Songcui Wu12？, Aiyou Huang1？, Baoyu Zhang1, Li Huan12, Peipei Zhao12, Apeng Lin1and Guangce Wang1，Corresponding author: Guangce Wang gcwang@qdio.ac.cn
Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Nanhai Road 7, Qingdao 266071, China
College of Earth Sciences, University of Chinese Academy of Science, Beijing 100049, China
Songcui Wu and Aiyou Huang contributed equally to this work.
The electronic version of this article is the complete one and can be found online at:http://www.biotechnologyforbiofuels.com/content/8/1/78
| Received: | 26 June 2014 |
| Accepted: | 20 May 2015 |
| Published: | 28 May 2015 |
Rising CO2 concentration was reported to increase phytoplankton growth rate as well as lipid productivity. This has raised questions regarding the NADPH supply for high lipid synthesis as well as rapid growth of algal cells.
In this study, growth, lipid content, photosynthetic performance, the activity, and expression of key enzymes in Calvin cycle and oxidative pentose phosphate pathway (OPPP) were analyzed in the marine diatom Phaeodactylum tricornutum under three different CO2 concentrations (low CO2(0.015 %), mid CO2 (atmospheric, 0.035 %) and high CO2 (0.15 %)). Both the growth rate and lipid content of P. tricornutum increased significantly under the high CO2 concentration. Enzyme activity and mRNA expression of three Calvin cycle-related enzymes (Rubisco, 3-phosphoglyceric phosphokinase (PGK), phosphoribulokinase (PRK)) were also increased under high CO2 cultivation, which suggested the enhancement of Calvin cycle activity. This may account for the observed rapid growth rate. In addition, high activity and mRNA expression of G6PDH and 6PGDH, which produce NADPH through OPPP, were observed in high CO2 cultured cells. These results indicate OPPP was enhanced and might play an important role in lipid synthesis under high CO2 concentration.
The oxidative pentose phosphate pathway may participate in the lipid accumulation in rapid-growthP. tricornutum cells in high CO2 concentration.
As a result of increased industrialization and human activities, global carbon dioxide (CO2) emission has increased dramatically and induces seawater acidification, which affects the growth and photosynthesis of marine phytoplankton [1]. Photosynthetic CO2 fixation by phytoplankton (e.g., eukaryotic microalga) mainly depends upon Calvin cycle, and ribulose-1, 5-bisphosphate carboxylase/oxygenase (Rubisco) catalyzes the initial step of CO2 fixing. The response of photosynthesis, metabolite including the Calvin cycle enzymes (especially Rubisco), and growth to elevated CO2 have been well studied in higher plants. It was shown that short-term CO2 elevating generally accelerates carbon fixation and leads to an increase of Rubisco activity followed by an enhancement of growth in C3 plants [2]–[6]. Yet, in the long-term, the photosynthesis decreased with increasing CO2, which is typically accompanied by a decline in the amount and activity of Rubisco and other enzymes in the Calvin cycle and a decrease of growth rates. Compared to high plants, the impacts of elevated CO2 concentration on microalgae and their response to CO2 levels, especially Calvin cycle enzyme (including Rubisco) activity and amount, have been learned much less extent and attentions mainly paid to photosynthesis and algal growth. Only in the study of Euglena gracilis Z, Nakano et al. focused on Rubisco and found Rubisco activity was higher in high CO2conditions [7]. Whereas, elevated CO2 concentration increased the efficiency of photosynthetic carbon fixation and growth of phytoplankton was gradually known as general phenomenon [8], [9]. Kim et al.[10] reported that the growth of Skeletonema costatum enhanced at higher concentrations of CO2. Tortell et al. [11] also found that rising CO2 can enhance Chaetoceros spp. growth. In addition, various studies have shown that rising CO2 concentration increases lipid productivity as well as phytoplankton growth rate, such as in Phaeodactylum tricornutum[12],Nannochloropsis oculata[13], and Chlorella vulgaris[14], high levels of CO2 concentration enhanced both biomass production and lipid content, thus shedding light on the potential for biodiesel production from microalgae. To select microalgae for obtaining a higher lipid productivity, even higher concentrations of CO2 (10 % CO2 and flue gas) were used to cultivate Botryococcus brauniiand Scenedesmus sp. [15]. At present, about 60 species of microalgae have been well domesticated with high concentration of CO2 for producing large biomass and achieving high biofuel yields [16]. However, most previous studies have only focused on microalgal growth rate, lipid content, and tolerance to high levels of CO2[17]–[20]. Little effort has been directed toward the analysis of the mechanism involved in lipid accumulation in microalgae and their simultaneous rapid growth rate.
Microalgal lipids, as a source of biofuel, are usually derived from long-chain fatty acids, which require NADPH for synthesis [21], [22]. For example, to produce an 18-carbon fatty acid, 16 NADPH molecules are required as electron donors. Therefore, enhanced lipid accumulation will surely increase metabolic demand in microalgae for NADPH. Microalgae have been demonstrated to grow rapidly in high CO2 concentrations. This suggests that quantity of NADPH, which is supplied by the light reaction, is required for photosynthetic carbon fixation which supplies substrates and energy for the synthesis of major constituents (proteins, nucleic acids, and carbohydrates) essential for algal growth. For effective CO2 fixation, ATP and NADPH produced by photosynthetic light reactions must be maintained at a molar ratio of 3:2 [23]. Once a large number of NADPH molecules are consumed, the ratio will be disrupted leading to a reduction in carbon fixation activity. An important question therefore remains about how NADPH is supplied for high fatty acid synthesis as well as rapid growth of algal cells cultured under high CO2 concentration. It is more likely that another pathway may contribute to providing this reductant.
In the present study, we evaluated the lipid content in the diatom P. tricornutum which was cultivated in three different CO2 conditions (0.015 %, atmospheric, and 0.15 %). Furthermore, we measured the activity of seven key enzymes and mRNA expression in P. tricornutum to explore the mechanism of rapid growth and the simultaneous increase in lipid accumulation in high-CO2 cultured algal cells. Our research showed that the pentose phosphate pathway may be incorporated in maintaining the NADPH supply under high CO2 concentrations.