LPCAT3/LPLAT12 deficiency in the liver ameliorates acetaminophen-induced acute liver injury.

Acetaminophen (APAP) is a double-edged sword, mainly depending on the dosage. A moderate dose of APAP is effective for fever and pain relief; however, an overdose induces acute liver injury. The mechanism underlying APAP-induced acute liver failure is unclear, and its treatment is limited. A recent report has shown that several oxidized phospholipids are associated with APAP-induced acute liver failure. Lysophosphatidylcholine acyltransferase 3 (Lpcat3, Lplat12), which is highly expressed in the liver, preferentially catalyzes the incorporation of arachidonate into lysophospholipids (PLs). In the present study, we investigated the roles of Lpcat3 on APAP-induced acute liver injury using liver-specific Lpcat3-knockout mice. Hepatic Lpcat3 deficiency reduced the degree of APAP-induced necrosis of hepatocytes around Zone 3 and ameliorated the elevation of hepatic injury serum marker levels, and prolonged survival. Lipidomic analysis showed that the accumulation of oxidized and hydroperoxidized phospholipids was suppressed in Lpcat3-knockout mice. The amelioration of APAP-induced acute liver injury was due not only to the reduction in the lipid synthesis of arachidonic acid PLs because of Lpcat3 deficiency, but also to the promotion of the APAP detoxification pathway by facilitating the conjugation of glutathione and N-acetyl-p-benzoquinone imine. Our findings suggest that Lpcat3 is a potential therapeutic target for treating APAP-induced acute liver injury.

Overexpression of lysophospholipid acyltransferase, LPLAT10/LPCAT4/LPEAT2, in the mouse liver increases glucose-stimulated insulin secretion.

Postprandial hyperglycemia is an early indicator of impaired glucose tolerance that leads to type 2 diabetes mellitus (T2DM). Alterations in the fatty acid composition of phospholipids have been implicated in diseases such as T2DM and nonalcoholic fatty liver disease. Lysophospholipid acyltransferase 10 (LPLAT10, also called LPCAT4 and LPEAT2) plays a role in remodeling fatty acyl chains of phospholipids; however, its relationship with metabolic diseases has not been fully elucidated. LPLAT10 expression is low in the liver, the main organ that regulates metabolism, under normal conditions. Here, we investigated whether overexpression of LPLAT10 in the liver leads to improved glucose metabolism. For overexpression, we generated an LPLAT10-expressing adenovirus (Ad) vector (Ad-LPLAT10) using an improved Ad vector. Postprandial hyperglycemia was suppressed by the induction of glucose-stimulated insulin secretion in Ad-LPLAT10-treated mice compared with that in control Ad vector-treated mice. Hepatic and serum levels of phosphatidylcholine 40:7, containing C18:1 and C22:6, were increased in Ad-LPLAT10-treated mice. Serum from Ad-LPLAT10-treated mice showed increased glucose-stimulated insulin secretion in mouse insulinoma MIN6 cells. These results indicate that changes in hepatic phosphatidylcholine species due to liver-specific LPLAT10 overexpression affect the pancreas and increase glucose-stimulated insulin secretion. Our findings highlight LPLAT10 as a potential novel therapeutic target for T2DM.

Lysophospholipid Acyltransferase 9 Promotes Emphysema Formation via Platelet-activating Factor.

