Dynamism of Metabolic Carbon Flow of Starch and Lipids in Chlamydomonas debaryana

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.

Mechanisms of Sodium-Acetate-Induced DHA Accumulation in a DHA-Producing Microalga, Crypthecodinium sp. SUN

“…The starch content of the G group increased dramatically after 24 h cultivation, reaching a maximum (36.79% of DW) at 72 h. As glucose was the direct substrate of starch biosynthesis, it is reasonable that starch was largely accumulated in G group. After 72 h cultivation, the starch content of the glucose group decreased to 35.39% of DW at 96 h. Many studies have shown that the reduction in starch content might be related to lipid accumulation in microalgae [ 9 , 10 , 12 ]. The decline in starch content after 72 h in group G might be due to the carbon flux flow into other storage components [ 13 ].…”

“…Studies have shown that the upregulation of PDH was a key factor stimulating acetyl-CoA production and lipid accumulation in Chlamydomonas sp. JSC4 [ 12 ]. However, the comparative transcriptome analysis showed unexpected results, where the expressions of genes coding three PDH isoenzymes had no significant differences among the three groups in the present study ( Figure 4 ).…”

“…When compared with the wild type (WT), a starch-deficient mutant of Crypthecodinium cohnii had a decrease in starch content but an increase in lipid and DHA contents by 1.12-fold and 1.13-fold, respectively, of that found in WT [ 9 ]. Besides, starch-to-lipid conversion is generally observed in microalgae under stress conditions [ 10 , 11 , 12 , 13 ]. High salinity was shown to promote carbon redistribution and starch conversion to lipid accumulation in Chlorella sorokiniana [ 10 ].…”

Application of ionizing radiation as an elicitor to enhance the growth and metabolic activities in Chlamydomonas reinhardtii

“…The radiation-induced growth stimulation of C. reinhardtii cells may be associated with enhanced metabolic activities, such as photosynthesis, chlorophyll, and protein synthesis. Considering the dynamic metabolic carbon flow that modulates the relative proportions of starch and lipids in a reverse relationship ( Sato and Toyoshima, 2021 ), the simultaneous enhancement of starch and lipid synthesis after X-irradiation with 6 Gy is likely to be attributed to the metabolic alterations associated with growth stimulation.…”

“…The photosynthetically assimilated carbon in Chlamydomonas is partly redirected from a pool of starch to neutral lipid synthesis ( Li et al., 2010 ). Therefore, the relative proportions of starch and lipids can be modulated in a reverse relationship by altering the dynamic metabolic carbon flow ( Sato and Toyoshima, 2021 ). In contrast, novel mixotrophic fed-batch cultivation, including systematic feeding of acetic acid and nutrients, is considered a commercially viable strategy applicable to high-density Chlamydomonas cultures, increasing biomass density, productivity, and the total amount of recombinant proteins ( Fields et al., 2018 ).…”