Phosphatidylethanolamine Biosynthesis Research Articles

A mutation of EPT1 (SELENOI) underlies a new disorder of Kennedy pathway phospholipid biosynthesis.

Mutations in genes involved in lipid metabolism have increasingly been associated with various subtypes of hereditary spastic paraplegia, a highly heterogeneous group of neurodegenerative motor neuron disorders characterized by spastic paraparesis. Here, we report an unusual autosomal recessive neurodegenerative condition, best classified as a complicated form of hereditary spastic paraplegia, associated with mutation in the ethanolaminephosphotransferase 1 (EPT1) gene (now known as SELENOI), responsible for the final step in Kennedy pathway forming phosphatidylethanolamine from CDP-ethanolamine. Phosphatidylethanolamine is a glycerophospholipid that, together with phosphatidylcholine, constitutes more than half of the total phospholipids in eukaryotic cell membranes. We determined that the mutation defined dramatically reduces the enzymatic activity of EPT1, thereby hindering the final step in phosphatidylethanolamine synthesis. Additionally, due to central nervous system inaccessibility we undertook quantification of phosphatidylethanolamine levels and species in patient and control blood samples as an indication of liver phosphatidylethanolamine biosynthesis. Although this revealed alteration to levels of specific phosphatidylethanolamine fatty acyl species in patients, overall phosphatidylethanolamine levels were broadly unaffected indicating that in blood EPT1 inactivity may be compensated for, in part, via alternate biochemical pathways. These studies define the first human disorder arising due to defective CDP-ethanolamine biosynthesis and provide new insight into the role of Kennedy pathway components in human neurological function.

Phosphatidylethanolamine Deficiency Impairs Escherichia coli Adhesion by Downregulating Lipopolysaccharide Synthesis, Which is Reversible by High Galactose/Lactose Cultivation.

As the initiation step of bacterial infection or biofouling, bacterial adhesion on cells or substrates is generally an optimal target for antibacterial design. Phosphatidylethanolamine (PE) is the principal phospholipid in bacteria, and its function in bacterial adhesion remains unclear. In this study, four E. coli strains including two PE-deficient mutants (PE-PC- and PE-PC+ strains) and two PE-containing wild-type controls (PE + PC- strains) were recruited to investigate the influence of PE deficiency on bacterial adhesion. We found that PE deficiency could impair E. coli adhesion on macrophages (human THP-1-derived and mouse RAW264.7 macrophages) or glass coverslips by downregulating lipopolysaccharide (LPS) biosynthesis, which could be reversible by high galactose/lactose but not glucose cultivation. The data imply that PE play important role in bacterial adhesion probably via affecting LPS biosynthesis and suggest that targeting PE biosynthesis is also a potential antibacterial strategy.

Genome-wide transcriptome analysis revealed organelle specific responses to temperature variations in algae.

Temperature is a critical environmental factor that affects microalgal growth. However, microalgal coping mechanisms for temperature variations are unclear. Here, we determined changes in transcriptome, total carbohydrate, total fatty acid methyl ester, and fatty acid composition of Tetraselmis sp. KCTC12432BP, a strain with a broad temperature tolerance range, to elucidate the tolerance mechanisms in response to large temperature variations. Owing to unavailability of genome sequence information, de novo transcriptome assembly coupled with BLAST analysis was performed using strand specific RNA-seq data. This resulted in 26,245 protein-coding transcripts, of which 83.7% could be annotated to putative functions. We identified more than 681 genes differentially expressed, suggesting an organelle-specific response to temperature variation. Among these, the genes related to the photosynthetic electron transfer chain, which are localized in the plastid thylakoid membrane, were upregulated at low temperature. However, the transcripts related to the electron transport chain and biosynthesis of phosphatidylethanolamine localized in mitochondria were upregulated at high temperature. These results show that the low energy uptake by repressed photosynthesis under low and high temperature conditions is compensated by different mechanisms, including photosystem I and mitochondrial oxidative phosphorylation, respectively. This study illustrates that microalgae tolerate different temperature conditions through organelle specific mechanisms.