Cigarette smoking is known to be the leading cause of chronic obstructive pulmonary disease (COPD). However, the detailed mechanisms have not been elucidated. PAF (platelet-activating factor), a potent inflammatory mediator, is involved in the pathogenesis of various respiratory diseases such as bronchial asthma and COPD. We focused on LPLAT9 (lysophospholipid acyltransferase 9), a biosynthetic enzyme of PAF, in the pathogenesis of COPD. LPLAT9 gene expression was observed in excised COPD lungs and single-cell RNA sequencing data of alveolar macrophages (AMs). LPLAT9 was predominant and upregulated in AMs, particularly monocyte-derived AMs, in patients with COPD. To identify the function of LPLAT9/PAF in AMs in the pathogenesis of COPD, we exposed systemic LPLAT9-knockout (LPALT9-/-) mice to cigarette smoke (CS). CS increased the number of AMs, especially the monocyte-derived fraction, which secreted MMP12 (matrix metalloprotease 12). Also, CS augmented LPLAT9 phosphorylation/activation on macrophages and, subsequently, PAF synthesis in the lung. The LPLAT9-/- mouse lung showed reduced PAF production after CS exposure. Intratracheal PAF administration accumulated AMs by increasing MCP1 (monocyte chemoattractant protein-1). After CS exposure, AM accumulation and subsequent pulmonary emphysema, a primary pathologic change of COPD, were reduced in LPALT9-/- mice compared with LPLAT9+/+ mice. Notably, these phenotypes were again worsened by LPLAT9+/+ bone marrow transplantation in LPALT9-/- mice. Thus, CS-induced LPLAT9 activation in monocyte-derived AMs aggravated pulmonary emphysema via PAF-induced further accumulation of AMs. These results suggest that PAF synthesized by LPLAT9 has an important role in the pathogenesis of COPD.

LPGAT1/LPLAT7 regulates acyl chain profiles at the sn-1 position of phospholipids in murine skeletal muscles.

Skeletal muscle consists of both fast- and slow-twitch fibers. Phospholipids are important structural components of cellular membranes, and the diversity of their fatty acid composition affects membrane characteristics. Although some studies have shown that acyl chain species in phospholipids differ among various muscle fiber types, the mechanisms underlying these differences are unclear. To investigate this, we analyzed phosphatidylcholine (PC) and phosphatidylethanolamine (PE) molecules in the murine extensor digitorum longus (EDL; fast-twitch) and soleus (slow-twitch) muscles. In the EDL muscle, the vast majority (93.6%) of PC molecules was palmitate-containing PC (16:0-PC), whereas in the soleus muscle, in addition to 16:0-PC, 27.9% of PC molecules was stearate-containing PC (18:0-PC). Most palmitate and stearate were bound at the sn-1 position of 16:0- and 18:0-PC, respectively, and 18:0-PC was found in type I and IIa fibers. The amount of 18:0-PE was higher in the soleus than in the EDL muscle. Peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) increased the amount of 18:0-PC in the EDL. Lysophosphatidylglycerol acyltransferase 1 (LPGAT1) was highly expressed in the soleus compared with that in the EDL muscle and was upregulated by PGC-1α. LPGAT1 knockout decreased the incorporation of stearate into PC and PE in vitro and ex vivo and the amount of 18:0-PC and 18:0-PE in murine skeletal muscle with an increase in the level of 16:0-PC and 16:0-PE. Moreover, knocking out LPGAT1 decreased the amount of stearate-containing phosphatidylserine (18:0-PS), suggesting that LPGAT1 regulated the acyl chain profiles of phospholipids, namely, PC, PE, and PS, in the skeletal muscle.

Visualization of Phospholipid Synthesis on Tissue Sections Using Functional Mass Spectrometry Imaging.

Functional mass spectrometry imaging (fMSI) is a potent tool for elucidating the spatial distribution of enzyme activities in tissues at high resolution. In this study, we applied fMSI to probe the intricate biosynthesis of phospholipids, which exist as thousands of molecular species in tissues and exhibit a unique distribution specific to cell type. By using deuterium- and 13C-labeled substrates, we visualized the activities of key enzymes involved in phospholipid synthesis, including glycerol 3-phosphate acyltransferase (GPAT), lysophosphatidic acid acyltransferases (LPAAT), lysophospholipid acyltransferases (LPLAT), and long-chain acyl-CoA synthetase (ACSL). Additionally, we were able to visualize a two-step sequential enzyme reaction involving ACSL and LPLAT. This novel approach unveiled significant variations in enzyme activity distribution depending on the type of fatty acids used as substrates. It will also help to reveal the mechanisms underlying the formation of numerous phospholipid species.

Multiomics identifies the link between intratumor steatosis and the exhausted tumor immune microenvironment in hepatocellular carcinoma.

