On the stabilization of plant lipid droplets: Dynamic interplay between oleosins and phospholipids.

As central organelles for lipid and energy homeostasis in cells, lipid droplets (oleosomes) consist of a neutral lipid core - mainly triacylglycerols and sterol esters. Crucial to the droplet stability and dynamics, the droplet boundary is decorated by a dense phospholipid monolayer enriched in phosphatidylcholines (PCs) as well as surfactant-like proteins, which in plant cells, are predominantly oleosins (OLs). The monolayer keeps plant lipid droplets stable for years throughout seed dormancy yet delicately destabilizes them during germination to ensure the targeted release of their stored lipids. The respective contributions of PCs and OLs to the fine metastability of lipid droplets remain insufficiently understood. Here, we address this question using reconstituted oil-in-water emulsions stabilized by purified OLs and PCs in controlled microfluidic systems, enabling variation of interfacial OL:PC ratios and environmental conditions such as pH and ionic strength. Aided by super-resolution microscopy and dilatational rheology, we reveal that oleosins form a dense, elastic interfacial network protecting droplets from coalescence. The presence of PCs decreases the network density and interfacial elasticity, while their effect on coalescence resistance is condition-dependent. These findings indicate complementary roles of the two interfacial agents: OLs function as primary stabilizers that resist coalescence and provide interfacial mechanical robustness, whereas PCs reduce interfacial tension and attenuate the robustness by disrupting the OL organization. In conclusion, our reductionist approach provides insights into the metastable nature of seed lipid droplets by isolating the distinct interfacial-mechanical contributions of OLs and PCs and offers a basis for designing bioinspired emulsions with tunable and stimuli-responsive stability.