Project

Back to overview

Lipid Storage, Binding, and Export

English title Lipid Storage, Binding, and Export
Applicant Schneiter Roger
Number 173003
Funding scheme Project funding (Div. I-III)
Research institution Division de Biochimie Département de Biologie Université de Fribourg
Institution of higher education University of Fribourg - FR
Main discipline Biochemistry
Start/End 01.12.2017 - 30.11.2021
Approved amount 940'150.00
Show all

All Disciplines (2)

Discipline
Biochemistry
Cellular Biology, Cytology

Keywords (8)

lipid droplets; Saccharomyces cerevisiae; topology of lipid synthesis; endoplasmic reticulum; CAP proteins; lipid homeostasis; sterol-binding proteins; cholesterol

Lay Summary (German)

Lead
Dieses Forschungsprogramm behandelt zwei getrennte Themen: die Biogenese von Lipidtröpfchen und deren Assoziation mit der Membran des endoplasmatischen Retikulums sowie die Struktur und Funktion einer wichtigen und weitverbreiteten Protein-Superfamilie, die als CAP-Proteine bekannt ist.
Lay summary

Lipidtröpfchen sind in allen eukaryotischen Zellen vorhanden und sind als kugelige intrazelluläre Strukturen erkennbar. Sie dienen dazu, metabolische Energie in Form von neutralen Lipiden, allgemein bekannt als "Fett" zu speichern. Lipidtröpfchen sind also in vielen der pandemischen Erkrankungen wie Fettleibigkeit, Insulinresistenz, Atherosklerose und Lipotoxizität verwickelt. Wir und andere haben zuvor gezeigt, dass diese Lipidtröpfchen eng mit der Membran des endoplasmatischen Retikulums assoziiert sind. Die genaue Art dieser Assoziation zwischen den beiden Kompartimenten ist jedoch nur schlecht definiert. Ziel dieses Teils der Forschungsarbeiten ist es, die Art der Assoziation zwischen dem endoplasmatischen Retikulum und den Lipidtröpfchen genauer zu verstehen.

Die CAP- Superfamilie von Proteinen ist in viele grundlegende biologische Prozesse verwickelt und beinhaltet die Immunabwehr in Säugetieren und Pflanzen, Spermienreifung und -befruchtung, Pathogen-Virulenz, Gift-Toxizität und sogar Prostata- und Hirntumoren. CAP-Familienmitglieder sind meist sekretierte Glykoproteine, die im extrazellulären Raum agieren und sehr stabil sind. Die Wirkungsweise dieser Proteine ist jedoch noch unklar. Wir konnten zuvor zeigen, dass die CAP-Familienmitglieder in Hefe, bekannt als Pry Proteine, Sterole und verwandte hydrophoben Verbindungen binden und solubilisieren können. Neuere Resultate deuten darauf hin, dass diese Proteine auch Fettsäuren binden können. Wir werden prüfen, ob Pry-Proteine Fettsäuren binden und werden die entsprechende Bindungsstelle durch gezielte Mutagenese definieren. Diese Ergebnisse werden somit dazu beitragen, die molekulare Wirkungsweise dieser Proteine und deren physiologische Funktionen genauer zu verstehen.

Direct link to Lay Summary Last update: 18.04.2017

Lay Summary (English)

Lead
This research program addresses two separate topics: the biogenesis of lipid droplets and their association with the membrane of the endoplasmic reticulum, and the structure and function of an important and widespread protein superfamily, known as CAP proteins.
Lay summary

Lipid droplets are present in all eukaryotic cells and are discernible as round intracellular structures. They serve to store metabolic energy in form of neutral lipids, commonly known as “fat”. Lipid droplets are thus implicated in many of the pandemic diseases such as obesity, insulin resistance, atherosclerosis and lipotoxicity. We and others have previously shown that these lipid droplets are closely associated with the membrane of the endoplasmic reticulum. The precise nature of this association between the two compartments, however, has remained elusive. The aim of this part of the proposal thus is to define the nature of the association between the endoplasmic reticulum and lipid droplets.

