Metabolic engineering of biomass for high energy density: oilseed‐like triacylglycerol yields from plant leaves

Abstract

High biomass crops have recently attracted significant attention as an alternative platform for the renewable production of high energy storage lipids such as triacylglycerol (TAG). While TAG typically accumulates in seeds as storage compounds fuelling subsequent germination, levels in vegetative tissues are generally low. Here, we report the accumulation of more than 15% TAG (17.7% total lipids) by dry weight in Nicotiana tabacum (tobacco) leaves by the co-expression of three genes involved in different aspects of TAG production without severely impacting plant development. These yields far exceed the levels found in wild-type leaf tissue as well as previously reported engineered TAG yields in vegetative tissues of Arabidopsis thaliana and N. tabacum. When translated to a high biomass crop, the current levels would translate to an oil yield per hectare that exceeds those of most cultivated oilseed crops. Confocal fluorescence microscopy and mass spectrometry imaging confirmed the accumulation of TAG within leaf mesophyll cells. In addition, we explored the applicability of several existing oil-processing methods using fresh leaf tissue. Our results demonstrate the technical feasibility of a vegetative plant oil production platform and provide for a step change in the bioenergy landscape, opening new prospects for sustainable food, high energy forage, biofuel and biomaterial applications.

Arabidopsis Lipins, PDAT1 Acyltransferase, and SDP1 Triacylglycerol Lipase Synergistically Direct Fatty Acids toward β-Oxidation, Thereby Maintaining Membrane Lipid Homeostasis

“…To date, a major effort has focused on the ectopic expression of WRINKLED1 (WRI1), a key seed-specific transcriptional regulator (Cernac and Benning, 2004), to increase the supply of FAs for TAG assembly (Sanjaya et al, 2011;Kelly et al, 2013;Vanhercke et al, 2014). We previously reported that disruption of the eukaryotic galactolipid pathway in the tgd1-1 mutant results in substantial increases in the rates of both FA synthesis and turnover in leaves (Fan et al, 2013a).…”

Inference and prediction of metabolic network fluxes

“…This also leads to the conclusion that if one is attempting to engineer the biomass composition of crops (e.g. to increase the energy density of bioenergy crops; Vanhercke et al, 2014), a rather more subtle and coordinated engineering of central metabolism may be required than is usually achieved with the sledgehammer of transgenic or mutagenic interventions. Ultimately, it may be preferable to attempt to directly manipulate the controls on biomass demand (Morandini, 2013) and to rely on the inherent plasticity and flexibility of central metabolism to meet the altered demand.…”

Multigene Engineering of Triacylglycerol Metabolism Boosts Seed Oil Content in Arabidopsis

“…simultaneously alter the expression of multiple genes) and test empirically whether combinations provide a better effect. Surprisingly, there are currently no academic reports of gene stacking to enhance seed oil content, although several recent studies have successfully applied this approach to manipulate oil metabolism in leaves (Sanjaya et al, 2011;Fan et al, 2013;Kelly et al, 2013b;Vanhercke et al, 2013Vanhercke et al, , 2014Winichayakul et al, 2013). In general these studies suggest that a push-pull-protect strategy may be effective (Fig.…”