BACKGROUND AND AIMS: Immunotherapy has become the standard-of-care treatment for hepatocellular carcinoma (HCC), but its efficacy remains limited. To identify immunotherapy-susceptible HCC, we profiled the molecular abnormalities and tumor immune microenvironment (TIME) of rapidly increasing nonviral HCC. APPROACHES AND RESULTS: We performed RNA-seq of tumor tissues in 113 patients with nonviral HCC and cancer genome sequencing of 69 genes with recurrent genetic alterations reported in HCC. Unsupervised hierarchical clustering classified nonviral HCCs into three molecular classes (Class I, II, III), which stratified patient prognosis. Class I, with the poorest prognosis, was associated with TP53 mutations, whereas class III, with the best prognosis, was associated with cadherin-associated protein beta 1 (CTNNB1) mutations. Thirty-eight percent of nonviral HCC was defined as an immune class characterized by a high frequency of intratumoral steatosis and a low frequency of CTNNB1 mutations. Steatotic HCC, which accounts for 23% of nonviral HCC cases, presented an immune-enriched but immune-exhausted TIME characterized by T cell exhaustion, M2 macrophage and cancer-associated fibroblast (CAF) infiltration, high PD-L1 expression, and TGF-ß signaling activation. Spatial transcriptome analysis suggested that M2 macrophages and CAFs may be in close proximity to exhausted CD8+ T cells in steatotic HCC. An in vitro study showed that palmitic acid-induced lipid accumulation in HCC cells upregulated PD-L1 expression and promoted immunosuppressive phenotypes of cocultured macrophages and fibroblasts. Patients with steatotic HCC, confirmed by chemical-shift MR imaging, had significantly longer PFS with combined immunotherapy using anti-PD-L1 and anti-VEGF antibodies. CONCLUSIONS: Multiomics stratified nonviral HCCs according to prognosis or TIME. We identified the link between intratumoral steatosis and immune-exhausted immunotherapy-susceptible TIME.

Constitutively active GPR43 is crucial for proper leukocyte differentiation.

The G protein-coupled receptors, GPR43 (free fatty acid receptor 2, FFA2) and GPR41 (free fatty acid receptor 3, FFA3), are activated by short-chain fatty acids produced under various conditions, including microbial fermentation of carbohydrates. Previous studies have implicated this receptor energy homeostasis and immune responses as well as in cell growth arrest and apoptosis. Here, we observed the expression of both receptors in human blood cells and a remarkable enhancement in leukemia cell lines (HL-60, U937, and THP-1 cells) during differentiation. A reporter assay revealed that GPR43 is coupled with Gαi and Gα12/13 and is constitutively active without any stimuli. Specific blockers of GPR43, GLPG0974 and CATPB function as inverse agonists because treatment with these compounds significantly reduces constitutive activity. In HL-60 cells, enhanced expression of GPR43 led to growth arrest through Gα12/13 . In addition, the blockage of GPR43 activity in these cells significantly impaired their adherent properties due to the reduction of adhesion molecules. We further revealed that enhanced GPR43 activity induces F-actin formation. However, the activity of GPR43 did not contribute to butyrate-induced apoptosis in differentiated HL-60 cells because of the ineffectiveness of the inverse agonist on cell death. Collectively, these results suggest that GPR43, which possesses constitutive activity, is crucial for growth arrest, followed by the proper differentiation of leukocytes.

Lysophosphatidylcholine acyltransferase 1 controls mitochondrial reactive oxygen species generation and survival of retinal photoreceptor cells.

Due to their high energy demands and characteristic morphology, retinal photoreceptor cells require a specialized lipid metabolism for survival and function. Accordingly, dysregulation of lipid metabolism leads to the photoreceptor cell death and retinal degeneration. Mice bearing a frameshift mutation in the gene encoding lysophosphatidylcholine acyltransferase 1 (Lpcat1), which produces saturated phosphatidylcholine (PC) composed of two saturated fatty acids, has been reported to cause spontaneous retinal degeneration in mice; however, the mechanism by which this mutation affects degeneration is unclear. In this study, we performed a detailed characterization of LPCAT1 in the retina and found that genetic deletion of Lpcat1 induces light-independent and photoreceptor-specific apoptosis in mice. Lipidomic analyses of the retina and isolated photoreceptor outer segment (OS) suggested that loss of Lpcat1 not only decreased saturated PC production but also affected membrane lipid composition, presumably by altering saturated fatty acyl-CoA availability. Furthermore, we demonstrated that Lpcat1 deletion led to increased mitochondrial reactive oxygen species levels in photoreceptor cells, but not in other retinal cells, and did not affect the OS structure or trafficking of OS-localized proteins. These results suggest that the LPCAT1-dependent production of saturated PC plays critical roles in photoreceptor maturation. Our findings highlight the therapeutic potential of saturated fatty acid metabolism in photoreceptor cell degeneration-related retinal diseases.

