In Vivo Packaging of Triacylglycerols Enhances Arabidopsis Leaf Biomass and Energy Density    

Abstract

Our dependency on reduced carbon for energy has led to a rapid increase in the search for sustainable alternatives and a call to focus on energy densification and increasing biomass yields. In this study, we generated a uniquely stabilized plant structural protein (cysteine [Cys]-oleosin) that encapsulates triacylglycerol (TAG). When coexpressed with diacylglycerol O-acyltransferase (DGAT1) in Arabidopsis (Arabidopsis thaliana), we observed a 24% increase in the carbon dioxide (CO 2 ) assimilation rate per unit of leaf area and a 50% increase in leaf biomass as well as approximately 2-, 3-, and 5-fold increases in the fatty acid content of the mature leaves, senescing leaves, and roots, respectively. We propose that the coexpression led to the formation of enduring lipid droplets that prevented the futile cycle of TAG biosynthesis/lipolysis and instead created a sustained demand for de novo lipid biosynthesis, which in turn elevated CO 2 recycling in the chloroplast. Fatty acid profile analysis indicated that the formation of TAG involved acyl cycling in Arabidopsis leaves and roots. We also demonstrate that the combination of Cys-oleosin and DGAT1 resulted in the highest accumulation of fatty acids in the model single-cell eukaryote, Saccharomyces cerevisiae. Our results support the notion that the prevention of lipolysis is vital to enabling TAG accumulation in vegetative tissues and confirm the earlier speculation that elevating fatty acid biosynthesis in the leaf would lead to an increase in CO 2 assimilation. The Cysoleosins have applications in biofuels, animal feed, and human nutrition as well as in providing a tool for investigating fatty acid biosynthesis and catabolism.

Understanding the Biochemical Basis of Temperature-Induced Lipid Pathway Adjustments in Plants

“…Triacylglycerols (TAGs), although representing <1% of total lipids in Arabidopsis leaf tissues, has been implicated in shortterm glycerolipid intermediate provision during membrane lipid remodeling (James et al, 2010;Fan et al, 2013;Winichayakul et al, 2013). The breakdown of TAG in plants can be mediated by multiple TAG lipases (El-Kouhen et al, 2005;Eastmond, 2006).…”

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.…”

“…1A). For example, expression of the WRINKLED1 (WRI1) transcription factor can be used to drive glycolysis and fatty acid synthesis (Cernac and Benning, 2004), expression of DIACYLGLYCEROLACYL TRANSFERASE1 (DGAT1) can be used to draw fatty acids into TAG (Jako et al, 2001), and turnover can also be minimized by disrupting lipolysis (Fan et al, 2013;Kelly et al, 2013b;Vanhercke et al, 2013;Winichayakul et al, 2013). Here, we investigate the effects of stacking these gene modifications on seed oil content and yield in Arabidopsis.…”

Breaking through the feed barrier: options for improving forage genetics

“…A technology to increase plant lipid concentrations and plant biomass has been recently reported (Winichayakul et al 2013). A synthetic gene encoding a novel lipid-encapsulation protein, cysteine oleosin, was constitutively co-expressed with a gene encoding diacylglycerol acyltransferase (DGAT1), to increase energy density and reduce photorespiration of the model species Arabidopsis thaliana via the production and encapsulation of neutral lipids (Winichayakul et al 2013).…”

“…A synthetic gene encoding a novel lipid-encapsulation protein, cysteine oleosin, was constitutively co-expressed with a gene encoding diacylglycerol acyltransferase (DGAT1), to increase energy density and reduce photorespiration of the model species Arabidopsis thaliana via the production and encapsulation of neutral lipids (Winichayakul et al 2013). The leaf lipid concentration was doubled to~8% DM, and lipid concentration in the plant root also increased to 8% of the DM.…”