Abstract
Microalgae have the potential to recycle CO2 as starch and triacylglycerol (TAG), which provide alternative source of biofuel and high added-value chemicals. Starch accumulates in the chloroplast, whereas TAG accumulates in the cytoplasmic lipid droplets (LD). Preferential accumulation of starch or TAG may be achieved by switching intracellular metabolic carbon flow, but our knowledge on this control remains limited. Are these two products mutually exclusive? Or, does starch act as a precursor to TAG synthesis, or vice versa? To answer these questions, we analyzed carbon flow in starch and lipids using a stable isotope 13C in Chlamydomonas debaryana NIES-2212, which accumulates, without nutrient limitation, starch in the exponential growth phase and TAG in the stationary phase. Pulse labeling experiments as well as pulse labeling and chase experiments were conducted, and then, gas chromatography-mass spectrometry (GC-MS) analysis was performed on starch-derived glucose and lipid-bound fatty acids. We exploited the previously developed method of isotopomer analysis to estimate the proportion of various pools with different isotopic abundance. Starch turned over rapidly to provide carbon for the synthesis of fatty acids in the exponential phase cells. Most fatty acids showed rapid and slow components of metabolism, whereas oleic acid decayed according to a single exponential curve. Highly labeled population of fatty acids that accumulated during the initial labeling decreased rapidly, and replaced by low abundance population during the chase time, indicating that highly labeled fatty acids were degraded and the resulting carbons were re-used in the re-synthesis with about 9-fold unlabeled, newly fixed carbons. Elongation of C16–C18 acids in vivo was indicated by partially labeled C18 acids. The accumulation of TAG in the stationary growth phase was accounted for by both de novo synthesis and remodeling of membrane lipids. These results suggest that de novo synthesis of starch and TAG was rapid and transient, and also almost independent to each other, but there is a pool of starch quickly turning over for the synthesis of fatty acids. Fatty acids were also subject to re-synthesis. Evidence was also provided for remodeling of lipids, namely, re-use of acyl groups in polar lipids for TAG synthesis.
Highlights
Photosynthetic organisms, such as plants, algae, cyanobacteria and anoxygenic bacteria, recycle CO2 under the sunlight to provide organic matters for the biosphere and human life
The overall picture of the complex metabolic pathway leading to TAG synthesis was elucidated in plants (Lung and Weselake, 2006; Li-Beisson et al, 2013; Bates, 2016), but this might not be adapted to microalgae, because microalgae are unicellular organisms that accumulate both starch and TAG within a single cell, whereas the sink and source organs are separated in plants
TAG is typically accumulated as lipid droplets (LD) in the cytosol of developing seeds, LD is supposed to emerge by budding from the endoplasmic reticulum (ER)
Summary
Photosynthetic organisms, such as plants, algae, cyanobacteria and anoxygenic bacteria, recycle CO2 under the sunlight to provide organic matters for the biosphere and human life. Plant and microalgal oil is mainly triacylglycerol (TAG) consisting of long chain fatty acids. TAG is synthesized by two major pathways (Li-Beisson et al, 2013): namely, the acylation of glycerol-3-phosphate with acyl-CoA in the de novo pathway called Kennedy pathway, and the remodeling pathway in which phosphatidylcholine (PC) acts as an intermediate that provides fatty acids and/or diacylglycerol (DAG) for the synthesis of TAG (Bates et al, 2013). LD is typically a spherical globule of 0.1–1 μm in diameter, mainly consisting of TAG and surrounded by a half unit membrane containing oleosin and other specific proteins (Chapman et al, 2012). TAG is known to be present as a minor component of plastoglobules, which are small lipid droplets present in chloroplasts, consisting of plastoquinone, tocopherol, and phytyl esters (Bréhélin et al, 2007; Lohscheider and Bártulos, 2016)
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