The dynamics of leaf lipid droplets and the identification of a novel lipid droplet protein in Arabidopsis thaliana
Lipid droplets (LDs) are unique, ER-derived organelles found in all eukaryotic organisms. Structurally, LDs contain a core of energy-rich neutral lipids, such as triacylglycerols (TAGs), that is enclosed by a single phospholipid monolayer and decorated with a diverse array of ‘coat’ proteins. In plants, LDs are found in high abundance in pollen and oilseeds, and are ultimately mobilized in the latter as a carbon and energy source during post-germinative growth. While some of the proteins associated with seed LDs have been well-characterized, relatively little is known about LDs in other plant tissue/cell types, particularly in vegetative organs, such as leaves, and the proteins that govern their formation and activity. In this study, Arabidopsis thaliana was used as a model organism to investigate LD activity and to identify and characterize novel LD coat proteins. Using a variety of imaging techniques, the abundance of LDs in leaf cells was found to vary throughout the diurnal cycle and increased in response to both heat and cold stress treatments. Extended light and dark treatments were also shown to affect leaf LD abundance. The number of LDs in leaves appears to be regulated, at least in part, by lipid droplet associated proteins (LDAPs), a three-member family of constitutively-expressed LD coat proteins. Loss-of-function Arabidopsis ldap mutants were found to produce fewer LDs in leaf cells and exhibited a marked decrease in LD proliferation during temperature stress treatments. Putative interactors of LDAP3 were also identified using a Y2H assay and investigated to determine their possible involvement in LD-related processes. One of these interactors, termed LDAP-interacting protein (LDIP) was determined to be a ubiquitously- expressed, bona fide LD coat protein that regulates LD size and neutral lipid homeostasis in both leaves and seeds. Subsequent localization experiments indicated that LDAP3 recruits LDIP to the LD surface during biogenesis and that LDIP may modulate the phospholipid composition of the LD monolayer. This research has allowed for the generation of a model for plant LD formation in non-seed tissues.