Enhanced triacylglycerol production in oleaginous microalga Neochloris oleoabundans by co-overexpression of lipogenic genes: Plastidial LPAAT1 and ER-located DGAT2

Abstract

Microalgae accumulate lipid triacylglycerol (TAG), a promising feedstock for production of natural edible oils and biofuels. To make products derived from microalgal TAG economically viable, increasing TAG content and productivity are of high importance. To increase TAG content, two endogenous key enzymes of TAG biosynthesis: plastidial lysophosphatidic acid acyltransferase (NeoLPAAT1) and endoplasmic reticulum-located diacylglycerol acyltransferase 2 (NeoDGAT2) were co-overexpressed in oleaginous microalga Neochloris oleoabundans. The neutral lipid content in NeoLPAAT1-NeoDGAT2 co-overexpressing transformant detected by Nile red staining increased 2-fold without compromising cell growth. The transcriptional levels of NeoLPAAT1 and NeoDGAT2 levels were 1.9-fold higher in the transformant than wild type. Considerably higher lipid accumulation was found in the transformant than wild type: total lipid content (73.72 ± 4.17 % DCW) increased 1.6-fold, TAG content (50.63 ± 6.15 % DCW) increased 2.1-fold, total lipid productivity (16.84 ± 0.66mg/L/day) increased 1.9-fold, and TAG productivity (11.68 ± 0.90mg/L/day) increased 2.1-fold. Fatty acid composition was slightly altered in the transformant compared to wild type; saturated fatty acid C16:0 increased to 26% from 20%, whereas C18:0 was reduced to 7% from 14%. Long-term stability of NeoLPAAT1-NeoDGAT2 co-overexpression was observed in the transformant continuously maintained on solid medium in a period of 4 years. The results suggested that targeted engineering of genes in pathway located at different organelles should be possible in microalgal lipid metabolism.