The inhibition of TOR in the model diatom Phaeodactylum tricornutum promotes a get-fat growth regime

Abstract

The target of rapamycin (TOR) signaling pathway regulates fundamental intracellular functions critical for cell viability and proliferation. Manipulation of TOR in high lipid-producing microalgae may help overcome the trade-off between biomass production and lipid yield that still impairs the viable production of biofuel from microalgae. In this study, we inhibited the TOR kinase in the model diatom Phaeodactylum tricornutum using the selective TOR inhibitor AZD-8055, and analyzed cell proliferation, chlorophyll content, lipid synthesis and carbon metabolism. AZD-8055 inhibits cell proliferation in a dose-dependent manner compared to N deprivation which stops growth. Microscopy, flow cytometry, and quantitative analyses of lipids also demonstrated that AZD-8055 treatment strongly promotes triacylglycerol (TAG) accumulation while decreasing the quantity of sterols. The TAG productivity of AZD-8055 treated cultures was significantly higher than for N deprived cultures. Measurement of the activities of the key metabolic enzymes glyceraldehyde phosphate dehydrogenase (GAPDH), glucose-6-phosphate dehydrogenase (G6PDH) and malate dehydrogenase (MDH) revealed opposite effects for AZD-8055 treatment and N-starvation on the activity of the glycolytic enzyme GAPDH. This suggests that TOR inhibition and N starvation may have distinct impacts on general metabolism and lipid accumulation. Our main finding is that treating cultures with AZD-8055 results in higher TAG productivity than N starvation in P. tricornutum. The chemical or genetic manipulation of the TOR signaling pathway in P. tricornutum and other diatoms may lead to the development of strains or approaches suitable for the enhanced production of TAGs for biofuel

Engineering the unicellular alga Phaeodactylum tricornutum for high‐value plant triterpenoid production

“…Substantial advances will need to be made for algae with regard to both gene expression control and cultivation before this platform could become commercially feasible for the production of small molecules. To date, the regulation of the lipid and sterol metabolism of diatoms is largely unknown; a recent study has shown that sterol biosynthesis can be chemically induced in a lipid‐independent manner (Prioretti et al ., ). This study, therefore, represents pioneering work validating the potential of these eukaryotic microalgae as a chassis for the photo production of value‐added molecules.…”

ppGpp influences protein protection, growth and photosynthesis inPhaeodactylum tricornutum

“…The fluorescent probe Nile red was used to detect neutral lipids by flow cytometry as specified in (Prioretti et al , 2017). The cells at different growth phases were fixed with 2% (v/v) glutaraldehyde (Prioretti et al , 2017) and 1 ml of cell suspension was incubated with 2 μl of Nile red solution (0.25 mg ml −1 ) for 5 minutes before the analysis with a bench flow cytometer (BD Accuri C6, BD Biosciences). To determine Nile red fluorescence, the trigger signal was set to FL2 fluorescence and combined with the side scatter (SSC) signal.…”

“…Like plants, diatoms possess signalling pathways with prokaryotic origins such as the bacterial histidine-kinase-based two-component systems (Bowler et al , 2008), as well as pathways of eukaryotic origins such as the Target Of Rapamycin (TOR) kinase (Prioretti et al , 2017; Prioretti et al , 2019) and the G protein-coupled receptor signalling pathway (Port et al 2013).…”

Storage Compound Accumulation in Diatoms as Response to Elevated CO2 Concentration

“…In Chaetoceros gracilis, a combined mild Biology 2020, 9, 5 2 of 17 deprivation of both silicon and sodium chloride produced a lipid accumulation reaching up to 73% of the total cell dry weight [10]. Other approaches to enhance lipid production in diatoms involve the use of phytohormones and other hormone-derived compounds [19,20], as well as the pharmacological modulation of signaling pathways involved in the synthesis of lipids [20,21]. In addition, thanks to the new molecular engineering approaches in diatoms, it is also possible to enhance lipid accumulation in these organisms by genetic transformation.…”