Impaired biosynthesis of the non-bilayer lipids phosphatidylethanolamine or cardiolipin does not affect peroxisome biogenesis and proliferation in Saccharomyces cerevisiae

The non-bilayer forming lipids cardiolipin (CL) and phosphatidylethanolamine (PE) modulate membrane curvature, facilitate membrane fusion and affect the stability and function of membrane proteins. Yeast peroxisomal membranes contain significant amounts of CL and PE.We analysed the effect of CL deficiency and PE depletion on peroxisome biogenesis and proliferation in Saccharomyces cerevisiae. Our data indicate that deletion of CRD1, which encodes cardiolipin synthase, does not affect peroxisome biogenesis or abundance, both at peroxisome repressing (glucose) or inducing (oleate) growth conditions. Analysis of strains deficient in one of the three PE biosynthesis pathways (psd1, psd2 or the triple deletion strain eki1 cki1 dpl1) revealed that in all three strains peroxisome numbers were reduced upon growth of cells on oleic acid, whereas the psd1 strain also showed a reduction in peroxisome abundance upon growth on glucose. Because PE is an intermediate of the phosphatidylcholine (PC) biosynthesis pathway, PE depletion affects PC formation. PC however can be synthesized by an alternative pathway when choline is supplemented to the growth medium. Because the addition of choline resulted in suppression of the peroxisome phenotypes in phosphatidylserine decarboxylase mutant strains, we conclude that peroxisome biogenesis and proliferation are not crucially dependent on CL or PE.

Mitochondrial acyltransferases and glycerophospholipid metabolism

Our understanding of the synthesis and remodeling of mitochondrial phospholipids remains incomplete. Two isoforms of glycerol-3-phosphate acyltransferase (GPAT1 and 2) and two isoforms of acylglycerol-3-phosphate acyltransferase (AGPAT4 and 5) are located on the outer mitochondrial membrane, suggesting that both lysophosphatidic acid and phosphatidic acid are synthesized in situ for de novo glycerolipid biosynthesis. However, it is believed that the phosphatidic acid substrate for cardiolipin and phosphatidylethanolamine biosynthesis is produced at the endoplasmic reticulum whereas the phosphatidic acid synthesized in the mitochondria must be transferred to the endoplasmic reticulum before it undergoes additional steps to form the mature phospholipids that are trafficked back to the mitochondria. It is unclear whether mitochondrial phospholipids are remodeled by mitochondrial acyltransferases or whether lysophospholipids must return to the endoplasmic reticulum or to the mitochondrial associated membrane for reesterification. In this review we will focus on the few glycerolipid acyltransferases that are known to be mitochondrial. This article is part of a Special Issue entitled: Lipids of Mitochondria edited by Guenther Daum.

Specific requirements of nonbilayer phospholipids in mitochondrial respiratory chain function and formation.

Mitochondrial membrane phospholipid composition affects mitochondrial function by influencing the assembly of the mitochondrial respiratory chain (MRC) complexes into supercomplexes. For example, the loss of cardiolipin (CL), a signature non-bilayer-forming phospholipid of mitochondria, results in disruption of MRC supercomplexes. However, the functions of the most abundant mitochondrial phospholipids, bilayer-forming phosphatidylcholine (PC) and non-bilayer-forming phosphatidylethanolamine (PE), are not clearly defined. Using yeast mutants of PE and PC biosynthetic pathways, we show a specific requirement for mitochondrial PE in MRC complex III and IV activities but not for their formation, whereas loss of PC does not affect MRC function or formation. Unlike CL, mitochondrial PE or PC is not required for MRC supercomplex formation, emphasizing the specific requirement of CL in supercomplex assembly. Of interest, PE biosynthesized in the endoplasmic reticulum (ER) can functionally substitute for the lack of mitochondrial PE biosynthesis, suggesting the existence of PE transport pathway from ER to mitochondria. To understand the mechanism of PE transport, we disrupted ER-mitochondrial contact sites formed by the ERMES complex and found that, although not essential for PE transport, ERMES facilitates the efficient rescue of mitochondrial PE deficiency. Our work highlights specific roles of non-bilayer-forming phospholipids in MRC function and formation.

The Glycerol-3-Phosphate Acyltransferase TbGAT is Dispensable for Viability and the Synthesis of Glycerolipids in Trypanosoma brucei.

