARα and γ). Furthermore, p-CA promoted adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) phosphorylation and enhanced the expression of the mammalian suppressor of Sec4 (MSS4), a critical protein that can inhibit lipid droplet growth. Thus, p-CA can decrease lipid accumulation and inhibit lipid droplet fusion, which are correlated with the enhancement of liver lipases and genes related to fatty acid oxidation as an activator of PPARs. Therefore, p-CA is capable of regulating lipid metabolism and is a potential therapeutic drug or health care product for hyperlipidemia and fatty liver.

START domain-mediated nuclear translocation of Them1 regulates the expression of metabolic genes relative to thermogenesis in brown adipocytes

Thioesterase superfamily member 1 (Them1) is a long-chain fatty acyl-CoA thioesterase that is highly expressed in brown adipose tissue (BAT), where it limits energy expenditure. Them1 is comprised of tandem enzymatic domains and a C-terminal lipid-binding steroidogenic acute regulatory protein (StAR)-related lipid transfer (START) domain. Fatty acid binding to the START domain stabilizes the enzymatic domains and increases activity, whereas lysophosphatidylcholine (LPC) acts in the opposite manner. In brown adipocytes, Them1 localizes to puncta within the cytoplasm, but diffuses in response to β-adrenergic stimulation after which it is also found in the nucleus. The aim of this study was to determine the mechanism of Them1 nuclear trafficking and the metabolic consequences. METHODS: Identification of a putative nuclear localization signal (NLS) was determined by computational analysis. Full-length, NLS-deleted, or truncated Them1 cDNA constructs were cloned and linked to EGFP. Plasmid or adenoviral vectors were transfected into immortalized brown adipose cells (iBAs) that do not express Them1. Them1 was visualized by live cell confocal imaging via EGFP and with NucSpot nuclear staining. Signal quantification was performed with Imaris software. Them1 nuclear localization was evaluated in the presence or absence of norepinephrine (NE), leptomycin B, cycloheximide, or lipids that bind the START domain. Them1 nuclear localization was also quantified in BAT from mice treated with a β3-adrenergic receptor agonist, CL316,243. mRNA expression in iBAs expressing full-length or NLS-mutant Them1 in response to NE was determined by RNA-Seq. RESULTS: In the diffuse state, Them1 localized to the nucleus in a time-dependent manner. The NLS resides in a hinge region between the thioesterase domains and the START domain. Deletion of either the NLS or the START domain blocked nuclear targeting. Conversely, truncated Them1 containing only the START domain was targeted to the nucleus. Entry into the nucleus was balanced by trafficking out of the nucleus and new protein synthesis was not required, as evidenced by using leptomycin B to block nuclear efflux and cycloheximide to block protein synthesis, respectively. Incubation of NE-stimulated iBAs with LPC promoted nuclear Them1 translocation. Them1 protein was also detected in the nucleus of BAT from wild-type mice in response to CL administration. Nuclear Them1 localization in iBAs was associated with down-regulated mRNA expression of genes that control lipid and glucose metabolism during cold adaptation. CONCLUSIONS: Our data suggest that LPC generated during thermogenesis binds and destabilizes the START domain, leading to exposure of the NLS and nuclear trafficking of Them1 in response to β-adrenergic stimulation. Once inside the nucleus, Them1 regulates the transcription of lipid and glucose metabolic enzymes that are required for chronic cold adaptation. This work was supported by NIH DK103046 to DEC, SJH, and EAO This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Skeletal Muscle Consequences of Phosphatidylethanolamine Synthesis Deficiency.

The maintenance of phospholipid homeostasis is increasingly being implicated in metabolic health. Phosphatidylethanolamine (PE) is the most abundant phospholipid on the inner leaflet of cellular membranes, and we have previously shown that mice with a heterozygous ablation of the PE synthesizing enzyme, Pcyt2 (Pcyt2+/-), develop obesity, insulin resistance, and NASH. Skeletal muscle is a major determinant of systemic energy metabolism, making it a key player in metabolic disease development. Both the total PE levels and the ratio of PE to other membrane lipids in skeletal muscle are implicated in insulin resistance; however, the underlying mechanisms and the role of Pcyt2 regulation in this association remain unclear. Here, we show how reduced phospholipid synthesis due to Pcyt2 deficiency causes Pcyt2+/- skeletal muscle dysfunction and metabolic abnormalities. Pcyt2+/- skeletal muscle exhibits damage and degeneration, with skeletal muscle cell vacuolization, disordered sarcomeres, mitochondria ultrastructure irregularities and paucity, inflammation, and fibrosis. There is intramuscular adipose tissue accumulation, and major disturbances in lipid metabolism with impaired FA mobilization and oxidation, elevated lipogenesis, and long-chain fatty acyl-CoA, diacylglycerol, and triacylglycerol accumulation. Pcyt2+/- skeletal muscle exhibits perturbed glucose metabolism with elevated glycogen content, impaired insulin signaling, and reduced glucose uptake. Together, this study lends insight into the critical role of PE homeostasis in skeletal muscle metabolism and health with broad implications on metabolic disease development.