The CAP superfamily of proteins are implicated in many fundamental biological processes, ranging from immune defense in mammals and plants, sperm maturation and fertilization, pathogen virulence, venom toxicity and even prostate and brain cancer. CAP family members are mostly secreted glycoproteins that are stable in the extracellular space. The mode of action of these proteins, however, has remained elusive. We could previously show that the CAP family members in yeast, known as Pathogen Related in Yeast (Pry), bind and thereby solubilize sterols and related hydrophobic compounds. Recent evidence indicates that these proteins not only are capable of binding sterols, but they independently also can bind fatty acids. We will test whether Pry proteins bind fatty acids and define the binding site by site-directed mutagenesis. These results will thus help to define the molecular mode of action of these proteins and thereby improve our understanding of the physiological function these proteins exert in both health and disease.

Direct link to Lay Summary Last update: 18.04.2017

Responsible applicant and co-applicants

Employees

Publications

Publication
Lipid droplet biogenesis from specialized ER subdomain.
Choudhary V, Schneiter R (2020), Lipid droplet biogenesis from specialized ER subdomain., in Microbial Cell, 7, 218-221.
Lipid droplet biogenesis is driven by liquid-liquid phase separation..
Zoni V, Khaddaj R, Campomanes P, Thiam AR, Schneiter R, Vanni S (2020), Lipid droplet biogenesis is driven by liquid-liquid phase separation.., in Submitted, submitted.
Mitochondrial sphingosine-1-phosphate lyase is essential for phosphatidylethanolamine synthesis and survival of Trypanosoma brucei.
Dawoody Nejad L, Stumpe M, Rauch M, Hemphill A, Schneiter R, Bütikofer P, Serricchio M (2020), Mitochondrial sphingosine-1-phosphate lyase is essential for phosphatidylethanolamine synthesis and survival of Trypanosoma brucei., in Sci Rep, 10(1), 8268-8268.
Necator americanus Ancylostoma secreted protein-2 (Na-ASP-2) selectively binds an ascaroside (ascr#3)
El Atab O, Darwiche R, Truax NJ, Schneiter R, Hull KG, Romo D, Asojo OA (2020), Necator americanus Ancylostoma secreted protein-2 (Na-ASP-2) selectively binds an ascaroside (ascr#3), in Int. J. Parasit, submitted.
Seipin and Nem1 establish discrete ER subdomains to initiate yeast lipid droplet biogenesis.
Choudhary V, El Atab O, Mizzon G, Prinz WA, Schneiter R (2020), Seipin and Nem1 establish discrete ER subdomains to initiate yeast lipid droplet biogenesis., in J. Cell. Biol., 219(7), e201910177.
Targeting of integral membrane proteins to the surface of lipid droplets.
Khaddaj R, Mari M, Cottier S, Reggiori F, Schneiter R (2020), Targeting of integral membrane proteins to the surface of lipid droplets., in Submitted, Submitted, submitted.
The yeast cell wall protein Pry3 inhibits mating through highly conserved residues within the CAP domain.
Cottier S, Darwiche R, Meyenhofer F, Debelyy MO, Schneiter R (2020), The yeast cell wall protein Pry3 inhibits mating through highly conserved residues within the CAP domain., in Biol Open, 9, bio053470.
Architecture of Lipid Droplets in Endoplasmic Reticulum Is Determined by Phospholipid Intrinsic Curvature.
Choudhary V, Golani G, Joshi AS, Cottier S, Schneiter R, Prinz WA, Kozlov MM (2018), Architecture of Lipid Droplets in Endoplasmic Reticulum Is Determined by Phospholipid Intrinsic Curvature., in Current Biology, 28, 915-926e9.
Crystal structure of Brugia malayi venom allergen-like protein-1 (BmVAL-1), a vaccine candidate for lymphatic filariasis.
Darwiche R, Lugo F, Drurey C, Varossieau K, Smant G, Wilbers RHP, Maizels RM, Schneiter R, Asojo OA (2018), Crystal structure of Brugia malayi venom allergen-like protein-1 (BmVAL-1), a vaccine candidate for lymphatic filariasis., in Int. J. Parasit., 48(5), 371-378.
Heligmosomoides polygyrus Venom Allergen-like Protein-4 (HpVAL-4) is a sterol binding protein.
