Enhanced triacylglycerol (TAG) and protein accumulation in transgenic diatomThalassiosira pseudonanawith altered photosynthetic pigmentation

Abstract

Microalgal productivity in mass cultures is limited by the inefficiency with which available light energy is utilized. In dense cultures, cells closest to the light source absorb more light energy than they can use and dissipate the excess, while light penetrance into the culture is steeply attenuated. Reducing microalgal light harvesting and/or dissipating capacity per cell may improve total light utilization efficiency in mass cultures. In this study, two transgenic lines of the diatom Thalassiosira pseudonana with altered photosynthetic pigment content are evaluated with respect to photosynthetic parameters, growth, and macromolecule accumulation. In one line, violaxanthin de-epoxidase-like 2 (VDL2) is overexpressed (OE), resulting in a reduction of the diadinoxanthin cycle pigments, which are involved in light energy dissipation (non-photochemical quenching, NPQ), accompanied by a stoichiometric increase in the light-harvesting pigment fucoxanthin. No differences in the maximum potential quantum yield of photosystem II (Fv/Fm) or light-limited photosynthetic rate (α) were found. However, when adapted to 30 µmol photons m -2 sec -1 , the VDL2 OE maximum relative electron transport rate (rETRmax) upon exposure to saturating light intensities was 86-95% of wild type (WT). When adapted to 300 µmol photons m -2 sec -1 , VDL2 OE saturated photosynthesis at 62-71% of the light intensity needed to saturate WT (Ek). NPQ was substantially lower at and below 300 µmol photons m -2 sec -1 . VDL2 OE accumulated up to 3.4 times as much triacylglycerol (TAG) as WT during exponential growth, and up to twice as much protein. Growth in terms of culture density was up to 7% slower. TAG and protein accumulation inversely correlated with NPQ. The second line evaluated was obtained by using antisense RNA to simultaneously silence or knock down (KD) both LUT1-like (LTL) genes, hypothesized to catalyze an intermediate carotenoid biosynthesis step of converting β-carotene to zeaxanthin. Overall reduction of photosynthetic pigment content without altering the relative abundance of individual pigments resulted. No significant differences in photosynthetic parameters compared to WT were found. LTL KD grew at a rate comparable to WT and accumulated up to 40% more TAG during exponential growth, while protein content was reduced by 11-19%. LTL KD cells were elongated and 5-10% smaller than WT, and cultures contained auxospores, indicating stress that may relate to a cell cycle progression defect.Conception and design (OG, MH), experimental work (OG, DPY), data analysis and interpretation (OG), drafting of the manuscript (OG), critical revision for important intellectual content and final approval (OG, DPY). MH is deceased.

Engineering the Unicellular Alga Phaeodactylum tricornutum for Enhancing Carotenoid Production

“…Vdr is a high light-induced gene and appears to be generally present in all chlorophytes [ 24 ]; it supposedly participates in de-epoxidation reactions in addition to VDE at a certain threshold of light and lumen acidification [ 30 ]. VDL2 has been recently overexpressed in the diatom Thalassiosira pseudonana , leading to an increase in fucoxanthin and a reduction in Dd-Dt, without a net change in their sum nor in β-carotene, suggesting that VDL2 is required in the step leading from the Dd-Dt pigments to fucoxanthin [ 31 ]. Reactions from violaxanthin to fucoxanthin and enzymes involved are still unknown, but it had been hypothesized that the reactions are catalysed by VDLs proteins.…”