Active CO₂ Transport by the Green Alga Chlamydomonas reinhardtii

Abstract

Mass spectrometric measurements of dissolved free 13CO2 were used to monitor CO2 uptake by air grown (low CO2) cells and protoplasts from the green alga Chlamydomonas reinhardtli. In the presence of 50 micromolar dissolved inorganic carbon and light, protoplasts which had been washed free of extemal carbonic anhydrase reduced the 13CO2 concentration in the medium to close to zero. Similar resuits were obtained with low CO2 cells treated with 50 micromolar acetazolamide. Addition of carbonic anhydrase to protoplasts after the period of rapid CO2 uptake revealed that the removal of CO2 from the medium in the light was due to selective and active CO2 transport rather than uptake of total dissolved inorganic carbon. demonstrated for these organisms (2, 4, 9, 15). In the case of cyanobacteria, both HCO3-and CO2 are substrates for active transport (2,3,6,7,14,15) with CO2 being selectively and preferentially used by the cells (2,6,16). In Chlamydomonas, HC03-is actively transported (4, 25, 29), but CO2 uptake has been considered to be passive (18,20). Carbon dioxide, however, is taken up from the medium faster than HCO3-by Chlamydomonas (13,28,29) and several authors (13,29) have considered the possibility of active CO2 transport.Studies on the DIC transport mechanism of green algae are complicated by their cellular compartmentation. Recently, it was shown that isolated chloroplasts of low C02 Chlamydomonas reinhardtii were able to accumulate DIC (19) and a model was presented where the only active DIC transport mechanism was located on the chloroplast envelope (18,19). In that model the plasma membrane was suggested to be only a diffusion barrier for CO2 generated by external carbonic anhydrase. In contrast, by comparison of the apparent affinities for DIC of whole cells and purified chloroplasts, Suiltemeyer et al. (26) came to the conclusion that active transport by the chloroplast alone may not be responsible for the photosynthetic characteristics of whole cells.Another difficulty in examining the DIC species taken up by whole cells is the presence of an external carbonic anhydrase (10) which catalyzes the rapid equilibrium between CO2 and HCO3-, thus making a direct discrimination between CO2 and HCO3-uptake impossible (7, 13). However, using inhibitors for external carbonic anhydrase or the cell-wall less mutant CW-15, some authors came to the conclusion that CO2 and not HCO3-(13) or that both C02 and HCO3- (29) were actively transported.Confusion about which DIC species is actively taken up from the medium may also be caused by methods which only measure total rates of transport rather than transport of CO2 or HC03-individually. Using MS, which measures free dissolved gases in liquid, several authors presented direct eviGreen algae and cyanobacteria possess a high apparent affinity for DIC3 when grown at low DIC concentrations (low CO2 cells: 2,5,9,17), and DIC accumulation has been '

Carbon dioxide availability, intracellular pH and growth rate of the coccolithophore Emiliania huxleyi

“…When external pH was buffered at pH 8.3 and CO2 levels increased from 0.03 to 0.1 % (v/v) C02/air, pH, and growth rate decreased while at 0.5 % (v/v) C02/air high-calcifying cells failed to grow, in part because of the decrease in pHi. Many species of marine phytoplankton restrict intracellular pH to between 7.0 and 7.4 by ion transport mechanisms and a high buffering capacity of the cytosol and these show enhanced growth at elevated CO2 concentrations (Beardall & Raven 1981, Gehl & Colrnan 1985, Sultemeyer et al 1989. The inhibition of growth of the high-calcifying cells of E. huxleyi at elevated CO2 may be related to the inability to generate adequate OH-to neutralize the protons produced by calcification and to counteract the acidification of the cytosol resulting from the passive diffusion of CO2 at elevated ambient concentrations.…”

Gross and net primary production: closing the gap between concepts and measurements

“…First, CO 2 fixation by Rubisco can increase internal pH within the chloroplast stroma (in eukaryotes) or in the cytosol (in Cyanobacteria) by removing CO 2 . Many species of marine microalgae show enhanced growth at elevated CO 2 concentrations (Gehl & Colman 1985, Sültemeyer et al 1989. However, increasing CO 2 leads to a decline in pH.…”

Light-Induced Carbonic Anhydrase Expression in Chlamydomonas reinhardtii

“…The periplasmic CA has been shown to be important in the supply to the cells of free CO2 from the dehydration of HCO3- (1,25), and has been identified to be a glycoprotein with a subunit molecular mass estimated at 35 to 37 kD (6,29,32). The induction of CA activity under low-CO2 conditions was shown to be inhibited by the protein translation inhibitor, cycloheximide (22,24,32), suggesting that the enzyme was synthesized de novo when the CO2 concentration was lowered.…”