Limited fatty-acid supply from the plastid and active catabolism of triacylglycerol prevent the accumulation of triacylglycerol in Coccomyxa sp. strain Obi grown under nitrogen-replete conditions

Abstract

The Coccomyxa sp. strain Obi is a unicellular green alga that accumulates >40% triacylglycerol (TAG) per cell dry weight under nitrogen (N)-limiting conditions. In this study, we analyzed the de novo synthesis of TAG under N-replete and N-limiting conditions using 14C tracers and quantitative reverse transcription PCR (RT-qPCR) approaches. In cells grown under N-replete conditions, the incorporation of photosynthetically fixed 14C into TAG was <5% of that incorporated into polar lipids, whereas the incorporation of exogenous 14C-fatty acids into TAG was approximately 50% of that incorporated into polar lipids. These results demonstrated that enzyme activities that convert fatty acids to TAG are appreciably expressed under N-replete conditions; however low fatty acid supply limits the TAG synthesis in N-replete cells. In agreement with this inference, the expression of FAT1 that encodes acyl-ACP thioesterase, which releases free fatty acids from acyl-ACP, was poorly expressed under N-replete conditions but induced under N-limiting conditions. Meanwhile, pulse-chase labeling experiments demonstrated a significant turnover of TAG under N-replete conditions. Taken together, we propose that both the limited carbon flux into TAG and active catabolism of TAG prevented the accumulation of TAG under N-replete conditions, whereas the increased carbon flux into TAG and possibly decreased catabolism of TAG contributed to the enhanced accumulation of TAG under N-limiting conditions. Furthermore, pulse-chase experiments demonstrated that the rate of TAG synthesis was much slower than that of polar lipid synthesis under N-limiting conditions, partly because of the slow conversion of diacylglycerol to TAG.