Asojo OA, Darwiche R, Gebremedhin S, Smant G, Lozano-Torres JL, Drurey C, Pollet J, Maizels RM, Schneiter R, Wilbers RHP (2018), Heligmosomoides polygyrus Venom Allergen-like Protein-4 (HpVAL-4) is a sterol binding protein., in Int. J. Parasit., 48(5), 359-369.
Localization and functional characterization of the pathogenesis-related proteins Rbe1p and Rbt4p in Candida albicans.
Bantel Y, Darwiche R, Rupp S, Schneiter R, Sohn K (2018), Localization and functional characterization of the pathogenesis-related proteins Rbe1p and Rbt4p in Candida albicans., in PLoS One, 13(8), 0201932-0201932.
Secreted venom allergen-like proteins of helminths: Conserved modulators of host responses in animals and plants.
Wilbers RHP, Schneiter R, Holterman MHM, Drurey C, Smant G, Asojo OA, Maizels RM, Lozano-Torres JL (2018), Secreted venom allergen-like proteins of helminths: Conserved modulators of host responses in animals and plants., in PLoS Pathog, 14(10), 1007300-1007300.
A Ligand-Binding Assay to Measure the Affinity and Specificity of Sterol-Binding Proteins In Vitro.
Darwiche R, Schneiter R (2017), A Ligand-Binding Assay to Measure the Affinity and Specificity of Sterol-Binding Proteins In Vitro., in Methods Mol Biol, 1645, 361-368.
Chemical crosslinking and mass spectrometry to elucidate the topology of integral membrane proteins.
Debelyy MO, Waridel P, Quadroni M, Schneiter R, Conzelmann A (2017), Chemical crosslinking and mass spectrometry to elucidate the topology of integral membrane proteins., in PLoS One, 12(10), 0186840-0186840.
Crystal Structure of Borrelia turicatae protein, BTA121, a differentially regulated  gene in the tick-mammalian transmission cycle of relapsing fever spirochetes.
Luo Z, Kelleher AJ, Darwiche R, Hudspeth EM, Shittu OK, Krishnavajhala A, Schneiter R, Lopez JE, Asojo OA (2017), Crystal Structure of Borrelia turicatae protein, BTA121, a differentially regulated  gene in the tick-mammalian transmission cycle of relapsing fever spirochetes., in Sci Rep, 7(1), 15310-15310.
Crystal Structure of MpPR-1i, a SCP/TAPS protein from Moniliophthora perniciosa, the fungus that causes Witches' Broom Disease of Cacao.
Baroni RM, Luo Z, Darwiche R, Hudspeth EM, Schneiter R, Pereira GAG, Mondego JMC, Asojo OA (2017), Crystal Structure of MpPR-1i, a SCP/TAPS protein from Moniliophthora perniciosa, the fungus that causes Witches' Broom Disease of Cacao., in Sci Rep, 7(1), 7818-7818.
Plant pathogenesis-related proteins of the cacao fungal pathogen Moniliophthora perniciosa differ in their lipid-binding specificities.
Darwiche R, El Atab O, Baroni RM, Teixeira PJPL, Mondego JMC, Pereira GAG, Schneiter R (2017), Plant pathogenesis-related proteins of the cacao fungal pathogen Moniliophthora perniciosa differ in their lipid-binding specificities., in J. Biol. Chem., 292(50), 20558-20569.
Sphingolipid accumulation causes mitochondrial dysregulation and cell death.
Knupp J, Martinez-Montañés F, Van Den Bergh F, Cottier S, Schneiter R, Beard D, Chang A (2017), Sphingolipid accumulation causes mitochondrial dysregulation and cell death., in Cell Death Differ, 24, 2044-2053.
The function of yeast CAP family proteins in lipid export, mating, and pathogen defense.
Darwiche R, El Atab O, Cottier S, Schneiter R (2017), The function of yeast CAP family proteins in lipid export, mating, and pathogen defense., in FEBS Lett, 592(8), 1304-1311.
The pathogen-related yeast protein Pry1, a member of the CAP protein superfamily, is a fatty acid-binding protein.
Darwiche R, Mène-Saffrané L, Gfeller D, Asojo OA, Schneiter R (2017), The pathogen-related yeast protein Pry1, a member of the CAP protein superfamily, is a fatty acid-binding protein., in J. Biol. Chem., 292(20), 8304-8314.
The sterol-binding activity of PATHOGENESIS-RELATED PROTEIN 1 reveals the mode of action of an antimicrobial protein.
Gamir J, Darwiche R, Van't Hof P, Choudhary V, Stumpe M, Schneiter R, Mauch F (2017), The sterol-binding activity of PATHOGENESIS-RELATED PROTEIN 1 reveals the mode of action of an antimicrobial protein., in Plant J, 89(3), 502-509.

