Lipid droplets (LDs) are ubiquitous organelles that play a critical role in lipid homeostasis by regulating the storage and mobilization of fatty acids. When nutrient supplies are limited, the fatty acids stored in LDs are transferred to the mitochondria for the generation of energy. As starvation persists however, autophagy is triggered to degrade and recycle damaged organelles. Autophagy releases FAs into the cytosol to replenishing LDs with new FAs, increasing LD abundance. Interestingly, the autophagy-dependent LDs cluster in close proximity to mitochondria, forming membrane contact sites (MCSs). MCSs are areas of close apposition between the membranes of two organelles mediated by tethering forces that arise from protein-protein interaction. However, the identity of a LD-mitochondrial tethering complex and the functional importance of the LD-mitochondrial contacts in FA transfer and energy homeostasis are unknown.

In chapter one, we observed an increase in LD abundance during prolonged nutrient deprivation. Starvation-induced LDs are not required for fatty acid delivery to mitochondria, but instead LDs sequester fatty acids that are released during autophagy to prevent lipotoxic dysregulation of mitochondrial function. In chapter two, we established an inducible heterodimer tethering system to temporally and spatially regulate LD-mitochondrial contact to study the functional importance of the interorganelle contact. Finally, in chapter three we employed proximity-labeling proteomics to identify protein tethers that mediate the close positioning of LDs and mitochondria. Together, our studies advance our understanding of organelle communication in the maintenance of lipid and energy homeostasis.