Photosynthesis and Inorganic Carbon Transport in Isolated Asparagus Mesophyll Cells

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The species of inorganic carbon (CO2 or HCO3-) taken up a a source of substrate for photosynthetic fixation by isolated Asparagus qprengeri mesophyll ceUls is investigated. Discrimination between CO2 or HC03-transport, during steady state photosynthesis, is achieved by monitoring the changes (by "C fixation) which occur in the specific activity of the intracellular pool of inorganic carbon when the inorganic carbon present in the suspending medium is in a state of isotopic disequilibrium. Quantitative comparisons between theoretical (CO2 or HCO3-transport) and experimental time-courses of "C incorporation, over the pH range of 5.2 to 7.5, indicate that the specific activity of extracellular C02, rather than HCO3, is the appropriate predictor of the intracellular specific activity.It is concluded, therefore, that CO2 is the major source of exogenous inorganic carbon taken up by Asparagus cells. However, at high pH (8.5), a component of net DIC uptake may be attributable to HCO3-transport, as the incorporation of "C during isotopic disequilibrium exceeds the maximum possible incorporation predicted on the basis of CO2 uptake alone. The contribution of HCO3-to net inorganic carbon uptake (pH 8.5) is variable, ranging from 5 to 16%, but is independent of the extracellular HCO3-concentration. The evidence for direct HC03-transport is subject to alternative explanations and must, therefore, be regarded as equivocal. Nonlinear regression analysis of the rate of "C incorporation as a function of time indicates the presence of a smaUl extracellular resistance to the diffusion of C02, which is partially alleviated by a high extracellular concentration of HC03-.It is now apparent that there are a number of specialized mechanisms by which plant cells acquire DIC4 from their surroundings for photosynthetic fixation (15) and that the various means of acquisition dramatically affect the efficiency of photosynthetic C assimilation (3-5, 7, 13). In cyanobacteria, for example, HC03-is actively transported across the plasmalemma and accumulated within the cells (2,13,16

contrasting

confidence: 62%

Inorganic Carbon Uptake during Photosynthesis

Espie¹,

Owttrim²,

Colman³

1986

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“…Other environmental stimuli have also been shown to influence the cytoplasmic pH. Light was shown, using 14C labeled weak acids, to increase the cytoplasmic pH in Asparagus mesophyll cells (10). In maize roots, hypotoxia resulted in the reduction of cytoplasmic pH by 0.3 to 0.6 units (20).…”

Section: Resultsmentioning

confidence: 99%

³¹P NMR Study of Elicitor Treated Phaseolus vulgaris Cell Suspension Cultures

Ojalvo

Rokem²,

Navon³

et al. 1987

ABSTRACIThe addition of an elicitor (glucan) to Phaseolus vulgaris cell suspension cultures increased the formation of the phytoalexin phaseollin. Intracellular pH and phosphate concentrations were studied with 31P nuclear magnetic resonance spectroscopy on elicitor-treated cells which were aerated during the nuclear magnetic resonance measurement. The pH of the vacuole and to a lesser extent the pH of the cytoplasm were affected at 10 minutes after elicitor addition; a decrease in pH from 53 to 4.8 was noted in the vacuole and from 7A6 to 7.28 in the cytoplasm. The ratio between the amount of Pi in the vacuole to that in the cytoplasm also changed within 10 minutes after elicitor addition. The signal for ATP (8-ATP) was low after elicitor addition and was high again 23 hours after elicitation. tions and pH observed in plant cells after elicitor addition (24). In that study the plant cells were not aerated during the NMR measurements. It was shown that the amounts of inorganic phosphate in the vacuole increased at the expense of the phosphate in the cytoplasm of P. hortense cells after the addition of fungal cell wall fragments. It was briefly mentioned that the vacuolar pH decreased from 5.5 to 5.1 within 1 h after elicitor addition.The present study was carried out to establish more definitely the changes in cytoplasmic and vacuolar pH after the addition of an elicitor to P. vulgaris cell suspension cultures. These cells were studied under aerated conditions in the NMR tube. MATERIALS AND METHODSGrowth Conditions. Cells of Phaseolus vulgaris strain BRTIL-WAX (commercial) were maintained in the dark on SH medium (23) containing: 2,4-D, 0.5 mg/L; p-chlorophenoxyacetic acid, 2 mg/L; kinetin, 0.1 mg/L; nitrilotriacetic acid, 40 mg/L; and enzymatic casein hydrolyzate (Sigma, Cat. No. C-0626, St. Louis, MO), 2 g/L. An inoculum (10% by volume) of homogeneous suspension was transferred every 5 d to fresh medium. For 31P NMR experiments cells were transferred to SH medium without auxins (2,4-D and pCPA) and without the following medium components: Na7EDTA, nitrilotriacetic acid, FeSO4. 7H20, CoC12* 6H20, Na2MoO4 * 2H20, CuS04. 6H20, ZnSO4* 7H20, H3BO3, and MnSO444H20. Sodium chloride (20 mg/L) was added to this medium. All experiments were performed in 250 ml cotton plugged Erlenmeyer flasks with 100 ml medium. Growth was in a rotary shaker (G-25