Collaboration

Group / person Country
Types of collaboration
Choudhary/AIIMS India (Asia)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Exchange of personnel
Reggiori/UMCG Netherlands (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Bütikofer/IBMM Bern Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Exchange of personnel
Reinhardt/UniFr Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Asojo/Hampton University United States of America (North America)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Vanni/UniFr Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Van Boghaert/Ghent Belgium (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Exchange of personnel
Wilbers/Wageningen Netherlands (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
EMBO Workshop on Lipid function in health and disease Poster Lipid droplets are formed at pre-defined discrete sites in the ER 27.09.2019 Dresden, Germany Schneiter Roger;
Centre for Integrative Genomics (CIG) symposium, Metabolism: molecules, diseases, and personalized health Talk given at a conference Lipid droplets are formed at pre-defined discrete sites in the ER 13.06.2019 Lausanne, Switzerland Schneiter Roger; Khaddaj Rasha;
Summer school: Molecular Organization, Function and Dynamics of Biomembranes, at the Institut d'Etudes Scientifiques in Cargèse, France. 10-20 June 2019 Talk given at a conference The function of CAP proteins in sterol binding and transport is affected by their interaction with PSP94 and A1BG 10.06.2019 Cargèse, France El Atab Ola;
Levures, Modèles et Outil Talk given at a conference Tracking lipid droplet fate during yeast mating 11.09.2018 Rheinau, Switzerland El Atab Ola; Khaddaj Rasha;
45th Annual Conference on Yeast Talk given at a conference Tracking lipid droplet fate during yeast mating 15.05.2018 Smolenice, Slovakia Schneiter Roger;


Communication with the public

Communication Title Media Place Year
Media relations: print media, online media LD_Biogenesis Cordis European Comission (in preperation) International 2020

Awards

Title Year
Dr. Stéphanie Cottier was featured in a "first person" article in Biol Open. https://bio.biologists.org/content/9/6/bio054171 2020
Dr. Vineet Choudhary received a call as Assistant Professor at the Department of Biotechnology, All India Institute of Medical Sciences, New Delhi. 2020
The JCB article by Dr. Vineet Choudhary was featured in a SPOTLIGHT commentary entitled "building the lipid droplet assembly complex" in JCB. https://rupress.org/jcb/article/219/7/e202006025/151882/Building-the-lipid-droplet-assembly The same article was featured in a Microreview in Microbial Cell. http://microbialcell.com/advance-publications/ 2020
Dr. Rabih Darwiche received a fellowship as beginning scientist from the SNF to start his postdoctoral work at the Harvard Medical School. 2017
Dr. Vineet Choudhary received a Marie Curie Fellowship from the European Comission Horizon 2020 Program to work on lipid droplet biogenesis. 2017

