Regulation of cellular sphingomyelin dynamics – SPHINGODYNA

Our body contains thousands of different lipid species. These lipids are not randomly distributed in a cell. Even in one membrane, the lipid composition of the outer and inner leaflets of the lipid bilayer is different. However, how this asymmetry is built, maintained and regulated remains poorly understood as is the physiological significance of lipid asymmetry. Sphingomyelin (SM) is a major mammalian sphingolipid, which localizes almost exclusively to the outer leaflet of the plasma membrane. There, SM forms specific lipid domains together with cholesterol. These lipid domains are postulated to be involved in a number of pathophysiological events as diverse as membrane traffic to signal transduction. SM is also a reservoir of a bioactive lipid, ceramide. Outer leaflet SM is degraded by acid SMase (aSMase), which is a secreted enzyme. aSMase activity on the cell surface is involved in a number of events including apoptosis, viral infection and tumor metastasis. Interestingly enough, in addition to aSMase, cells contain cytosolic SMase, neutral SMase (nSMase). nSMase is also involved in a diverse set of signaling pathways, including apoptosis and exosome release. This raises an open question in the field of membrane biology. Since SM is predominantly at the outer leaflet, how can it be transferred to the inner leaflet?
Today, it is assumed that the asymmetric distribution of glycerophospholipids in the PM is controlled on site by flippases, floppases, and scramblases. Bulk SM is synthesized on the lumenal side of the Golgi apparatus by SM synthase 1. SM is then transported to the outer leaflet of the PM by vesicular traffic, whereas local SM is synthesized on the extracellular side of the PM by SM synthase 2. Thus, the asymmetric distribution of SM is established by the SM synthases. No protein that catalyzes the transbilayer movement of SM has been identified, however evidences show that SM is also present at the inner leaflet of the PM. Using a dedicated genome-wide screen, we identified protein candidates potentially regulating the asymmetric distribution of SM at the PM. We establish for one candidate PMP2 that it functions at dynamic deformation of the PM to induce transbilayer lipid movement.. We have identified a second protein candidate GGA1, an intracellular membrane traffic protein and we hypothesize that GGA1 functions at the organelle level to induce transbilayer lipid movement of SM. This project will address the mechanism by which GGA1 and accessory protein(s) regulate the transbilayer movement of SM.
Up to recently SM was supposed to be only at the extracellular side of the PM, very little is known about its role on the intracellular side. Several lines of evidence suggest that the presence of inner leaflet SM is important in physiopathology. We are in a unique position to address more precisely the biological role of SM since genetic manipulation of identified proteins can alter the transbilayer distribution of SM. Together with overexpression of the cytosolic form of bacterial SMase (bSMase), we have new tools to study the function of SM. This project will study the effect of the alteration of SM asymmetry on membranes lipid content, composition and distribution. We will also clarify the effect on cytosolic nSMase-mediated exosome release. Our results will reveal the function of SM asymmetry in pathophysiology.