Glycerolipids are the main constituents of biological membranes in Trypanosoma brucei, which causes sleeping sickness in humans. Importantly, they occur as a structural component of the glycosylphosphatidylinositol lipid anchor of the abundant cell surface glycoproteins procyclin in procyclic forms and variant surface glycoprotein in bloodstream form, that play crucial roles for the development of the parasite in the insect vector and the mammalian host, respectively. The present work reports the characterization of the glycerol-3-phosphate acyltransferase TbGAT that initiates the biosynthesis of ester glycerolipids. TbGAT restored glycerol-3-phosphate acyltransferase activity when expressed in a Leishmania major deletion strain lacking this activity and exhibited preference for medium length, unsaturated fatty acyl-CoAs. TbGAT localized to the endoplasmic reticulum membrane with its N-terminal domain facing the cytosol. Despite that a TbGAT null mutant in T.brucei procyclic forms lacked glycerol-3-phosphate acyltransferase activity, it remained viable and exhibited similar growth rate as the wild type. TbGAT was dispensable for the biosynthesis of phosphatidylcholine, phosphatidylinositol, phosphatidylserine, and GPI-anchored protein procyclin. However, the null mutant exhibited a slight decrease in phosphatidylethanolamine biosynthesis that was compensated with a modest increase in production of ether phosphatidylcholine. Our data suggest that an alternative initial acyltransferase takes over TbGAT's function in its absence.

Sp1 and Sp3 Are the Transcription Activators of Human ek1 Promoter in TSA-Treated Human Colon Carcinoma Cells.

BackgroundEthanolamine kinase (EK) catalyzes the phosphorylation of ethanolamine, the first step in the CDP-ethanolamine pathway for the biosynthesis of phosphatidylethanolamine (PE). Human EK exists as EK1, EK2α and EK2β isoforms, encoded by two separate genes, named ek1 and ek2. EK activity is stimulated by carcinogens and oncogenes, suggesting the involvement of EK in carcinogenesis. Currently, little is known about EK transcriptional regulation by endogenous or exogenous signals, and the ek gene promoter has never been studied.Methodology/Principal FindingsIn this report, we mapped the important regulatory regions in the human ek1 promoter. 5’ deletion analysis and site-directed mutagenesis identified a Sp site at position (-40/-31) that was essential for the basal transcription of this gene. Treatment of HCT116 cells with trichostatin A (TSA), a histone deacetylase inhibitor, significantly upregulated the ek1 promoter activity through the Sp(-40/-31) site and increased the endogenous expression of ek1. Chromatin immunoprecipitation assay revealed that TSA increased the binding of Sp1, Sp3 and RNA polymerase II to the ek1 promoter in HCT116 cells. The effect of TSA on ek1 promoter activity was cell-line specific as TSA treatment did not affect ek1 promoter activity in HepG2 cells.Conclusion/SignificanceIn conclusion, we showed that Sp1 and Sp3 are not only essential for the basal transcription of the ek1 gene, their accessibility to the target site on the ek1 promoter is regulated by histone protein modification in a cell line dependent manner.

Transcriptional suppression of CTP:phosphoethanolamine cytidylyltransferase by 25-hydroxycholesterol is mediated by nuclear factor-Y and Yin Yang 1.

Pcyt2 (CTP:phosphoethanolamine cytidylyltransferase) is the rate-limiting enzyme in mammalian PE (phosphatidylethanolamine) biosynthesis. Previously, we reported that Pcyt2 mRNA levels increased in several types of cells after serum starvation, an effect that could be suppressed by supplementation with low-density lipoprotein or 25-HC (25-hydroxycholesterol). Transcription of Hmgcr, which encodes 3-hydroxy-3-methylglutaryl-CoA reductase, is also suppressed by 25-HC in the same dose-dependent manner. Nevertheless, a sterol-regulatory element was not detected in the Pcyt2 promoter region. The important element for transcriptional control of Pcyt2 by 25-HC (1.25 μM) was determined to reside between -56 and -36 on the basis of analysis with several Pcyt2 promoter deletion-luciferase reporters in NIH 3T3 cells. Using the yeast one-hybrid system, we found that NF-Y (nuclear factor-Y) binds at C(-37)CAAT(-41) and YY1 (Yin Yang1) binds at C(-42)AT(-40) in the Pcyt2 promoter. Endogenous NF-Y and YY1 bind clearly and competitively to these sites and are important for basal Pcyt2 transcription. Moreover, NF-Y binds to the Hmgcr promoter at C(-14)CA(-12) in gel-shift analysis, and suppression of the basal luciferase activity of the Hmgcr promoter-reporter construct (-30/+61) by 25-HC was abolished when C(-14)CA(-12) was mutated. Furthermore, transcriptional suppression of Pcyt2 by 25-HC was reduced following knockdown targeting of NF-YA or YY1. ChIP analysis revealed that 25-HC inhibited the interaction between NF-Y and RNA polymerase II on the Pcyt2 and Hmgcr promoters. On the basis of these results, we conclude that NF-Y and YY1 are important for the basal transcription of Pcyt2 and that NF-Y is involved in the inhibitory effects of 25-HC on Pcyt2 transcription.