Associated projects

Number Title Start Funding scheme
177088 Characterization of Biomolecules by Mass Spectrometry 01.01.2018 R'EQUIP
182725 Deciphering pathways of fatty acid efflux and their use to optimize microbial cell factories 01.02.2019 Project funding (Div. I-III)
153416 Lipid Storage and Export 01.10.2014 Project funding (Div. I-III)

Abstract

This research program addresses two separate topics: the biogenesis of lipid droplets and their association with the membrane of the endoplasmic reticulum, and the structure and function of an important and widespread protein superfamily, known as CAP proteins. Lipid droplets are present in all eukaryotic cells and are discernible as round intracellular structures. They serve to store metabolic energy in form of neutral lipids, commonly known as “fat”. This energy is stored primarily in the fatty acids that are esterified in triacylglycerols and steryl esters, and is released upon ?-oxidation of these fatty acids. Lipid droplets are thus implicated in many of the pandemic diseases such as obesity, insulin resistance, atherosclerosis and lipotoxicity. The hydrophobic core of lipid droplets is covered by a phospholipid monolayer and they harbor a specific set of proteins, many of which function in neutral lipid metabolism, such as lipases or acyltransferases. We and others have previously shown that these lipid droplets are closely associated with the membrane of the endoplasmic reticulum. This association is functionally important as it allows the transfer of integral membrane as well as that of lipids between the two compartments. The precise nature of this association between the two compartments, however, has remained elusive. Recent results from our laboratory indicate that lipid droplets are accessible to proteins from within the luminal compartment of the endoplasmic reticulum, indicating that lipid droplets may form inside this luminal compartment. Should this indeed be the case, the biogenesis of lipid droplets would be similar to that of lipoprotein particles, which in essence are a functionalized, secreted form of lipid droplets. The aim of this part of the proposal thus is to define the nature of the association between the endoplasmic reticulum and lipid droplets. Therefore, we will determine whether endoplasmic reticulum resident integral membrane proteins can laterally move to localize over lipid droplets. In addition, we will compare the structure and topology of integral membrane proteins between their localization in the endoplasmic reticulum and their lipid droplet localization. The results of this study will thus provide new essential information to define and understand how exactly proteins and lipids can be exchanged between the endoplasmic reticulum and the lipid droplet storage compartment. This information is essential for an improved understanding of the etiology of lipid-related diseases.The CAP superfamily of proteins is named after the three founding members of this family, Cysteine-rich secretory proteins (CRISP), Antigen 5, and Pathogenesis-related 1 (PR-1). CAP family members are implicated in many fundamental biological processes, ranging from immune defense in mammals and plants, sperm maturation and fertilization, pathogen virulence, venom toxicity and even prostate and brain cancer. CAP family members are mostly secreted glycoproteins that are stable in the extracellular space. The mode of action of these proteins, however, has remained elusive. We could previously show that the CAP family members in yeast, known as Pathogen Related in Yeast (Pry), bind and thereby solubilize sterols and related hydrophobic compounds. Sterol binding is a conserved feature of many CAP proteins, including human CRISP2, a CAP superfamily member that is expressed in the testis and epididymis and participates in sperm-egg interaction during fertilization, and the plant PR-1 protein, which is synthesized in response to infections of plants with pathogens. Recent evidence indicates that these proteins not only are capable of binding sterols, but they independently also can bind fatty acids. We will test whether Pry proteins bind fatty acids, define the binding site by site-directed mutagenesis and test whether lipid binding of these proteins is important for their membrane association, permeabilization, and in vivo function. These results will thus help to define the molecular mode of action of these proteins and thereby improve our understanding of the physiological function these proteins exert in both health and disease.
-