A hallmark of eukaryotic cells is the compartmentalization of specific functions within different membrane-enclosed organelles. The unique lipid composition of the various intracellular organelles is essential to preserve their distinct structural and functional identities, and requires tight regulation of lipid sorting, metabolism and transport. Disruption of lipid trafficking and metabolism leads to human pathologies such as obesity, diabetes, cancer and neurodegenerative disorders. Lipid transport can be mediated by vesicular trafficking but also by less well-understood vesicle-independent routes, the latter often occurring at sites of close contact between organelles (membrane contact sites, MCS). Increasing lines of evidence suggest that lipid transfer proteins (LTPs) play a major role in regulating lipid composition of membranous organelles and are particularly enriched at MSC, where they are thought to facilitate exchange of lipids. Non-vesicular lipid transport at MCS is particularly important in the case of the mitochondria that are largely excluded from the classical vesicular pathways.
The endoplasmic reticulum (ER) is the major site of lipid synthesis in eukaryotic cells and forms an extensive and dynamic membrane network that spreads throughout the cell and is engaged in MCS with nearly every other organelle, including mitochondria, lipid droplets and plasma membrane. Some of the components of MCS have started to be identified and the list is rapidly growing. However, MCS are still not well understood because most of their components remain to be identified and their visualization within the narrow space of MCS is a challenging problem, given that they are presents on membranes only a few dozen nanometers apart.
Our team studies the role of lipid trafficking at MCS in the maintenance of organelle structural and functional identities in mammalian cells. In particular we are interested in understanding how lipids are exchanged at ER-mitochondria MCS, and how these transport activities impact on mitochondria morphology, function and dynamics.
Mitochondria are dynamic organelles that participate in a plethora of biological processes including energy conversion, metabolism, signaling and apoptosis, and are therefore of utmost importance for cell viability. To fulfill their multiple tasks, mitochondria adopt a variety of morphologies resulting from a constant reshaping by fusion and fission events. This dynamic behavior imposes the necessity to maintain a defined protein and lipid composition of both outer and inner mitochondria membranes in order to preserve the functional integrity of mitochondria and the spatial organization of complex biochemical reactions. Since mitochondria do not have the enzymes to synthesize all their lipids and lipid precursors, they require a continuous and coordinated supply of membrane lipids from the ER to carry out their physiological processes and maintain their membrane integrity. However, the mechanisms and the LTPs mediating these lipid transfer activities are still largely unknown. Several proteins have been proposed to tether ER-mitochondria membranes in mammalian cells but none of them has been shown to play a direct role in phospholipid trafficking. Moreover, whether and how lipid exchange at these MCS impact on mitochondria functions and dynamics, is still unknown.
We have recently found that the lipid transfer proteins (LTPs) ORP5 and ORP8 [members of the Oxysterol-binding protein (OSBP)-related protein (ORP) family] are novel components of ER-mitochondria MCS, in addition to ER-PM contacts. We have also shown that these proteins are required for mitochondria morphology and respiratory function and we hypothesize that their primary function is to mediate lipid transport (i.e. Phosphatidylserine) at ER-mitochondria MCS. Studying the role of these proteins and of their binding partners offers the unique opportunity to functionally characterize ER-mitochondria MCS involved in lipid transport.
By using a combination of in-situ (using mammalian cells) imaging and morphological approaches including electron microscopy (EM), biochemical and subcellular fractionation techniques, as well as cell-free lipid transport assays we aim :
1. To biochemically, morphologically and functionally characterize ER-mitochondria MCS involved in lipid transport
2. To study the impact of lipid transport at ER-mitochondria MCS on mitochondria morphology, activity and dynamics.
3. To study how lipid transport activities at ER-mitochondria MCS are coordinated with intracellular lipid trafficking pathways at other MCS (i.e. MCS with lipid droplets).
Understanding the functions of ER-mitochondria MCS is not just important for comprehending fundamental physiological processes but also for understanding pathogenic processes in various diseases (such as neurodegenerative disorders) induced by disruption of these MCS.