“…6). A theoretical prediction of this disequilibrium data can be obtained using the models of Espie and Colman (9,10) (21,22).…”

mentioning

confidence: 99%

“…Inorganic carbon accumulation was determined using the silicone fluid centrifugation technique as described elsewhere (5,9,14). Terminating solution (100 MAl of2 N NaOH in 20% methanol) was placed in the bottom of a 400 ,l microfuge tube and layered with 100 ,l of silicone fluid (Ar2O:Ar200 = 13.2:5, Wacker Chemical Co; FDR).…”

mentioning

confidence: 99%

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The Role of External Carbonic Anhydrase in Inorganic Carbon Acquisition by Chlamydomonas reinhardii at Alkaline pH

Williams¹,

Turpin²

1987

The role of external carbonic anhydrase in inorganic carbon acquisition and photosynthesis by Chiamydomonas reinhardii at alkaline pH (8.0) was studied. Acetazolamide (50 micromolar) completely inhibited external carbonic anhydrase (CA) activity as determined from isotopic disequilibrium experiments. Under these conditions, photosynthetic rates at low dissolved inorganic carbon (DIC) were far greater than could be maintained by CO2 supplied from the spontaneous dehydration of HC03-thereby showing that C. reinhardii has the ability to utilize exogenous HCO3-. Acetazolamide increased the concentration of DIC required to half-saturate photosynthesis from 38 to 80 micromolar, while it did not affect the maximum photosynthetic rate. External CA activity was also removed from the cell-wall-less mutant (CW-15) by washing. This had no effect on the photosynthetic kinetics of the algae while the addition of acetazolamide to washed cells (CW-15) increased the K?'c from 38 to 80 micromolar. Acetazolamide also caused a buildup of the inorganic carbon pool upon NaHCO3 addition, indicating that this compound partially inhibited internal CA activity. The effects of acetazolamide on the photosynthetic kinetics of C. reinhardii are likely due to the inhibition of internal rather than a consequence of the inhibition of external CA.Further analysis of the isotopic disequilibrium experiments at saturating concentration of DIC provided evidence consistent with active CO2 transport by C. reinhardii. The observation that C. reinhardii has the ability to take up both CO2 and bicarbonate throws into question the role of external CA in the accumulation of DIC in this alga.of carbon for photosynthesis (23). Findenegg (12) (26) then the function of external CA is called into question.Internal CA appears to facilitate the CO2 supply to Rubisco from the internal HCO3 pool (24, 25). Spalding et al. (24,25) showed that a mutant of C. reinhardii lacking internal CA exhibited high rates of photorespiration and accumulated large concentrations of intracellular inorganic carbon. Their conclusion was that an internal CA was required to facilitate CO2 supply to Rubisco from actively transported HCO3 . Proponents of the theory that CO2 is the only species of carbon to cross the plasmalemma suggest that active transport ofbicarbonate occurs at the chloroplast envelope and that the internal CA is located within the chloroplast (21,22).In this report we provide evidence that C. reinhardii has the ability to take up either CO2 or bicarbonate and that the magnitude of each flux is dependent upon the experimental conditions. Our results suggest that both CO2 and HCO3 accumulation are active and occur at the plasmalemma. We also provide evidence that external CA is not necessary for efficient photosynthesis at alkaline pH.