40 research outputs found
Neuronal Cholesterol Accumulation Induced by Cyp46a1 Down-Regulation in Mouse Hippocampus Disrupts Brain Lipid Homeostasis
Impairment in cholesterol metabolism is associated with many neurodegenerative disorders including Alzheimer's disease (AD). However, the lipid alterations underlying neurodegeneration and the connection between altered cholesterol levels and AD remains not fully understood. We recently showed that cholesterol accumulation in hippocampal neurons, induced by silencing Cyp46a1 gene expression, leads to neurodegeneration with a progressive neuronal loss associated with AD-like phenotype in wild-type mice. We used a targeted and non-targeted lipidomics approach by liquid chromatography coupled to high-resolution mass spectrometry to further characterize lipid modifications associated to neurodegeneration and cholesterol accumulation induced by CYP46A1 inhibition. Hippocampus lipidome of normal mice was profiled 4 weeks after cholesterol accumulation due to Cyp46a1 gene expression down-regulation at the onset of neurodegeneration. We showed that major membrane lipids, sphingolipids and specific enzymes involved in phosphatidylcholine and sphingolipid metabolism, were rapidly increased in the hippocampus of AAV-shCYP46A1 injected mice. This lipid accumulation was associated with alterations in the lysosomal cargoe, accumulation of phagolysosomes and impairment of endosome-lysosome trafficking. Altogether, we demonstrated that inhibition of cholesterol 24-hydroxylase, key enzyme of cholesterol metabolism leads to a complex dysregulation of lipid homeostasis. Our results contribute to dissect the potential role of lipids in severe neurodegenerative diseases like AD
Shotgun lipidomics and mass spectrometry imaging unveil diversity and dynamics in Gammarus fossarum lipid composition
Sentinel species are playing an indispensable role in monitoring environmental pollution in aquatic ecosystems. Many pollutants found in water prove to be endocrine disrupting chemicals that could cause disruptions in lipid homeostasis in aquatic species. A comprehensive profiling of the lipidome of these species is thus an essential step toward understanding the mechanism of toxicity induced by pollutants. Both the composition and spatial distribution of lipids in freshwater crustacean Gammarus fossarum were extensively examined herein. The baseline lipidome of gammarids of different sex and reproductive stages was established by high throughput shotgun lipidomics. Spatial lipid mapping by high resolution mass spectrometry imaging led to the discovery of sulfate-based lipids in hepato-pancreas and their accumulation in mature oocytes. A diverse and dynamic lipid composition in G. fossarum was uncovered, which deepens our understanding of the biochemical changes during development and which could serve as a reference for future ecotoxicological studies.Approches Protéomique et Lipidomique pour la compréhension des mécanismes moléculaires de toxicité en lien avec l'altération du métabolisme lipidique chez l'espÚce sentinelle Gammarus fossarum durant le cycle de reproductio
Approches multi-omiques pour la comprĂ©hension des mĂ©canismes de toxicitĂ© en lien avec lâaltĂ©ration du mĂ©tabolisme lipidique chez lâespĂšce sentinelle Gammarus fossarum
National audienc
Traitement des données brutes de la spectrométrie de masse - infusion directe. Application à la lipidomique
Institut de Chimie CNRS action convergenceNational audienc
Characterization of lysolipid acyltransferases in S. cerevisiae - Contribution of Mass Spectrometry
En plus de leurs propriĂ©tĂ©s structurales comme constituants majeurs des membranes biologiques des cellules, les lipides jouent de nombreux rĂŽles dans la signalisation cellulaire, le stockage dâĂ©nergie et le transport de protĂ©ines. Leurs importances biologiques ont menĂ© Ă une augmentation accrue des mĂ©thodes analytiques pour la caractĂ©risation dâespĂšces molĂ©culaires uniques. De rĂ©cents progrĂšs en spectromĂ©trie de masse ont amenĂ© Ă la caractĂ©risation et Ă la quantification des espĂšces molĂ©culaires des lipides dans des extraits lipidiques bruts (Han and Gross, 2005; Murphy et al., 2001). Par exemple, les espĂšces molĂ©culaires de phospholipides peuvent ĂȘtre identifiĂ©es spĂ©cifiquement par leur tĂȘte polaire, la nature de leurs chaĂźnes dâacide gras et leur positionnement au niveau du squelette glycĂ©rol.In addition to their structural properties as main constituents of biological membranes, lipids play a multitude of roles such as in cell signalling, energy storage, and protein transport. Their biological importance has led to an increasing focus on analytical methods for the characterisation of their individual molecular species. Improvements in mass spectrometric technology has provided a great advantage for the characterisation and quantification of molecular lipid species in total lipid extracts (Han and Gross, 2005; Murphy et al., 2001). For instance, phospholipid molecular species can be identified on the basis of a characteristic fragment of the lipid class, the nature of the acyl chains and their positions on the glycerol backbone.A method allowing the quantitative profiling of the yeast lipidome was developed in a recent study using automated shotgun infusion strategy (Ejsing et al., 2009). We applied this method to characterise several lysophospholipid acyltransferase yeast mutants produced using reverse-genetics. These enzymes are involved in essential biological processes like de novo synthesis or remodelling of the phospholipid membrane component (Testet et al., 2005; Le Guedard et al., 2009). The comparative analysis of phospholipid molecular species from the wild-type strain and the corresponding deletion mutants has allowed us to identify lipid compositional changes, and has given us significant indications about the in vivo function of the encoded lysophospholipid acyltransferases
Caractérisation de lysolipide acyltransférases chez S. cerevisiae. Apport de la Spectrométrie de Masse.