ACADL suppresses PD-L1 expression to prevent cancer immune evasion by targeting Hippo/YAP signaling in lung adenocarcinoma.

Lung cancer is the leading cause of cancer-related death. Cancer immune evasion is a key barrier in the treatment of lung cancer and the development of effective anticancer therapeutics. Long-chain Acyl-CoA dehydrogenase (ACADL), a key enzyme that regulates β-oxidation of long-chain fatty acyl-CoAs, has been found to act as a tumor suppressor in cancers. However, the role of ACADL in lung adenocarcinoma (LUAD) has not been explored. In the current study, we find that ACADL functions as a tumor suppressor in LUAD to inhibit proliferation and enhanced chemotherapeutic drug-induced apoptosis. Interestingly, ACADL prevents tumor immune evasion by suppressing PD-L1 expression in LUAD. ACADL is critical for Hippo/YAP pathway-mediated PD-L1 regulation. Moreover, YAP activation is essential for ACADL suppression of PD-L1 transcription. In addition, ACADL increases the protein stability and kinase activity of LATS kinase to inhibit YAP activation and PD-L1 transcription. Furthermore, we show that ACADL expression is positively correlated with a better OS and FP in LUAD. Our data reveals that ACADL could be a promising target for regulating Hippo/YAP pathway to prevent tumor immune evasion in LUAD.

Heterologous Production of Calendic Acid Naturally Found in Calendula officinalis by Recombinant Fission Yeast.

Calendic acid (CA) is a conjugated fatty acid with anti-cancer properties that is widely present in seed oil of Calendula officinalis. Using the co-expression of C. officinalis fatty acid conjugases (CoFADX-1 or CoFADX-2) and Punica granatum fatty acid desaturase (PgFAD2), we metabolically engineered the synthesis of CA in the yeast Schizosaccharomyces pombe without the need for linoleic acid (LA) supplementation. The highest CA titer and achieved accumulation were 4.4 mg/L and 3.7 mg/g of DCW in PgFAD2 + CoFADX-2 recombinant strain cultivated at 16 °C for 72 h, respectively. Further analyses revealed the accumulation of CA in free fatty acids (FFA) and downregulation of the lcf1 gene encoding long-chain fatty acyl-CoA synthetase. The developed recombinant yeast system represents an important tool for the future identification of the essential components of the channeling machinery to produce CA as a high-value conjugated fatty acid at an industrial level.

Cyclic stretch promotes vascular homing of endothelial progenitor cells via Acsl1 regulation of mitochondrial fatty acid oxidation

Endothelial progenitor cells (EPCs) play an important role in vascular repair and re-endothelialization after vessel injury. EPCs in blood vessels are subjected to cyclic stretch (CS) due to the pulsatile pressure, but the role of CS in metabolic reprogramming of EPC, particularly its vascular homing and repair, is largely unknown. In the current study, physiological CS applied to EPCs at a magnitude of 10% and a frequency of 1 Hz significantly promoted their vascular adhesion and endothelial differentiation. CS enhanced mitochondrial elongation and oxidative phosphorylation (OXPHOS), as well as adenosine triphosphateproduction. Metabolomic study and Ultra-high performance liquid chromatography-mass spectrometry assay revealed that CS significantly decreased the content of long-chain fatty acids (LCFAs) and markedly induced long-chain fatty acyl-CoA synthetase 1 (Acsl1), which in turn facilitated the catabolism of LCFAs in mitochondria via fatty acid β-oxidation and OXPHOS. In a rat carotid artery injury model, transplantation of EPCs overexpressing Acsl1 enhanced the adhesion and re-endothelialization of EPCs invivo. MRI and vascular morphology staining showed that Acsl1 overexpression in EPCs improved vascular repair and inhibited vascular stenosis. This study reveals a mechanotransduction mechanism by which physiological CS enhances endothelial repair via EPC patency.