Update and nomenclature proposal for mammalian lysophospholipid acyltransferases, which create membrane phospholipid diversity.

The diversity of glycerophospholipid species in cellular membranes is immense and affects various biological functions. Glycerol-3-phosphate acyltransferases (GPATs) and lysophospholipid acyltransferases (LPLATs), in concert with phospholipase A1/2s enzymes, contribute to this diversity via selective esterification of fatty acyl chains at the sn-1 or sn-2 positions of membrane phospholipids. These enzymes are conserved across all kingdoms, and in mammals four GPATs of the 1-acylglycerol-3-phosphate O-acyltransferase (AGPAT) family and at least 14 LPLATs, either of the AGPAT or the membrane-bound O-acyltransferase (MBOAT) families, have been identified. Here we provide an overview of the biochemical and biological activities of these mammalian enzymes, including their predicted structures, involvements in human diseases, and essential physiological roles as revealed by gene-deficient mice. Recently, the nomenclature used to refer to these enzymes has generated some confusion due to the use of multiple names to refer to the same enzyme and instances of the same name being used to refer to completely different enzymes. Thus, this review proposes a more uniform LPLAT enzyme nomenclature, as well as providing an update of recent advances made in the study of LPLATs, continuing from our JBC mini review in 2009.

Dietary omega-3 fatty acid does not improve male infertility caused by lysophospholipid acyltransferase 3 (LPLAT3/AGPAT3) deficiency.

Docosahexaenoic acid (DHA), an omega-3 fatty acid, usually presents as a constituent of phospholipids in the cellular membrane. Lysophospholipid acyltransferase 3 (LPLAT3; AGPAT3) is the primary enzyme that incorporates DHA into phospholipids. LPLAT3-KO mice show male infertility and visual dysfunction accompanied by decreased phospholipids (PLs) containing DHA (PL-DHA) in the testis and retina, respectively. In this study, we evaluated the effect of diets consisting mainly of triacylglycerol-bound DHA (fish oil) and PL-bound DHA (salmon roe oil) on the amount of PL-DHA in a broad range of tissues and on reproductive functions. Both diets elevated phosphatidylcholines (PCs)-containing DHA in most tissues of wild type (WT) mice. Although LPLAT3-KO mice acquired a minimal amount of PC-DHA in the testes and sperm by eating either of the diets, reproductive function did not improve. The present study suggests that DHA-rich diets do not restore sufficient PL-DHA to improve male infertility in LPLAT3-KO mice. Alternatively, PL-DHA can be biosynthesized by LPLAT3 but not by external supplementation, which may be necessary for normal reproductive function.

Sphingosine-1-phosphate promotes tumor development and liver fibrosis in mouse model of congestive hepatopathy.

BACKGROUND AND AIMS: Chronic liver congestion reflecting right-sided heart failure (RHF), Budd-Chiari syndrome, or Fontan-associated liver disease (FALD) is involved in liver fibrosis and HCC. However, molecular mechanisms of fibrosis and HCC in chronic liver congestion remain poorly understood. APPROACH AND RESULTS: Here, we first demonstrated that chronic liver congestion promoted HCC and metastatic liver tumor growth using murine model of chronic liver congestion by partial inferior vena cava ligation (pIVCL). As the initial step triggering HCC promotion and fibrosis, gut-derived lipopolysaccharide (LPS) appeared to induce LSECs capillarization in mice and in vitro. LSEC capillarization was also confirmed in patients with FALD. Mitogenic factor, sphingosine-1-phosphate (S1P), was increased in congestive liver and expression of sphingosine kinase 1, a major synthetase of S1P, was increased in capillarized LSECs after pIVCL. Inhibition of S1P receptor (S1PR) 1 (Ex26) and S1PR2 (JTE013) mitigated HCC development and liver fibrosis, respectively. Antimicrobial treatment lowered portal blood LPS concentration, LSEC capillarization, and liver S1P concentration accompanied by reduction of HCC development and fibrosis in the congestive liver. CONCLUSIONS: In conclusion, chronic liver congestion promotes HCC development and liver fibrosis by S1P production from LPS-induced capillarized LSECs. Careful treatment of both RHF and liver cancer might be necessary for patients with RHF with primary or metastatic liver cancer.

