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Role of lipids in COPII vesicle formation

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Defense Thèse de doctorat : Univ. Genève, 2016 - Sc. 4896 - 2016/01/22
Abstract Inside the cell, transport of proteins and lipids between organelles is mediated by membrane carriers known as vesicles. The formation of these vesicles requires the participation of coat proteins which are capable to change the shape of membranes into extremely curved structures, ultimately catalyzing the scission and release of a free vesicle to the cytoplasm. Vesicular transport from the endoplasmic reticulum (ER) to the Golgi apparatus is mediated by Coated Protein complex II (COPII). COPII vesicles are formed in discrete areas known as ER exit sites (ERES). This coat complex comprises several proteins which bind sequentially to the membrane. Initially, the transmembrane protein Sec12 recruits the small GTPase Sar1, which on its GTP form release an insert an alpha-helix on the ER surface. Then the heterodimer Sec23/24 is recruited to the membrane, which in coordination with Sar1 and cargo proteins form a pre-budding complex. On a latter step, the heterotetramer Sec13/31 scaffolds over the pre-budding complex and completes membrane deformation to form a bud. The last step requires the scission of the vesicle from the ER membrane, a process that is not yet well characterized but that may depend on Sar1. The lipid requirements to deform the ER membrane are largely unknown. In this study it has been found a novel link between lipids and COPII vesicle formation in Saccharomyces cerevisiae. A mutation that affects COPII proteins recruitment, sec12-4, can be rescued by the overexpression of phospholipases, which are enzymes that hydrolyze glycerophospholipids into lysophospholipids and free fatty acids. Lysophospholipids are conical lipids whose accumulation may induce changes in curvature or membrane fluidity. Lipidomics have revealed that lysophospholipids accumulate in rescued sec12-4 mutants overexpressing these phospholipases. Furthermore, COPII vesicles have been generated with in vitro budding techniques and the lipidome analyzed by mass spectrometry. Interestingly, these vesicles had four fold more lysophospholipids than the the proportion of lysophospholipids present in the ER. Finally, in vitro reconstitutions have been used to explore the effect of lysophospholipids on the binding of COPII proteins to artificial liposomes. Confocal microscopy images have revealed that lysophospholipids increase the binding of Sec13/31 to the membrane and affects the organization of the coats over the surface of giant liposomes. The results obtained in this study establish a new link between lysophospholipids and COPII vesicle formation. Their extreme conical shape may cause changes in membrane rigidity or packing defects and by that help COPII coats to deform ER membrane.
Keywords CellEndoplasmic reticulumVesicleCopiiLipidomicsBiophysicsLysophospholipidConical lipidMembraneMembrane deformationMembrane trafficCell biologyYeastSaccharomyces cerevisiaeIn vitro reconstitution
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URN: urn:nbn:ch:unige-852586
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Research groups Groupe Riezman
Groupe Roux
Project NCCR Chemical Biology
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MELERO CARRILLO, Alejandro. Role of lipids in COPII vesicle formation. Université de Genève. Thèse, 2016. https://archive-ouverte.unige.ch/unige:85258

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Deposited on : 2016-07-13

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