Antiphosphatidylethanolamine Antibodies: New Pieces to the Puzzle of the Antiphospholipid Syndrome?

Objective: To show the contribution of antiphosphatidylethanolamine antibodies (aPE) screening in Antiphospholipid antibody syndrome (APS) diagnosis, especially in Antiphosphopholipid Syndrome (SNAPS) patients, based on litterature data. Methods: A comprehensive literature review of English-language articles from January 1st 2000 to December 31th 2014 was performed using PudMed. Keywords used for the search were: seronegative antiphospholipid antibody OR phosphatidylethanolamine function, biosynthesis and physiological roles OR antiphosphatidylethanolamine antibody OR seronegative antiphospholipid syndrome. Results: Although aPE was not included in the Sapporo criteria for the classification of APS, many studies described a significant association between aPE and the main clinical events of APS: thrombosis and obstetrical complications. Conclusion: Because of aPL family’s heterogeneity, it appears necessary to look for non-criteria aPL in order to improve the diagnostic yield in APS, especially in patients with SNAPS. One of the most promising of this heterogeneous aPL family is aPE. Although these molecules may allow early detection of APS, their clinical relevance is still debatable and needs standardized aPE ELISA, in addition to well-designed clinical studies to confirm its diagnostic value, such as the association between recursion of thrombotic events and the presence of aPE.

Masking of β(1-3)-glucan in the cell wall of Candida albicans from detection by innate immune cells depends on phosphatidylserine.

The virulence of Candida albicans in a mouse model of invasive candidiasis is dependent on the phospholipids phosphatidylserine (PS) and phosphatidylethanolamine (PE). Disruption of the PS synthase gene CHO1 (i.e., cho1Δ/Δ) eliminates PS and blocks the de novo pathway for PE biosynthesis. In addition, the cho1Δ/Δ mutant's ability to cause invasive disease is severely compromised. The cho1Δ/Δ mutant also exhibits cell wall defects, and in this study, it was determined that loss of PS results in decreased masking of cell wall β(1-3)-glucan from the immune system. In wild-type C. albicans, the outer mannan layer of the wall masks the inner layer of β(1-3)-glucan from exposure and detection by innate immune effector molecules like the C-type signaling lectin Dectin-1, which is found on macrophages, neutrophils, and dendritic cells. The cho1Δ/Δ mutant exhibits increases in exposure of β(1-3)-glucan, which leads to greater binding by Dectin-1 in both yeast and hyphal forms. The unmasking of β(1-3)-glucan also results in increased elicitation of TNF-α from macrophages in a Dectin-1-dependent manner. The role of phospholipids in fungal pathogenesis is an emerging field, and this is the first study showing that loss of PS in C. albicans results in decreased masking of β(1-3)-glucan, which may contribute to our understanding of fungus-host interactions.

Lipid metabolism in mitochondrial membranes.