Lysophosphatidic acid receptor1/3 antagonist inhibits the activation of satellite glial cells and reduces acute nociceptive responses.

Lysophosphatidic acid (LPA) exerts various biological activities through six characterized G protein-coupled receptors (LPA1-6 ). While LPA-LPA1  signaling contributes toward the demyelination and retraction of C-fiber and induces neuropathic pain, the effects of LPA-LPA1  signaling on acute nociceptive pain is uncertain. This study investigated the role of LPA-LPA1  signaling in acute nociceptive pain using the formalin test. The pharmacological inhibition of the LPA-LPA1 axis significantly attenuated formalin-induced nociceptive behavior. The LPA1  mRNA was expressed in satellite glial cells (SGCs) in dorsal root ganglion (DRG) and was particularly abundant in SGCs surrounding large DRG neurons, which express neurofilament 200. Treatment with LPA1/3 receptor (LPA1/3 ) antagonist inhibited the upregulation of glial markers and inflammatory cytokines in DRG following formalin injection. The LPA1/3 antagonist also attenuated phosphorylation of extracellular signal-regulated kinase, especially in SGCs and cyclic AMP response element-binding protein in the dorsal horn following formalin injection. LPA amounts after formalin injection to the footpad were quantified by liquid chromatography/tandem mass spectrometry, and LPA levels were found to be increased in the innervated DRGs. Our results indicate that LPA produced in the innervated DRGs promotes the activation of SGCs through LPA1 , increases the sensitivity of primary neurons, and modulates pain behavior. These results facilitate our understanding of the pathology of acute nociceptive pain and demonstrate the possibility of the LPA1 on SGCs as a novel target for acute pain control.

Identification and characterization of LPLAT7 as an sn-1-specific lysophospholipid acyltransferase.

The main fatty acids at the sn-1 position of phospholipids (PLs) are saturated or monounsaturated fatty acids such as palmitic acid (C16:0), stearic acid (C18:0), and oleic acid (C18:1) and are constantly replaced, like unsaturated fatty acids at the sn-2 position. However, little is known about the molecular mechanism underlying the replacement of fatty acids at the sn-1 position, i.e., the sn-1 remodeling. Previously, we established a method to evaluate the incorporation of fatty acids into the sn-1 position of lysophospholipids (lyso-PLs). Here, we used this method to identify the enzymes capable of incorporating fatty acids into the sn-1 position of lyso-PLs (sn-1 lysophospholipid acyltransferase [LPLAT]). Screenings using siRNA knockdown and recombinant proteins for 14 LPLATs identified LPLAT7/lysophosphatidylglycerol acyltransferase 1 (LPGAT1) as a candidate. In vitro, we found LPLAT7 mainly incorporated several fatty acids into the sn-1 position of lysophosphatidylcholine (LPC) and lysophosphatidylethanolamine (LPE), with weak activities toward other lyso-PLs. Interestingly, however, only C18:0-containing phosphatidylcholine (PC) and phosphatidylethanolamine (PE) were specifically reduced in the LPLAT7-mutant cells and tissues from knockout mice, with a concomitant increase in the level of C16:0- and C18:1-containing PC and PE. Consistent with this, the incorporation of deuterium-labeled C18:0 into PLs dramatically decreased in the mutant cells, while deuterium-labeled C16:0 and C18:1 showed the opposite dynamic. Identifying LPLAT7 as an sn-1 LPLAT facilitates understanding the biological significance of sn-1 fatty acid remodeling of PLs. We also propose to use the new nomenclature, LPLAT7, for LPGAT1 since the newly assigned enzymatic activities are quite different from the LPGAT1s previously reported.

Mapping membrane lipids in the developing and adult mouse retina under physiological and pathological conditions using mass spectrometry.

