​Thesis presented May 05, 2023

Abstract:
ukaryotic cells are divided into several compartments delineated by one or more membranes. Organelle membranes are composed of a specific arrangement of lipids and proteins that ensure the identity of each compartment. As only a few cell compartments are able to synthesize their whole set of lipids, a massive exchange of lipids occurs between organelles. The transport of lipids can be mediated by vesicular pathways and/or by non-vesicular pathways. Non-vesicular lipid transport can be mediated by lipid transport proteins (LTPs) that facilitate the direct transfer of lipids from one membrane to another at membrane contact sites (MCSs). While LTPs have already been well investigated in yeast and human cells, only a small number have been characterized in plants. Phosphate (Pi) starvation is an abiotic stress commonly encountered by plants in nature during which a partial degradation of phospholipids occurs to increase the intracellular Pi availability. The degraded phospholipids are dynamically replaced by digalactosyl diacylglycerol (DGDG), a non-phosphorous plastid-synthetized lipid exclusively synthetized in the plastid. Lipid remodeling likely uses non-vesicular lipid transport routes highlighting its importance in the maintenance of membrane integrity during stress. Here, we characterize two actors involved in phosphate starvation-induced membrane remodeling: the mitochondrial transmembrane lipoprotein (MTL) complex and AtVPS13M1.
The MTL complex is a huge complex enriched in lipids and mainly composed of proteins located in both mitochondrial membranes. The MTL complex is important for intramitochondrial lipid trafficking during plant adaptation to Pi starvation. We attempt to purify the MTL complex by several biochemical techniques to allow the refinement of its composition and the study of its architecture and structure. Our results highlight the modular nature of the MTL complex that can presumably be found as a variable arrangement of different components. AtVPS13M1 is a vacuolar protein sorting 13 (VPS13) homolog in Arabidopsis thaliana. Recent data show that VPS13 proteins are huge LTPs implied in bulk lipid transport localized at multiple membrane contact sites between organelles. We identify six novel splice variants for AtVPS13M1 and show that the transcriptional landscape is widely dominated by only one presumably biologically relevant isoform. We show that AtVPS13M1 is able to bind multiple phosphoglycerolipids in vitro and participates in lipid homeostasis in Pi replete and deplete conditions. AtVPS13M1 localization remains to be further investigated but our preliminary studies suggest an association at least with the endoplasmic reticulum.
Overall, we provide an original approach in which we identify and characterize novel non-vesicular lipid transport routes important for Pi starvation-induced membrane lipid remodeling in plants.

Keywords:
MTL complex, VPS13, Phosphate starvation, Non-vesicular lipid transport, Mitochondria, Membrane contact sites