Three Distinctive Diacylglycerol Acyltransferases (DGAT1, DGAT2, and DGAT3) from <i>Perilla frutescens</i> and Their Potential in Metabolic Engineering for Designed Oil Production

Diacylglycerol acyltransferase (DGAT) catalyzes final acylation in triacylglycerol (TAG) biosynthesis. However, the mechanism underlying polyunsaturated fatty acids (PUFAs) in TAGs remains unclear for <i>Perilla frutescens</i>. Here, three novel UFA-preferring PfDGAT enzymes (PfDGAT1A, 2A, and 3B) were systematically examined <i>ex vivo</i> in <i>Saccharomyces cerevisiae</i> and <i>in vitro</i>. Genetic modulation of each of these <i>PfDGAT</i> in <i>Nicotiana tabacum</i> and <i>Chlamydomonas reinhardtii</i> significantly increased the oil content and accumulation of the target UFA-TAG (LA-, OA-, or ALA-TAG) in tobacco leaves and seeds as well as algal cells. Synergetic manipulation on two of them (PfDGAT1A+2A, 1A+3B, or 2A+3B) resulted in great enhancement of total oil and predominant synthesis of the two target UFA-TAGs (LA/OA-, LA/ALA-, or OA/ALA-TAGs) at about a 1:1 ratio in heterogeneous hosts. Moreover, the transgenic lines showed no negative phenotype. These findings demonstrate the functional TAG distribution pattern of three PfDGATs, providing valuable metabolic engineering targets in both higher plant and industrial microorganism systems for designed oil production.