Cell membranes, enveloping either cells or organelles, are fundamental structural elements in all kingdoms of life. Biological membranes are made of proteins inserted in a lipidic bilayer composed of phospholipids, sterols and sphingolipids. In addition to their role as a hydrophobic and semi-permeable barrier, they are involved in many physiological processes such as cell signalling, energy storage and membrane transport. Indeed, cell membranes allow a tight regulation of energy, information, nutrients and metabolites flows. The importance of membrane structures is such that the cell is obliged to ensure quantitatively and qualitatively their integrity, diversity and specificity. However, the cellular mechanisms that control this membrane homeostasis remain poorly understood. To better understand this phenomenon, it is important to focus on the modalities of membrane lipid supply and the phenomena involved in their remodeling. It is in this context that the Membrane Biogenesis laboratory has been interested in the study and characterization of enzymes involved in lipid metabolism, namely lysophospholipid acyltransferases. These enzymes are able to acylate a lysophospholipid (a phospholipid with a single fatty acid chain generally positioned in the sn-1 position of the glycerol skeleton), and thus allow the synthesis of a phospholipid sensu stricto (i.e. with two fatty acid chains). These enzymes could play an important role in the construction of membranes, the transfer of lipids between two subcellular compartments and the "remodeling" of membrane lipids. Because of the central role of lipids in physiological and pathological processes, their study has required the development of specific and sensitive analytical methods for the characterization of single molecular species. For more than two decades, advances in mass spectrometry have enabled the characterization and absolute quantification of lipid molecular species from crude lipid extracts from complex biological matrices (Han & Gross, 2005; Wenk, 2005). Indeed, the classes of phospholipids can be specifically identified by their polar head and the molecular species by the nature and positioning of the fatty acid chains at the glycerol backbone. Mass spectrometry is therefore the technique of choice to describe this molecular diversity in detail.Les membranes cellulaires, enveloppant soit les cellules soit les organelles, sont des Ă©lĂ©ments structurels fondamentaux dans tous les rĂšgnes de la vie. Les membranes biologiques sont constitueÌes de proteÌines inseÌreÌes dans une bicouche lipidique composĂ©e de phospholipides, de steÌrols et de sphingolipides. En plus de leur rĂŽle de barriĂšre hydrophobe et semi-permeÌable, elles sont impliqueÌes dans de treÌs nombreux processus physiologiques comme la signalisation cellulaire, le stockage dâeÌnergie et le transport membranaire. En effet, les membranes cellulaires permettent une rĂ©gulation Ă©troite des flux d'Ă©nergie, d'informations, de nutriments et de mĂ©tabolites. Lâimportance des structures membranaires est telle que la cellule est contrainte dâassurer quantitativement et qualitativement leur inteÌgriteÌ, leur diversiteÌ et leur speÌcificiteÌ. Cependant, les meÌcanismes cellulaires assurant le controÌle de cette homeÌostasie membranaire restent encore peu connus. Pour mieux comprendre ce pheÌnomeÌne, il est important de sâinteÌresser aux modaliteÌs dâapprovisionnement des membranes en lipides et aux pheÌnomeÌnes impliqueÌs dans leurs remodelages. Câest dans ce contexte que le laboratoire de BiogeneÌse Membranaire sâest inteÌresseÌ aÌ lâeÌtude et la caracteÌrisation dâenzymes impliqueÌes dans le meÌtabolisme des lipides, Ă savoir les lysophospholipides acyltransfeÌrases. Ces enzymes sont capables dâacyler un lysophospholipide (phospholipide comportant une seule chaiÌne dâacide gras positionneÌe geÌneÌralement en position sn-1 du squelette glyceÌrol), et de permettre ainsi la syntheÌse dâun phospholipide sensu stricto (câest-Ă -dire avec deux chaĂźnes dâacides gras). Ces enzymes pourraient jouer un roÌle important dans lâeÌdification des membranes, le transfert de lipides entre deux compartiments subcellulaires et le « remodelage » des lipides membranaires. En raison du roÌle central des lipides dans les processus physiologiques et pathologiques, leur eÌtude a nĂ©cessitĂ© le deÌveloppement de meÌthodes analytiques spĂ©cifiques et sensibles pour la caracteÌrisation dâespeÌces moleÌculaires uniques. Depuis plus d'une vingtaine dâannĂ©es, les progreÌs en spectromeÌtrie de masse ont permis la caracteÌrisation et la quantification absolue des espeÌces moleÌculaires des lipides aÌ partir d'extraits lipidiques bruts issus de matrices biologiques complexes (Han & Gross, 2005; Wenk, 2005). En effet, les classes de phospholipides peuvent eÌtre identifieÌes speÌcifiquement par leur teÌte polaire et les espeÌces moleÌculaires par la nature et le positionnement des chaiÌnes d'acides gras au niveau du squelette glyceÌrol. La spectromeÌtrie de masse se reÌveÌle donc eÌtre la technique de choix pour dĂ©crire finement cette diversiteÌ moleÌculaire
