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Our lab is working on lipid metabolism, using Baker’s yeast as a unicellular model organism. We are looking at two different questions to understand how cells regulate and adapt their lipid composition to changes in environmental conditions to maintain a homeostatic balance:
Lipid droplets are intracellular globular shaped structures that serve to store metabolic energy in form of neutral lipids (fat). They dynamically form and grow in size under conditions of energy excess and they shrink upon energy demand. Lipid droplets are composed of a hydrophobic core made of triacylglycerol and steryl esters. This neutral lipid core is covered by a phospholipid monolayer and harbors a set of proteins that are specifically targeted to the lipid droplet surface. While yeast cells typically harbor up to a dozen of these droplets, adipocytes are filled with one large lipid droplet. As the core of the lipid droplet is highly hydrophobic and not soluble in the aqueous environment, the synthesis, transport and storage of these lipids has to be orchestrated with the biogenesis of lipid droplets. We are using live cell microscopy to understand how these processes are coordinated and where exactly they occur in the cell. Our data indicate that lipid droplets are in close proximity to the endoplasmic reticulum (ER) allowing for rapid and bidirectional exchange of both lipids and proteins between the ER membrane and the lipid droplets.
Commentary: J Cell Sci 2016 129: e2005
A close link between lipid droplets and the ER lumen
The cell stores metabolic energy in globular intracellular structures called lipid droplets; these associate with a number of organelles, including mitochondria, vacuoles, peroxisomes and the endoplasmic reticulum (ER). The droplets contain neutral lipids, such as triacylglycerols and steryl esters, which are synthesised by enzymes located in the ER. Moreover, an exchange of proteins and lipids between lipid droplets and the ER has been observed. However, the exact nature of the interface between the ER and lipid droplets is still not understood. In this issue (p. 3803) Roger Schneiter and colleagues demonstrate that lipid droplets are accessible to ER proteins from within the ER luminal compartment by tagging cytosolic lipid droplet proteins with a signal sequence found in an ER lumen glycoprotein. Surprisingly, these engineered proteins are still targeted to lipid droplets. The tagged proteins are glycosylated and protected from protease-mediated cleavage, consistent with a location at luminal sites of the ER. Further, expression of the ER luminal proteins does not alter lipid droplet morphology, and, using a yeast mating assay, the authors show that proteins targeted to the ER lumen can access both mature and newly formed lipid droplets. Moreover, as demonstrated here, ER luminal proteins are also able to access lipid droplets in mammalian cells. With this work, the authors thus reveal that lipid droplets are accessible from both luminal and cytosolic sites of the ER.
© 2016. Published by The Company of Biologists Ltd
Mishra S., Khaddaj R., Cottier S., Stradalova V., Jacob C., and Schneiter R. (2016). Mature lipid droplets are accessible to ER luminal proteins. Journal Cell Science 129: 3803-3815 (link).
Jacquier N., Mishra S., Choudhary V., and Schneiter R. (2013). Expression of oleosin and perilipins in yeast promote formation of lipid droplets from the endoplasmatic reticulum. J. Cell Science 126: 5198-5209 (link).
Jacquier N., Choudhary V., Mari M., Toulmay A., Reggiori F., and Schneiter, R. (2011). Lipid droplets are functionally connected to the endoplasmic reticulum in Saccharomyces cerevisiae. J. Cell. Sci. 124: 2424-2437 (link).
Proteins belonging to the CAP superfamily are found in all kingdoms of life (Pfam PF00188). This superfamily was named after three founding members: Cysteine-rich secretory proteins (CRISP) found in mammals, antigen 5 (Ag5) in stinging insects and pathogenesis-related protein 1 (PR-1) in plants. The human genome encodes for 32 family members, whereas yeast has 3 CAP members, termed Pathogen Related in Yeast, Pry1-3. These proteins have been implicated in many different physiological processes ranging from immune defense in mammals and plants, sperm maturation and fertilization, prostate and brain cancer, pathogen virulence, and venom toxicity. They are most often secreted glycoproteins that adapt a unique alpha-beta-alpha sandwich fold. We found that these proteins bind both sterols and fatty acids in two independent lipid binding sites and they promote the export of these lipids in vivo. We are now testing whether the ability of these proteins to sequester lipids can account for their physiological functions reported for different organism.
Structural model of a possible open-loop conformation of the yeast Pry1 protein that could accommodate small hydrophobic molecules such as cholesterol. For illustration purposes, the cholesterol molecule (green) was manually positioned so as to fill optimally the open cavity.
Darwiche R., El Atab O., Cottier S., and Schneiter R. (2018): The function of yeast CAP family proteins in lipid export, mating and pathogen defense. FEBS Letters 592: 1304-1311 (link).
Darwiche R., Mène-Saffrané L., Gfeller D., Asojo O.A., and Schneiter R. (2017). The pathogen-related yeast protein Pry1, a member of the CAP protein superfamily, is a fatty acid-binding protein. Journal of Biological Chemistry 292: 8304-8314 (link).
Choudhary V., Darwiche R., Gfeller D., Zoete V., Michielin O., and Schneiter R. (2014). The caveolin-binding motif of the pathogen related yeast protein Pry1, a member of the CAP protein superfamily, is required for in vivo export of cholesteryl acetate. Journal of Lipid Research 55: 883-894 (link).
Choudhary V., and Schneiter R. (2012). Pathogen-Related Yeast (PRY) proteins and members of the CAP superfamily are secreted sterol-binding proteins. PNAS 109: 16882-16887 (link).