We have recently identified the lipid droplet organization (LDO) machinery in yeast, and its human homolog promethin/TMEM159. The LDO machinery is strongly enriched on a subpopulation of lipid droplets (LDs) located at the nucleus-vacuole junction contact site (NVJ).These LDs have a unique surface proteome, a feature that depends on the LDO component Ldo45. Nutrient stress results in pronounced accumulation of LDs at the NVJ, a process that depends on the presence of Ldo16. Despite these striking phenotypical observations, the exact functions of the LDO proteins are currently enigmatic. Here, we will functionally characterize the LDO-enriched LDs. Furthermore, we will dissect the mechanistic roles of the LDO proteins. Intriguingly, both LDOs and promethin are linked to the LD biogenesis protein seipin, a disease component related to the lipid storage disease BSCL2, and we will therefore include seipin in our analyses. Upon completion of this project, we will have detailed insights into how the conserved LDO machinery links contact site formation to biogenesis of specialized organelle subpopulations.