Background Lobosphaera incisa (L. incisa) is an oleaginous microalga that stores triacylglycerol (TAG) rich in arachidonic acid in lipid bodies (LBs). This organelle is gaining attention in algal research, since evidence is accumulating that proteins attached to its surface fulfill important functions in TAG storage and metabolism.ResultsHere, the composition of the LB proteome in L incisa was investigated by comparing different cell fractions in a semiquantitative proteomics approach. After applying stringent filters to the proteomics data in order to remove contaminating proteins from the list of possible LB proteins (LBPs), heterologous expression of candidate proteins in tobacco pollen tubes, allowed us to confirm 3 true LBPs: A member of the algal Major Lipid Droplet Protein family, a small protein of unknown function and a putative lipase. In addition, a TAG lipase that belongs to the SUGAR DEPENDENT 1 family of TAG lipases known from oilseed plants was identified. Its activity was verified by functional complementation of an Arabidopsis thaliana mutant lacking the major seed TAG lipases.ConclusionsHere we describe 3 LBPs as well as a TAG lipase from the oleaginous microalga L. incisa and discuss their possible involvement in LB metabolism. This study highlights the importance of filtering LB proteome datasets and verifying the subcellular localization one by one, so that contaminating proteins can be recognized as such. Our dataset can serve as a valuable resource in the identification of additional LBPs, shedding more light on the intriguing roles of LBs in microalgae.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-017-1042-2) contains supplementary material, which is available to authorized users.

BackgroundOleaginous microalgae are promising sources of energy-rich triacylglycerols (TAGs) for direct use for food, feed and industrial applications. Lobosphaera incisa is a fresh water unicellular alga, which in response to nutrient stress accumulates a high amount of TAGs with a high proportion of arachidonic acid (ARA). The final committed step of de novo TAG biosynthesis is catalyzed by acyl-CoA:diacylglycerol acyltransferases (DGATs), which add a fatty acid (FA) to the final sn-3 position of diacylglycerol (DAG).ResultsGenome analysis revealed the presence of five putative DGAT isoforms in L. incisa, including one DGAT of type 1, three DGATs of type 2 and a single isoform of a type 3 DGAT. For LiDGAT1, LiDGAT2.1, LiDGAT2.2 and LiDGAT2.3 enzyme activity was confirmed by expressing them in the TAG-deficient yeast strain H1246. Feeding experiments of yeast transformants with fatty acids suggest a broad substrate specificity spectrum for LiDGAT1. A significant TAG production in response to exogenous ARA was found for LiDGAT2.2. Cellular localization of the four type 1 and type 2 DGATs expressed in yeast revealed that they all localize to distinct ER domains. A prominent association of LiDGAT1 with ER domains in close proximity to forming lipid droplets (LDs) was also observed.ConclusionsThe data revealed a distinct molecular, functional and cellular nature of type 1 and type 2 DGATs from L. incisa, with LiDGAT1 being a major contributor to the TAG pool. LiDGATs of type 2 might be in turn involved in the incorporation of unusual fatty acids into TAG and thus regulate the composition of TAG. This report provides a valuable resource for the further research of microalgae DGATs oriented towards production of fresh-water strains with higher oil content of valuable composition, not only for oil industry but also for human and animal nutrition.Electronic supplementary materialThe online version of this article (10.1186/s12870-018-1510-3) contains supplementary material, which is available to authorized users.

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