Membrane phospholipids play pivotal roles in various cellular processes, and their levels are tightly regulated. In the retina, phospholipids had been scrutinized because of their distinct composition and requirement in visual transduction. However, how lipid composition changes during retinal development remains unclear. Here, we used liquid chromatography-mass spectrometry (LC-MS) to assess the dynamic changes in the levels of two main glycerophospholipids, phosphatidylcholine (PC) and phosphatidylethanolamine (PE), in the developing mouse retina under physiological and pathological conditions. The total levels of PC and PE increased during retinal development, and individual lipid species exhibited distinct level changes. The amount of very-long-chain PC and PE increased dramatically in the late stages of retinal development. The mRNA levels of Elovl2 and Elovl4, genes encoding enzymes essential for the synthesis of very-long-chain polyunsaturated fatty acids, increased in developing photoreceptors. Cell sorting based on CD73 expression followed by LC-MS revealed distinct changes in PC and PE levels in CD73-positive rod photoreceptors and CD73-negative retinal cells. Finally, using the NaIO3-induced photoreceptor degeneration model, we identified photoreceptor-specific changes in PC and PE levels from 1 day after NaIO3 administration, before the outer segment of photoreceptors displayed morphological impairment. In conclusion, our findings provide insight into the dynamic changes in PC and PE levels in the developing and adult mouse retina under physiological and pathological conditions. Furthermore, we provide evidence that cell sorting followed by LC-MS is a promising approach for investigating the relevance of lipid homeostasis in the function of different retinal cell types.

α-Synuclein V15A Variant in Familial Parkinson's Disease Exhibits a Weaker Lipid-Binding Property.

BACKGROUND: The α-Synuclein (α-Syn) V15A variant has been found in two Caucasian families with Parkinson's disease (PD). However, the significance of this missense variant remained unclear. OBJECTIVE: We sought to elucidate whether V15A could increase aggregation or change phospholipid affinity. METHODS: A sequencing analysis for the SNCA encoding α-Syn from 875 patients with PD and 324 control subjects was performed. Comparing with known pathogenic missense variants of α-Syn, A30P, and A53T, we analyzed the effects of V15A on binding to phospholipid membrane, self-aggregation, and seed-dependent aggregation in cultured cells. RESULTS: Genetic screening identified SNCA c.44 T>C (p.V15A) from two Japanese PD families. The missense variant V15A was extremely rare in several public databases and predicted as pathogenic using in silico tools. The amplification activity of α-Syn V15A fibrils was stronger than that of wild-type α-Syn fibrils. CONCLUSIONS: The discovery of the V15A variant from Japanese families reinforces the possibility that the V15A variant may be a causative variant for developing PD. V15A had a reduced affinity for phospholipids and increased propagation activity compared with wild-type. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Mutagenesis and homology modeling reveal a predicted pocket of lysophosphatidylcholine acyltransferase 2 to catch Acyl-CoA.

Platelet-activating factor (PAF) is a potent proinflammatory phospholipid mediator that elicits various cellular functions and promotes several pathological events, including anaphylaxis and neuropathic pain. PAF is biosynthesized by two types of lyso-PAF acetyltransferases: lysophosphatidylcholine acyltransferase 1 (LPCAT1) and LPCAT2, which are constitutive and inducible forms of lyso-PAF acetyltransferase, respectively. Because LPCAT2 mainly produces PAF under inflammatory stimuli, understanding the structure of LPCAT2 is important for developing specific drugs against PAF-related inflammatory diseases. Although the structure of LPCAT2 has not been determined, the crystal structure was reported for Thermotoga maritima PlsC, an enzyme in the same gene family as LPCAT2. Here, we identified residues in mouse LPCAT2 essential for its enzymatic activity and a potential acyl-coenzyme A (CoA)-binding pocket, based on homology modeling of mouse LPCAT2 with PlsC. We also found that Ala115 of mouse LPCAT2 was important for acyl-CoA selectivity. In conclusion, these results predict the three-dimensional (3D) structure of mouse LPCAT2. Our findings have implications for the future development of new drugs against PAF-related diseases.

Parkinson's disease-associated iPLA2-VIA/PLA2G6 regulates neuronal functions and α-synuclein stability through membrane remodeling.

