Carbonic anhydrase activity and inorganic carbon fluxes in low- and high-C1 cells of Chlamydomonas reinhardtü and Scenedesmus obliquus

Carbonic anhydrase (CA) activity associated with high- and low-dissolved inorganic carbon (C₁) grown cells was examined in whole cells by measuring ¹⁸O exchange from doubly labeled CO₂ (¹³C¹⁸O¹⁸O). Both algal species showed the presence of extracellular (periplasmic) as well as intracellular CA activity, which were both greatly increased in low-C₁ cells. The periplasmic CA activity was at least 40-fold higher in lowcompared to high-C₁ cells in both C. reinhardtii and S. obliquus. while low-C₁ cells of S. obliquus showed the highest activity of internal CA. The CA inhibitor ethoxyzolamide showed a strong inhibition of the C₁ uptake process in both C. reinhardtii and S. obliquus as in cyanobacteria. which may indicate that the nature of the primary uptake process is similar in both green algae and cyanobacteria. By using a mass spectrometnc disequilibrium technique it was possible to separate the C₁ fluxes of net HCO^?₃-uptake and net CO₂-uptake during steady-state photosynthesis in high- and Sow-C₁ grown cells of Chlamydomonas reinhardtii (WT. 2137+) and Scenedesmus obliquus (WT. D3). It was found that both high- and low-C₁ cells of the two algae can utilize both CO₂ and HCO^?₃ for photosynthesis, although low-C₁ cells have a higher affinity for the uptake of both C₁ species. Induction at low-C₁ causes an increase in the affinity of both species for HCO^?₃ and CO₂; changes in net CO₂-uptake were, however, significantly greater.     

蓝藻高CO2需求突变株的研究进展

对蓝藻CO2浓缩机制(CO2 concentrating mechanism,CCM)的认识,最终要揭示其作用的生理生化和分子生物学基础,蓝藻高CO2需求突变株的研究为这个目标的实现提供了一条最为有效的途径,1986年,Marcus等第一次把筛选光合作用突变株的方法加以修改,利用化学诱变筛选到第一个单细胞蓝藻Synechococcus PCC7942的高CO2需求突变株,为这一领域的研究在方法学上开辟了一个新的途径。     

绿藻CO2浓缩机制的研究进展

单细胞绿藻是淡水水体中浮游植物的重要组成部分,也是淡水生态系统中主要的初级生产者,其在适应外界CO2浓度变化的过程中,细胞内形成了一种主动转移无机碳的机制－CO2浓缩机制（CO2 concentrating mechanism,CCM）。该机制能使细胞在核酮糖－2－磷酸羧化氧化酶（rubiscol)固碳位点提高CO2浓度,以增加光合作用和减少光吸收。本文综述了这种机制中的无机碳转移模型和不同环境因子（光,温度,CO2浓度和营养水平）对它的调控作用,以期促进深入开展浮游植物对大气CO2浓度升高响应的研究。     

Photosynthetic inorganic carbon uptake and accumulation in two marine diatoms

Some physiological characteristics of photosynthetic inorganic carbon uptake have been examined in the marine diatoms Phaeodactylum tricornutum and Cyclotella sp. Both species demonstrated a high affinity for inorganic carbon in photosynthesis at pH7.5, having K_1/2(CO₂) in the range 1.0 to 4.0mmol m^?3 and O_2? and temperature-insensitive CO₂ compensation concentrations in the range 10.8 to 17.6 cm³ m^?3. Intracellular accumulation of inorganic carbon was found to occur in the light; at an external pH of 7.5 the concentration in P. tricornutum was twice, and that in Cyclotella 3.5 times, the concentration in the suspending medium. Carbonic anhydrase (CA) was detected in intact Cyclotella cells but not in P. tricornutum, although internal CA was detected in both species. The rates of photosynthesis at pH 8.0 of P. tricornutum cells and Cyclotella cells treated with 0.1 mol m^?3 acetazolamide, a CA inhibitor, were 1.5- to 5-fold the rate of CO₂ supply, indicating that both species have the capacity to take up HCO₃^? as a source of substrate for photosynthesis. No Na⁺ dependence for HCO₃^? could be detected in either species. These results indicate that these two marine diatoms have the capacity to accumulate inorganic carbon in the light as a consequence, in part, of the active uptake of bicarbonate.     

The CO2concentrating mechanism in cyanobactiria and microalgae

Over the past 10 years it has become clear that cyanobacteria and microalgae possess mechanisms for actively acquiring inorganic carbon from the external medium and are able to use this to elevate the CO₂ concentration around the active site of the primary photosynthetic carboxylating enzyme, ribulose bisphosphate carboxylase-oxygenase (Rubisco). This results in a vastly enhanced photosynthetic affinity for inorganic carbon (C_i) and improved photosynthetic efficiency. The CO₂ concentrating mechanism is dependent on the existence of membrane bound C_i transport systems, and a microenvironment within the cell where the accumulated C_i can be used to elevate CO₂ at the site of Rubisco. Evidence presented in this review suggests that in cyanobacteria this is achieved by the packaging of Rubisco and carbonic anhydrase (CA) into discrete structures, which are termed carboxysomes. Analogous structures in microalgae, termed pyrenoids, may perform a similar function. The recovery and analysis of high-CO₂-requiring mutants has greatly advanced our understanding of the mechanisms and genes underlying these systems, especially in cyanobacteria, and this review places particular emphasis on the contribution made by molecular genetic approaches.     

