Chloroplast pH values and buffer capacities in darkened leaves as revealed by CO2 solubilization in vivo

Hauser, Matthias; Eichelmann, Hillar; Heber, U.; Laisk, Agu

doi:10.1007/bf00201374

Cited by 20 publications

(15 citation statements)

References 23 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…This eliminated artifacts by fast intercellular CO 2 diusion between leaf parts outside and inside the cuvette. In this way, we realized that buer capacities measured earlier for chloroplasts in vivo (Hauser et al 1995b) were overestimates. This does not aect the regulatory aspects of the earlier work.…”

Section: Discussionmentioning

confidence: 86%

pH and buffer capacities of apoplastic and cytoplasmic cell compartments in leaves

Oja

Савченко

Jakob

et al. 1999

Planta

Self Cite

View full text Add to dashboard Cite

After opening the stomata in CO(2)-free air, darkened leaves of several plant species were titrated with CO(2) at concentrations between 1 and 16%, in air in order to reversibly decrease cellular pH values and to calculate buffer capacities from pH changes and bicarbonate accumulation using both gas-exchange and fluorescence methods for analysis. After equilibration with CO(2) for times ranging between 4.4 and 300 s, fast CO(2) release from bicarbonate indicated catalysis by highly active carbonic anhydrase. Its time constant was below 2.5 s. Additional CO(2) was released with time constants of about 5, 15 and approximately 300 s. With CO(2) as the acidifying agent, calculated buffer capacities depend on assumptions regarding initial pH in the absence of an acid load. At an initial stroma pH of 7.7, the stromal buffer capacity was about 20 mM pH-unit(-1 )in darkened spinach leaves. At an initial pH of 7.5 it would be only 12 mM pH-unit(-1), i.e. not higher than expected solely on the basis of known stromal concentrations of phosphate and phosphate esters, disregarding the contribution of other solutes. At a concentration of 16%, CO(2) reduced the stromal pH by about 1 pH unit. Buffering of the cytosol was measured by the CO(2)-dependent quenching of the fluorescence of pyranine which was fed to spinach leaves via the petiole. Brief exposures to high CO(2) minimized interference by effective cytosolic pH regulation. Cytosolic buffering appeared to be similar to or only somewhat higher than chloroplast buffering if the initial cytosolic pH was assumed to be 7.25, which is in accord with published cytosolic pH values. The difference from chloroplast pH values indicates the existence of a pH gradient across the chloroplast envelope even in darkened leaves. Apoplastic buffering was weak as measured by the CO(2)-dependent quenching of dextran-conjugated fluorescein isothiocyanate which was infiltrated together with sodium vanadate into potato leaves. In the absence of vanadate, the kinetics of apoplastic fluorescence quenching were more complex than in its presence, indicating fast apoplastic pH regulation which strongly interfered with the determination of apoplastic buffering capacities. At an apoplastic pH of 6.1 in potato leaves, apoplastic buffering as determined by CO(2) titration with and without added buffer was somewhat below 4 mM pH-unit(-1). Thus the apoplastic and cytosolic pH responses to additions of CO(2 )indicated that the observed cytoplasmic pH regulation under acid stress involves pumping of protons from the cytosol into the vacuole of leaf cells, but not into the apoplast.

show abstract

Section: Discussionmentioning

confidence: 86%

pH and buffer capacities of apoplastic and cytoplasmic cell compartments in leaves

Oja

Савченко

Jakob

et al. 1999

Planta

Self Cite

View full text Add to dashboard Cite

show abstract

“…Vol. 108, 1995 (IRGA) method (Oja et al, 1986;Laisk et al, 1989;Wagner et al, 1990;Hauser et al, 1995). In this study, we instead used changes in O, concentration that result when CO, is either taken up from or is added to a gas mixture containing O,.…”

