Nitrogen starvation-inducible promoter of microalga Neochloris oleoabundans lipogenic gene encoding diacylglycerol acyltransferase 2

Abstract

Microalgal lipid triacylglycerol (TAG) is a promising source for sustainable production of biofuels and edible oils. TAG biosynthesis in microalgae can be induced by nitrogen starvation (−N); however, regulation of the genes involved in this process is poorly known. To explore the regulation of gene encoding diacylglycerol acyltransferase 2 of oleaginous microalga Neochloris oleoabundans (NeoDGAT2) responsible for TAG biosynthesis, regulatory sequence of NeoDGAT2 gene (RDG) was cloned, and its functional regions were mapped by deletion analysis using the modified cyan fluorescent protein gene (CFP) as a reporter. The efficiency of CFP gene mTurquoise2 (Tq) without any intron, Tq1 and Tq2 with one and two copies of Chlamydomonas reinhardtii rbcS2 intron 1, respectively, was evaluated; Tq2 exhibited the highest CFP fluorescence activity in N. oleoabundans was therefore used as reporter for RDG deletion analysis. Deletion analysis of RDG revealed that the −N inducible region contained the predicted binding site of MYB transcription factor (MYB-bs). Specific binding between MYB-bs of RDG and the DNA-binding domain of MYB-related transcription factor ROC40 from C. reinhardtii was observed using electrophoretic mobility shift assay. Therefore, the corresponding MYB transcription factor in N. oleoabundans is probably the transcription factor regulating NeoDGAT2. The interaction between MYB transcription factor and the MYB-bs may play a role in regulating −N induced expression of NeoDGAT2, affecting TAG accumulation. MYB transcription factors can be the potential targets for engineering to increase TAG content. Increasing TAG content is essential for products derived from microalgal TAG to achieve economic viability.