Mutations in the iPLA2-VIA/PLA2G6 gene are responsible for PARK14-linked Parkinson's disease (PD) with α-synucleinopathy. However, it is unclear how iPLA2-VIA mutations lead to α-synuclein (α-Syn) aggregation and dopaminergic (DA) neurodegeneration. Here, we report that iPLA2-VIA-deficient Drosophila exhibits defects in neurotransmission during early developmental stages and progressive cell loss throughout the brain, including degeneration of the DA neurons. Lipid analysis of brain tissues reveals that the acyl-chain length of phospholipids is shortened by iPLA2-VIA loss, which causes endoplasmic reticulum (ER) stress through membrane lipid disequilibrium. The introduction of wild-type human iPLA2-VIA or the mitochondria-ER contact site-resident protein C19orf12 in iPLA2-VIA-deficient flies rescues the phenotypes associated with altered lipid composition, ER stress, and DA neurodegeneration, whereas the introduction of a disease-associated missense mutant, iPLA2-VIA A80T, fails to suppress these phenotypes. The acceleration of α-Syn aggregation by iPLA2-VIA loss is suppressed by the administration of linoleic acid, correcting the brain lipid composition. Our findings suggest that membrane remodeling by iPLA2-VIA is required for the survival of DA neurons and α-Syn stability.

Multiplex fatty acid imaging inside cells by Raman microscopy.

Visualizing intracellular fatty acids (including free and esterified form) is very useful for understanding how and where such molecules are incorporated, stored, and metabolized within cells. However, techniques of imaging multiple intracellular fatty acids have been limited by their small size, making it difficult to label and track without changing their biological and biophysical characteristics. Here, we present a new method for simultaneously visualizing up to five atomically labeled intracellular fatty acid species. For this, we utilized the distinctive Raman spectra depending on the labeling patterns and created a new, extensible opensource software to perform by-pixel analysis of extracting original spectra from mixed ones. Our multiplex imaging method revealed that fatty acids with more double bonds tend to concentrate more efficiently at lipid droplets. This novel approach contributes to reveal not only the spatial dynamics of fatty acids, but also of any other metabolites inside cells.

Lysophosphatidylethanolamine acyltransferase 2 (LPEAT2) incorporates DHA into phospholipids and has possible functions for fatty acid-induced cell death.

Glycerophospholipids, one of the main constituents of biological membranes, are synthesized from glycerol-3-phosphate through the de novo pathway, and are reconstituted through the remodeling pathway. Lysophosphatidylethanolamine acyltransferase 2 (LPEAT2), one of the enzymes that play a role in the remodeling pathway, has been previously reported to have LPEAT, lysophosphatidylcholine acyltransferase (LPCAT) and lysophosphatidylglycerol acyltransferase (LPGAT) activities with 16:0-CoA, 18:0-CoA, and 18:1-CoA as donors. In this study, we found that LPEAT2 is active with 22:6-CoA. Knockdown studies using Neuro 2A cells showed that LPEAT2 has endogenous LPEAT activity with 22:6-CoA, and that LPEAT2 has functions for modulating 22:6/20:4 ratios of phospholipids. In addition, we demonstrated that Neuro 2A cells overexpressing LPEAT2 underwent cell death with necrotic morphology when differentiated into neuron-like cells, with supplementation with 22:6 (DHA). These results suggest that LPEAT2 plays a role in inducing cell death DHA-dependently. This study will lead to better understand how DHA levels are regulated in phospholipids, especially in the brain where LPEAT2 is highly expressed. Our study also provides insight to understand the mechanism of cell death induced by DHA.

Conformation of ultra-long-chain fatty acid in lipid bilayer: Molecular dynamics study.

Ultra-long-chain fatty acids (ULCFAs) are biosynthesized in the restricted tissues such as retina, testis, and skin. The conformation of a single ULCFA, in which the sn-1 unsaturated chain has 32 carbons, in three types of phospholipid bilayers is studied by molecular dynamics simulations. It is found that the ultra-long tail of the ULCFA flips between two leaflets and fluctuates among an elongation into the opposite leaflet, lies between two leaflets, and turns back. As the number ratio of lipids in the opposite leaflet increases, the ratio of the elongated shape linearly decreases in all three cases. Thus, ULCFAs can sense the density differences between the two leaflets and respond to these changes.