Inorganic carbon acquisition in red tide dinoflagellates

Carbon acquisition was investigated in three marine bloom-forming dinollagellates-Prorocentrum minimum, Heterocapsa triquetra and Ceratium lineatum. In vivo activities of extracellular and intracellular carbonic anhydrase (CA), photosynthetic O2 evolution, CO2 and HCO3- uptake rates were measured by membrane inlet mass spectrometry (MIMS) in cells acclimated to low pH (8.0) and high pH (8.5 or 9.1). A second approach used short-term 14C-disequilibrium incubations to estimate the carbon source utilized by the cells. All three species showed negligible extracellular CA (eCA) activity in cells acclimated to low pH and only slightly higher activity when acclimated to high pH. Intracellular CA (iCA) activity was present in all three species, but it increased only in P. minimum with increasing pH. Half-saturation concentrations (K1/2) for photosynthetic O2 evolution were low compared to ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) kinetics. Moreover, apparent affinities for inorganic carbon (Ci) increased with increasing pH in the acclimation, indicating the operation of an efficient CO2 concentration mechanism (CCM) in these dinoflagellates. Rates of CO2 uptake were comparably low and could not support the observed rates of photosynthesis. Consequently, rates of HCO3- uptake were high in the investigated species, contributing more than 80% of the photosynthetic carbon fixation. The affinity for HCO3- and maximum uptake rates increased under higher pH. The strong preference for HCO3- was also confirmed by the 14C-disequilibrium technique. Modes of carbon acquisition were consistent with the 13C-fractionation pattern observed and indicated a strong species-specific difference in leakage. These results suggest that photosynthesis in marine dinoflagellates is not limited by Ci even at high pH, which may occur during red tides in coastal waters.     

Carbon balance during CAM: an assessment of respiratory CO2 recycling in the epiphytic bromeliads Aechmea nudicaulis and Aechmea fendleri

Abstract The regulation of crassulacean acid metabolism (CAM) under controlled environmental conditions has been investigated for two tropical epiphytes, relating plant water and carbon balance to growth form and habitat preference under natural conditions. Aechmea fendleri is restricted to wet, upper montane regions of Trinidad, while A. nudicaulis has a wider distribution extending into more arid regions of the island. Morphological characteristics of these plants are related to habitat preference in terms of leaf succulence (0.44 and 0.94 kg m^?2 for the two species respectively) and a distinct layer of water storage parenchyma in A. nudicaulis In contrast, the thinner leaves of A. fendleri contain little water-storage parenchyma and less chlorenchyma per unit area, but the plants have a more open leaf rosette. The two species differ in expression of CAM, since the proportion of respiratory CO₂ recycled as part of CAM had been found to be much lower in A. fendleri This study compared the efficiency of water use and role of respiratory CO₂ recycling under two PAR regimes (300 and 120 μnol m^?2 s^?1) and three night temperatures (12, 18 and 25 °C). Dark CO₂ uptake rates for both species were comparable to plants in the field (maximum of 2.3 ± 0.2 μmol m^?2s^?1± SD, n= 3). Total net CO₂ uptake at night increased on leaf area basis with temperature for both species under higher PAR, although under the low PAR regime CO₂ uptake was maximal at 18 °C. Water-use efficiency (WUE) increased at 18 °C and 25 °C during dark CO₂ uptake (Phase I) and also during late afternoon photosynthesis (Phase IV) in both species. For A. fendleri, dawn to dusk changes in titrable acidity (ΔH ⁺) were similar under high and low PAR, although ΔH⁺ was correlated to night temperature and PAR in A. nudicaulis. The proportion of ΔH⁺ derived from respiratory CO₂ also varied with experimental conditions. Thus percentage recycling was lower in A. fendleri under high PAR (0–10%), but was only reduced at 18 °C under low PAR. Recycling by A. nudicaulis ranged from 32–42% under high PAR, but was also reduced to 6% under low PAR at 18 °C; at 12 °C and 25 °C, recycling was 37% and 52% respectively. Previous studies have suggested a relationship between the proportion of recycling and degree of water stress. This study indicated that CAM as a CO₂ concentrating mechanism regulates both water-use efficiency and plant carbon balance in these epiphytes, in response to PAR and night temperature. However, the precise relationship between respiratory processes and the balance between external and internal sources of CO₂ is as yet unresolved.     

两株绿藻响应CO2浓度变化的生长和生理特性的研究

以小球藻FACHB-1580和栅藻FACHB-1618为研究对象, 比较了两株绿藻在0.04% CO₂、5% CO₂和20% CO₂ (v/v)三种通气培养条件下的生长和生理特性的响应, 试图阐述与无机碳利用相关生理参数和微藻利用CO₂能力的关系。结果表明, 两株绿藻均能高效利用CO₂, 在5%(v/v)条件下均表现出最大生物量积累、最大比生长速率和最大二氧化碳固定速率。小球藻FACHB-1580和栅藻FACHB-1618最大生物量分别为3.5和5.4 g/L, 分别是0.04% CO₂ (v/v)条件的1.41和1.46倍。在高达20% CO₂ (v/v)条件下, 两株绿藻的生物量均显著高于空气组(P<0.05)。随着CO₂浓度的增加, 两株绿藻的无机碳亲和力、胞内和胞外CA活性、初始Rubisco活性, 及Rubisco活化度均有下降趋势, 总的Rubisco活性变化不明显。另外, 小球藻FACHB-1580存在较高的胞外和胞内CA活性; 而栅藻FACHB-1618胞外CA活性几乎为零, 胞内CA活性显著低于小球藻FACHB-1580。由此推测, 小球藻FACHB-1580能同时吸收介质中的${rm{HCO}}_3^ - $和CO₂, 其胞内CA催化胞内${rm{HCO}}_3^ - $快速转化为CO₂, 从而为Rubisco提供充足的CO₂来源; 而栅藻FACHB-1618主要吸收介质中的CO₂, 其胞内CA活性较低, 推测其通过提高胞内CA含量, 或增强Rubisco对CO₂的亲和力等促进光合固碳作用。