Resumen de Role of Lipids in Protein Sorting From the Endoplasmic Reticulum
Protein sorting in the secretory pathway is crucial to maintain cellular compartmentalization and homeostasis. In addition to coat-mediated sorting, the role of lipids in driving protein sorting during secretory transport is a long-standing fundamental question that still remains unanswered. The present thesis aimed to directly address this central issue by investigating in yeast whether and how specific membrane lipids are involved in differential protein export from the endoplasmic reticulum (ER), the first essential transport step of the secretory pathway. We conduct 3D simultaneous multi-color high resolution live imaging to demonstrate that newly synthesized lipid-linked plasma membrane proteins, glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) having a very long chain ceramide (C26) lipid moiety, are clustered and sorted into specialized endoplasmic reticulum exit sites (ERES) that are distinct from those used by transmembrane plasma membrane proteins during their export from the ER. Furthermore, we show that the chain length (C26) of ceramide in the ER membrane is critical for this sorting selectivity. We also show that, in addition to the presence of C26 ceramide in the ER membrane, clustering of GPI-APs requires binding to the transmembrane p24 protein complex, the cargo receptor that specifically exports GPI-APs from the ER. Using molecular dynamics simulations, we found that membrane ceramides accumulate around the cytosolic leaflet of the p24 transmembrane helix. Based on our results we propose a mechanism for C26 ceramide-based sorting by which the local enrichment of C26 ceramide in both leaflets of the ER membrane upon multivalent p24 binding of GPI-APs with C26 ceramide in the luminal leaflet and p24 association of free C26 membrane ceramide in the cytosolic leaflet could promote concomitant protein clustering and membrane curvature by C26 acyl chain interdigitation. In sum, our study provides in vivo evidence for lipid chain length-based protein cargo sorting into selective export sites of the secretory pathway and better comprehension of potential mechanism for this lipid-based sorting.
