Identification and Characterization of Lipid Droplet-Associated Protein (LDAP) Isoforms from Tung Tree (Vernicia fordii)

Lipid droplets (LDs) are cytoplasmic organelles responsible primarily for the storage of neutral lipids, such as triacyclglycerols (TAGs). Derived from the endoplasmic reticulum bilayer, LDs are composed of a hydrophobic lipid core encased by a phospholipid monolayer and surface-associated proteins. To date, only a relatively few LD 'coat' proteins in plants have been identified and characterized, most of which come from studies of the model plant <i>Arabidopsis thaliana</i>. To expand our knowledge of the plant LD proteome, the LD-associated protein (LDAP) family from the tung tree (<i>Vernicia fordii</i>), whose seeds are rich in a commercially valuable TAG containing the conjugated fatty acid α-eleostearic acid (C18:3Δ^{9<i>cis</i>,11<i>trans</i>,13<i>trans</i>} [α-ESA]), was identified and characterized. Based on the tung tree transcriptome, three <i>LDAP</i> isoforms (<i>VfLDAP1-3</i>) were elucidated and the encoded proteins distinctly clustered into three clades along with their respective isoforms from other angiosperm species. Ectopic expression of the <i>VfLDAPs</i> in <i>Nicotiana benthamiana</i> leaves revealed that they localized specifically to LDs and influenced LD numbers and sizes, as well as increasing TAG content and altering TAG fatty acid composition. Interestingly, in a partially reconstructed TAG-ESA biosynthetic pathway, the co-expression of <i>VfLDAP3</i> and, to a lesser degree, <i>VfLDAP2</i>, significantly increased the content of α-ESA stored within the LDs. These results suggest that the VfLDAPs can influence the steady-state content and composition of TAG in plant cells and that certain LDAP isoforms may have evolved to more efficiently package TAGs into LDs containing unusual fatty acids, such as α-ESA.