Metabolic engineering of lipid catabolism increases microalgal lipid accumulation without compromising growth

Abstract

Biologically derived fuels are viable alternatives to traditional fossil fuels, and microalgae are a particularly promising source, but improvements are required throughout the production process to increase productivity and reduce cost. Metabolic engineering to increase yields of biofuel-relevant lipids in these organisms without compromising growth is an important aspect of advancing economic feasibility. We report that the targeted knockdown of a multifunctional lipase/phospholipase/acyltransferase increased lipid yields without affecting growth in the diatom Thalassiosira pseudonana. Antisense-expressing knockdown strains 1A6 and 1B1 exhibited wild-type-like growth and increased lipid content under both continuous light and alternating light/dark conditions. Strains 1A6 and 1B1, respectively, contained 2.4-and 3.3-fold higher lipid content than wild-type during exponential growth, and 4.1-and 3.2-fold higher lipid content than wild-type after 40 h of silicon starvation. Analyses of fatty acids, lipid classes, and membrane stability in the transgenic strains suggest a role for this enzyme in membrane lipid turnover and lipid homeostasis. These results demonstrate that targeted metabolic manipulations can be used to increase lipid accumulation in eukaryotic microalgae without compromising growth. metabolism | RNAi | algal biofuel | targeted manipulation | triacylglycerol

Two Glycerol-3-Phosphate Dehydrogenases from Chlamydomonas Have Distinct Roles in Lipid Metabolism

“…Some successful approaches have involved the inhibition of lipid catabolism pathways, bypass of carbohydrate synthesis, or modification of transcription factors (Trentacoste et al, 2013;Daboussi et al, 2014;Ngan et al, 2015), while other approaches, such as the overexpression of specific Kennedy pathway components like a DGAT gene, have been less successful in some instances (La Russa et al, 2012). Previous expression of a yeast GPDH in oilseed rape (Vigeolas et al, 2007) and the recent overexpression of a diatom GPDH (Yao et al, 2014) indicate the potential of GPDH manipulation.…”

Exploring Valuable Lipids in Diatoms

“…Two heterologous thioesterase genes were inserted into P. tricornutum and ratios of C12 and C14 fatty acids to total fatty acids increased (Radakovits et al, 2011). T. pseudonana exhibited increased lipid yields from the knockdown of a multi-functional lipase without affecting diatom growth (Trentacoste et al, 2013). Knocking down the genes encoding the decarboxylation enzyme phosphoenolpyruvate carboxykinase (PEPCK) and UDP-glucose pyrophosphorylase (UGPase) resulted in down-regulated decarboxylation and up-regulated lipid synthesis in P. tricornutum (Yang et al, 2016).…”

Nitrate Reductase Knockout Uncouples Nitrate Transport from Nitrate Assimilation and Drives Repartitioning of Carbon Flux in a Model Pennate Diatom

“…In NO 3 2 -limited conditions, diatoms and certain green algae redirect C from storage carbohydrates such as chrysolaminarin and starch to accumulation of neutral lipids, specifically triacylglycerol (TAG) (Rismani-Yazdi et al, 2012;Shifrin and Chisholm, 1981;Yu et al, 2009). Much recent work has focused both on characterizing the genes and metabolic pathways that induce lipid biosynthesis and on identifying routes and regulators that could be harnessed to substantially increase TAG production without negatively affecting growth (Alipanah et al, 2015;Ge et al, 2014;Hildebrand et al, 2012;Levitan et al, 2015;Radakovits et al, 2010;Trentacoste et al, 2013;Yang et al, 2013;Yoon et al, 2012). Deleterious impacts of NO 3 2 depletion include reduction of photosynthetic capacity, C-fixation, and N-assimilation and cessation of growth (Alipanah et al, 2015;Bender et al, 2014;Hockin et al, 2012;Syrett et al, 1986;Turpin, 1991).…”