Mitochondrial membranes have a unique lipid composition necessary for proper shape and function of the organelle. Mitochondrial lipid metabolism involves biosynthesis of the phospholipids phosphatidylethanolamine, cardiolipin and phosphatidylglycerol, the latter is a precursor of the late endosomal lipid bis(monoacylglycero)phosphate. It also includes mitochondrial fatty acid synthesis necessary for the formation of the lipid cofactor lipoic acid. Furthermore the synthesis of coenzyme Q takes place in mitochondria as well as essential parts of the steroid and vitamin D metabolism. Lipid transport and remodelling, which are necessary for tailoring and maintaining specific membrane properties, are just partially unravelled. Mitochondrial lipids are involved in organelle maintenance, fission and fusion, mitophagy and cytochrome c-mediated apoptosis. Mutations in TAZ, SERAC1 and AGK affect mitochondrial phospholipid metabolism and cause Barth syndrome, MEGDEL and Sengers syndrome, respectively. In these disorders an abnormal mitochondrial energy metabolism was found, which seems to be due to disturbed protein-lipid interactions, affecting especially enzymes of the oxidative phosphorylation. Since a growing number of enzymes and transport processes are recognised as parts of the mitochondrial lipid metabolism, a further increase of lipid-related disorders can be expected.

Fatty acid and phospholipid biosynthetic pathways are regulated throughout mammary epithelial cell differentiation and correlate to breast cancer survival

This work combined gene and protein expression, gas chromatography-flame ionization detector, and hydrophilic interaction liquid chromatography-tandem mass spectrometry to compare lipid metabolism changes in undifferentiated/proliferating vs. functionally differentiated mammary epithelial cells (MECs) and to study their correlation to breast cancer survival. Sixty-eight genes involved in lipid metabolism were changed in MEC differentiation. Differentiated cells showed induction of Elovl6 (2-fold), Scd1 (4-fold), and Fads2 (2-fold), which correlated with increased levels of C16:1 n-7 and C18:1 n-9 (1.5-fold), C20:3 n-6 (2.5-fold), and C20:4 n-6 (6-fold) fatty acids (FAs) and more phospholipids (PLs) containing these species. Further, increased expression (2- to 3-fold) of genes in phosphatidylethanolamine (PE) de novo biosynthesis resulted in a 20% PE increase. Proliferating/undifferentiated cells showed higher C16:0 (1.7-fold) and C18:2 n-6 (4.2-fold) levels and more PLs containing C16:0 FAs [PC(16:0/16:1), PG(16:0/18:2), PG(16:0/18:1), and SM(16:0/18:0)]. Kaplan-Meier analysis of data from 3455 patients with breast cancer disclosed a positive correlation for 59% of genes expressed in differentiated MECs with better survival. PE biosynthesis and FA oxidation correlated with better prognosis in patients with breast cancer, including the basal-like subtype. Therefore, genes involved in mammary gland FA and PL metabolism and their resulting molecular species reflect the cellular proliferative ability and differentiation state and deserve further studies as potential markers of breast cancer progression

Activation of peroxisome proliferator activator receptor delta in mouse impacts lipid composition and placental development at early stage of gestation.

Peroxisome proliferator-activated receptor delta (Ppard) activation has been implicated in regulating a multitude of biological processes in placental development. In this study, we employed the UPLC-ESI-TOFMS approach to investigate the metabolic traits in placenta from GW501516-treated mice at Embryonic Day 10.5. By analyzing the mass spectrum data, ions with the most significant differences between control and GW501516-treated groups were identified. Among these metabolites, the fatty acids, phospholipids, and sterol lipids were dramatically increased. Ingenuity Pathway Analysis (IPA) showed that phosphatidylethanolamine biosynthesis and glycolysis were the top two altered metabolic pathways involved in carbohydrate metabolism, energy production, and lipid metabolism. Subsequent immunoblotting experiments provided evidence for positive correlation of PPARD level and AKT and ERK signaling pathways upon GW501516 treatment. Furthermore, the stimulation of GW501516 increased trophoblast cell fusion gene syncytin-A (Syna), but not syncytin-B (Synb), expression, suggesting a potential role of Ppard activation in promoting cytotrophoblast differentiation. Our results reveal that Ppard activation elicits dramatic changes of the metabolic activities in placenta, which is correlated to AKT and ERK signaling.