Characterization of lysolipid acyltransferases in S. cerevisiae - Contribution of Mass Spectrometry
En plus de leurs propriĂ©tĂ©s structurales comme constituants majeurs des membranes biologiques des cellules, les lipides jouent de nombreux rĂŽles dans la signalisation cellulaire, le stockage dâĂ©nergie et le transport de protĂ©ines. Leurs importances biologiques ont menĂ© Ă une augmentation accrue des mĂ©thodes analytiques pour la caractĂ©risation dâespĂšces molĂ©culaires uniques. De rĂ©cents progrĂšs en spectromĂ©trie de masse ont amenĂ© Ă la caractĂ©risation et Ă la quantification des espĂšces molĂ©culaires des lipides dans des extraits lipidiques bruts (Han and Gross, 2005; Murphy et al., 2001). Par exemple, les espĂšces molĂ©culaires de phospholipides peuvent ĂȘtre identifiĂ©es spĂ©cifiquement par leur tĂȘte polaire, la nature de leurs chaĂźnes dâacide gras et leur positionnement au niveau du squelette glycĂ©rol.In addition to their structural properties as main constituents of biological membranes, lipids play a multitude of roles such as in cell signalling, energy storage, and protein transport. Their biological importance has led to an increasing focus on analytical methods for the characterisation of their individual molecular species. Improvements in mass spectrometric technology has provided a great advantage for the characterisation and quantification of molecular lipid species in total lipid extracts (Han and Gross, 2005; Murphy et al., 2001). For instance, phospholipid molecular species can be identified on the basis of a characteristic fragment of the lipid class, the nature of the acyl chains and their positions on the glycerol backbone.A method allowing the quantitative profiling of the yeast lipidome was developed in a recent study using automated shotgun infusion strategy (Ejsing et al., 2009). We applied this method to characterise several lysophospholipid acyltransferase yeast mutants produced using reverse-genetics. These enzymes are involved in essential biological processes like de novo synthesis or remodelling of the phospholipid membrane component (Testet et al., 2005; Le Guedard et al., 2009). The comparative analysis of phospholipid molecular species from the wild-type strain and the corresponding deletion mutants has allowed us to identify lipid compositional changes, and has given us significant indications about the in vivo function of the encoded lysophospholipid acyltransferases
Lipidomics and metabolomics multiplexed assay based on the innovative scout triggered MRM (stMRM) targeted MS mode
International audienc
Caractérisation de lysolipide acyltransférases chez S. cerevisiae - Apport de la Spectrométrie de Masse
En plus de leurs propriĂ©tĂ©s structurales comme constituants majeurs des membranes biologiques des cellules, les lipides jouent de nombreux rĂŽles dans la signalisation cellulaire, le stockage d Ă©nergie et le transport de protĂ©ines. Leurs importances biologiques ont menĂ© Ă une augmentation accrue des mĂ©thodes analytiques pour la caractĂ©risation d espĂšces molĂ©culaires uniques. De rĂ©cents progrĂšs en spectromĂ©trie de masse ont amenĂ© Ă la caractĂ©risation et Ă la quantification des espĂšces molĂ©culaires des lipides dans des extraits lipidiques bruts (Han and Gross, 2005; Murphy et al., 2001). Par exemple, les espĂšces molĂ©culaires de phospholipides peuvent ĂȘtre identifiĂ©es spĂ©cifiquement par leur tĂȘte polaire, la nature de leurs chaĂźnes d acide gras et leur positionnement au niveau du squelette glycĂ©rol.In addition to their structural properties as main constituents of biological membranes, lipids play a multitude of roles such as in cell signalling, energy storage, and protein transport. Their biological importance has led to an increasing focus on analytical methods for the characterisation of their individual molecular species. Improvements in mass spectrometric technology has provided a great advantage for the characterisation and quantification of molecular lipid species in total lipid extracts (Han and Gross, 2005; Murphy et al., 2001). For instance, phospholipid molecular species can be identified on the basis of a characteristic fragment of the lipid class, the nature of the acyl chains and their positions on the glycerol backbone.A method allowing the quantitative profiling of the yeast lipidome was developed in a recent study using automated shotgun infusion strategy (Ejsing et al., 2009). We applied this method to characterise several lysophospholipid acyltransferase yeast mutants produced using reverse-genetics. These enzymes are involved in essential biological processes like de novo synthesis or remodelling of the phospholipid membrane component (Testet et al., 2005; Le Guedard et al., 2009). The comparative analysis of phospholipid molecular species from the wild-type strain and the corresponding deletion mutants has allowed us to identify lipid compositional changes, and has given us significant indications about the in vivo function of the encoded lysophospholipid acyltransferases.BORDEAUX2-Bib. Ă©lectronique (335229905) / SudocSudocFranceF