Characterization of the diversification of phospholipid:diacylglycerol acyltransferases in the green lineage

Triacylglycerols have important physiological roles in photosynthetic organisms, and are widely used as food, feed and industrial materials in our daily life. Phospholipid:diacylglycerol acyltransferase (PDAT) is the pivotal enzyme catalyzing the acyl-CoA-independent biosynthesis of triacylglycerols, which is unique in plants, algae and fungi, but not in animals, and has essential functions in plant and algal growth, development and stress responses. Currently, this enzyme has yet to be examined in an evolutionary context at the level of the green lineage. Some fundamental questions remain unanswered, such as how PDATs evolved in photosynthetic organisms and whether the evolution of terrestrial plant PDATs from a lineage of charophyte green algae diverges in enzyme function. As such, we used molecular evolutionary analysis and biochemical assays to address these questions. Our results indicated that PDAT underwent divergent evolution in the green lineage: PDATs exist in a wide range of plants and algae, but not in cyanobacteria. Although PDATs exhibit the conservation of several features, phylogenetic and selection-pressure analyses revealed that overall they evolved to be highly divergent, driven by different selection constraints. Positive selection, as one major driving force, may have resulted in enzymes with a higher functional importance in land plants than green algae. Further structural and mutagenesis analyses demonstrated that some amino acid sites under positive selection are critically important to PDAT structure and function, and may be central in lecithin:cholesterol acyltransferase family enzymes in general.