Discovery of a bifunctional cardiolipin/phosphatidylethanolamine synthase in bacteria

Phosphatidylethanolamine (PE) and cardiolipin (CL) are major components of bacterial and eukaryotic membranes. In bacteria, synthesis of PE usually occurs via decarboxylation of phosphatidylserine (PS) by PS decarboxylases (Psd). CL is produced by various CL synthases (Cls). Membranes of the plant pathogen Xanthomonas campestris predominantly contain PE, phosphatidylglycerol (PG) and CL. The X. campestris genome encodes one Psd and six putative CLs. Deletion of psd resulted in loss of PE and accumulation of PS. The mutant was severely affected in growth and cell size. PE synthesis, growth and cell division were partially restored when cells were supplied with ethanolamine (EA) suggesting a previously unknown PE synthase activity. Via mutagenesis, we identified a Cls enzyme (Xc_0186) responsible for EA-dependent PE biosynthesis. Xanthomonas lacking xc_0186 not only lost its ability to utilize EA for PE synthesis but also produced less CL suggesting a bifunctional enzyme. Recombinant Xc_0186 in E. coli and in cell-free extracts uses cytidine diphosphate diacylglycerol (CDP-DAG) and PG for CL synthesis. It is also able to use CDP-DAG and EA for PE synthesis. Owing to its dual function in CL and PE production, we consider Xc_0186 the founding member of a new class of enzymes called CL/PE synthase (CL/PEs).

A novel murine CTP:phosphoethanolamine cytidylyltransferase splice variant is a post-translational repressor and an indicator that both cytidylyltransferase domains are required for activity

CTP:phosphoethanolamine cytidylyltransferase (Pcyt2) has an important regulatory function in biosynthesis of the membrane phospholipid phosphatidylethanolamine. We previously determined that the full-length Pcyt2α and its splice variant Pcyt2β are the main active isoforms of this enzyme. Here we report that mouse Pcyt2 could be spliced at Introns 7 and 8 to produce a unique third isoform, Pcyt2γ, in which the second cytidylyltransferase domain at the C-terminus becomes deleted. Pcyt2γ is ubiquitously expressed in embryonic and adult mouse tissues, and is the most abundant in the kidney, skeletal muscle and testis. Pcyt2γ splicing mechanism dominates over Pcyt2β exon-skipping mechanism in most examined tissues. Although Pcyt2γ maintains the N-terminal cytidylyltransferase domain as most cytidylyltransferases, the lack of the C-terminal cytidylyltransferase domain causes a complete loss of catalytic activity. However, Pcyt2γ interacts with the active isoform, Pcyt2α, and significantly reduces Pcyt2α homodimerization and activity. The inactive N-domain (H35Y, H35A) and C-domain (H244Y, H244A) mutants of Pcyt2α also reduce Pcyt2α homodimerization and activity. This study revealed the importance of both cytidylyltransferase (35)HYGH and (244)HIGH motifs for the activity of murine Pcyt2α and established that the naturally occurring splice variant Pcyt2γ has a function to restrain the enzyme activity through the formation of unproductive enzyme complexes.

Isoform-specific and Protein Kinase C-mediated Regulation of CTP:Phosphoethanolamine Cytidylyltransferase Phosphorylation

CTP:phosphoethanolamine cytidylyltransferase (Pcyt2) is the main regulatory enzyme for de novo biosynthesis of phosphatidylethanolamine by the CDP-ethanolamine pathway. There are two isoforms of Pcyt2, -α and -β; however, very little is known about their specific roles in this important metabolic pathway. We previously demonstrated increased phosphatidylethanolamine biosynthesis subsequent to elevated activity and phosphorylation of Pcyt2α and -β in MCF-7 breast cancer cells grown under conditions of serum deficiency. Mass spectroscopy analyses of Pcyt2 provided evidence for isoform-specific as well as shared phosphorylations. Pcyt2β was specifically phosphorylated at the end of the first cytidylyltransferase domain. Pcyt2α was phosphorylated within the α-specific motif that is spliced out in Pcyt2β and on two PKC consensus serine residues, Ser-215 and Ser-223. Single and double mutations of PKC consensus sites reduced Pcyt2α phosphorylation, activity, and phosphatidylethanolamine synthesis by 50-90%. The phosphorylation and activity of endogenous Pcyt2 were dramatically increased with phorbol esters and reduced by specific PKC inhibitors. In vitro translated Pcyt2α was phosphorylated by PKCα, PKCβI, and PKCβII. Pcyt2α Ser-215 was also directly phosphorylated with PKCα. Mapping of the Pcyt2α- and -β-phosphorylated sites to the solved structure of a human Pcyt2β showed that they clustered within and flanking the central linker region that connects the two catalytic domains and is a novel regulatory segment not present in other cytidylyltransferases. This study is the first to demonstrate differences in phosphorylation between Pcyt2 isoforms and to uncover the role of the PKC-regulated phosphorylation.