Section: Discussionmentioning

confidence: 99%

“…CO, has already been used as a pH probe in isolated chloroplasts (Werdan et al, 1975). With the development of rapid-response gas-exchange techniques (Oja et al, 1986), it became possible to measure the total amount of dissolved gaseous CO, and bicarbonate and its light-induced changes in the chloroplasts of leaves (Oja et al, 1986;Hauser et al, 1995;Oja and Laisk, 1995a). Results have been confirmed by other techniques with leaf protoplasts (Heineke and Heldt, 1988).…”

mentioning

confidence: 99%

Stimulation by Light of Rapid pH Regulation in the Chloroplast Stroma in Vivo as Indicated by CO2 Solubilization in Leaves

et al. 1995

View full text Add to dashboard Cite

Ju I i us-von-Sachs-l nstitute for B iosciences, U n iversity of W ü rzburg, D-97082 W ü rzbu rg, Germany (M. H ., U. H .);and Department of Plant Physiology, lnstitute of Molecular and Cell Biology, University of Tartu, Riia Street 181, EE2400 Tartu, Estonia (H.E., V.O., A.L.)Leaves of Brassica oleracea, Helianfbus annuus, and Nicofiana rustica were exposed for 20 s to high concentrations of CO,. C O , uptake by the leaf, which was very fast, was measured as a transient increase in the concentration of oxygen. Rapid solubilization of CO, in excess of that which is physically dissolved in aqueous phases is proposed to be caused by bicarbonate formation in the stroma of chloroplasts, which contain carbonic anhydrase. O n this basis, pH values and bicarbonate accumulation in the chloroplast stroma were calculated. Buffer capacities were far higher than expected on the basis of known concentrations in the chloroplast stroma. Moreover, apparent buffer capacities increased with the time of exposure to high CO,, and they were higher when the measurements were performed in the light than i n the dark. During prolonged exposure of leaves to 16% CO,, calculated bicarbonate concentrations i n the chloroplast stroma exceeded 90 mM in the dark and 120 mM in the light. l h e observations are interpreted as indicating that under acid stress protons are rapidly exported from the chloroplasts i n exchange for cations, which are imported. The data are discussed in terms of effective metabolic pH control by ion transport, first across the chloroplast envelope and, then, across the tonoplast of leaf mesophyll cells. l h e direct involvement of the vacuole in the regulation of the chloroplast pH in leaf cells is suggested.

show abstract

“…The volume of an oocyte is B500 nl. The concentrations of free CO 2 and bicarbonate in solutions were calculated using the Henderson-Hasselbalch equation (pH ¼ pK 1 þ log[HCO 3 À ]/[CO 2 ]) 57 . Bicarbonate microinjections were performed 20 min before voltage-clamp experiments.…”

Section: Methodsmentioning

confidence: 99%

A molecular pathway for CO2 response in Arabidopsis guard cells

et al. 2015

View full text Add to dashboard Cite

Increasing carbon dioxide (CO 2 ) levels in the atmosphere have caused global metabolic changes in diverse plant species. CO 2 is not only a carbon donor for photosynthesis but also an environmental signal that regulates stomatal movements and thereby controls plant-water relationships and carbon metabolism. However, the mechanism underlying CO 2 sensing in stomatal guard cells remains unclear. Here we report characterization of Arabidopsis RESISTANT TO HIGH CO 2 (RHC1), a MATE-type transporter that links elevated CO 2 concentration to repression of HT1, a protein kinase that negatively regulates CO 2 -induced stomatal closing. We also show that HT1 phosphorylates and inactivates OST1, a kinase which is essential for the activation of the SLAC1 anion channel and stomatal closing. Combining genetic, biochemical and electrophysiological evidence, we reconstituted the molecular relay from CO 2 to SLAC1 activation, thus establishing a core pathway for CO 2 signalling in plant guard cells.

show abstract

Chloroplast pH values and buffer capacities in darkened leaves as revealed by CO2 solubilization in vivo

Cited by 20 publications

References 23 publications

pH and buffer capacities of apoplastic and cytoplasmic cell compartments in leaves

pH and buffer capacities of apoplastic and cytoplasmic cell compartments in leaves

Stimulation by Light of Rapid pH Regulation in the Chloroplast Stroma in Vivo as Indicated by CO2 Solubilization in Leaves

A molecular pathway for CO2 response in Arabidopsis guard cells

Contact Info

Product

Resources

About