Mechanisms of ceramide sorting and transport between the endoplasmic reticulum and the Golgi apparatus in Saccharomyces cerevisiae.

• Applicant: Professor Dr. Florian Fröhlich
• Subject Area: Biochemistry
• Term: since 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 464504472

Ceramides are an important lipid class in all eukaryotic cells. They act as signaling molecules and are precursors for the complex sphingolipids which act as structural components of the plasma membrane. To be converted to complex sphingolipids, ceramides have to be transported from the endoplasmic reticulum to the Golgi apparatus. In mammalian cells, this step is mainly performed by non-vesicular transport via the ceramide transport protein CERT. In Saccharomyces cerevisiae, ceramides are transported by both, vesicular and non-vesicular transport. However, the molecular mechanisms underlying ceramide transport in yeast remain largely elusive. Analyzing high throughput chemical genetic interaction data, we have identified the yeast protein Svf1 as an important regulator of ceramide transport between the ER and the Golgi apparatus. Deletion of Svf1 results in decreased levels of complex sphingolipids and a concomitant accumulation of ceramides. In preliminary experiments we have identified an amphipathic helix at the N-terminus of Svf1 that targets Svf1 to the ER and the early Golgi apparatus. Based on these data we now propose to study the molecular mechanism of Svf1 interaction with membranes and especially ceramides in detail and analyze its role in ceramide sorting and transport. In the first aim we will analyze the interaction of Svf1 with model membranes in vitro and determine the lipid requirements for this interaction. In the second aim we will identify protein interaction partners necessary for the association of Svf1 with membranes and analyze their role in Svf1 dependent ceramide sorting. In the third aim we will identify the molecular mechanism of Svf1 dependent ceramide sorting and/or transport establishing in vitro ceramide sorting/transport assays. Completion of this project will shed light on the long standing question on how ceramides are transported between the ER and the Golgi in yeast. In addition, this project has the potential to uncover a completely novel mechanism on how lipids are sorted in cells.

• DFG Programme: Research Grants