Meclizine Inhibits Mitochondrial Respiration through Direct Targeting of Cytosolic Phosphoethanolamine Metabolism

We recently identified meclizine, an over-the-counter drug, as an inhibitor of mitochondrial respiration. Curiously, meclizine blunted respiration in intact cells but not in isolated mitochondria, suggesting an unorthodox mechanism. Using a metabolic profiling approach, we now show that treatment with meclizine leads to a sharp elevation of cellular phosphoethanolamine, an intermediate in the ethanolamine branch of the Kennedy pathway of phosphatidylethanolamine biosynthesis. Metabolic labeling and in vitro enzyme assays confirmed direct inhibition of the cytosolic enzyme CTP:phosphoethanolamine cytidylyltransferase (PCYT2). Inhibition of PCYT2 by meclizine led to rapid accumulation of its substrate, phosphoethanolamine, which is itself an inhibitor of mitochondrial respiration. Our work identifies the first pharmacologic inhibitor of the Kennedy pathway, demonstrates that its biosynthetic intermediate is an endogenous inhibitor of respiration, and provides key mechanistic insights that may facilitate repurposing meclizine for disorders of energy metabolism.

Phosphatidylethanolamine from Phosphatidylserine Decarboxylase2 is Essential for Autophagy Under Cadmium Stress in Saccharomyces cerevisiae

Cadmium (Cd) is a potent toxic element used in several industries and in the process contaminates air, soil, and water. Exposure of Saccharomyces cerevisiae to Cd increases the major phospholipids, and profound increase was observed in phosphatidylethanolamine (PE). In yeast, there are four different pathways contributing to the biosynthesis of PE, and contribution to PE pool through phosphatidylserine decarboxylase2 (psd2) is not significant in normal conditions. Upon Cd exposure, psd2Δ strain showed a significant decrease in major phospholipids including PE. When exposed to Cd, wild-type (WT) cells depicted an increase in ER stress and autophagy, whereas in psd2, ER stress was noted but autophagy process was impaired. The supplementation of ethanolamine did not overcome the Cd stress and also the autophagy process, whereas overexpression of PSD2 in psd2Δ increased the cellular tolerance, PE levels, and the autophagy process against Cd stress. From our studies, we can suggest that PSD2 of S. cerevisiae has an important role in PE synthesis and in autophagy process under Cd stress.

Regulation of Phosphatidylethanolamine Homeostasis — The Critical Role of CTP:Phosphoethanolamine Cytidylyltransferase (Pcyt2)

Phosphatidylethanolamine (PE) is the most abundant lipid on the protoplasmatic leaflet of cellular membranes. It has a pivotal role in cellular processes such as membrane fusion, cell cycle regulation, autophagy, and apoptosis. CTP:phosphoethanolamine cytidylyltransferase (Pcyt2) is the main regulatory enzyme in de novo biosynthesis of PE from ethanolamine and diacylglycerol by the CDP-ethanolamine Kennedy pathway. The following is a summary of the current state of knowledge on Pcyt2 and how splicing and isoform specific differences could lead to variations in functional properties in this family of enzymes. Results from the most recent studies on Pcyt2 transcriptional regulation, promoter function, autophagy, and cell growth regulation are highlighted. Recent data obtained from Pcyt2 knockout mouse models is also presented, demonstrating the essentiality of this gene in embryonic development as well as the major physiological consequences of deletion of one Pcyt2 allele. Those include development of symptoms of the metabolic syndrome such as elevated lipogenesis and lipoprotein secretion, hypertriglyceridemia, liver steatosis, obesity, and insulin resistance. The objective of this review is to elucidate the nature of Pcyt2 regulation by linking its catalytic function with the regulation of lipid and energy homeostasis.