Regulation of Vacuolar pH of Plant Cells

Mathieu, Yves; Guern, J.; Kurkdjian, Armen; Manigault, P; Manigault, Jeanne; Zielińska, Teresa; Gillet, Brigitte; Belœil, Jean‐Claude; Lallemand, Jean‐Yves

doi:10.1104/pp.89.1.19

Cited by 42 publications

(20 citation statements)

References 40 publications

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“…This ratio is, of course, strongly dependent on the value of the buffering capacity chosen for the calculation. The buffering capacity ,B ofthe vacuolar sap around pH 5.5 calculated from the malate, citrate, and phosphate contents was in good agreement with the one estimated from weak base-induced alkalinizations (22). Extrapolation of these calculations of ,B values in the range pH 5.0 to 5.5 showed that a mean value of 36 ,uEq.…”

supporting

confidence: 78%

“…Successive 31P NMR spectra of a vacuolar preparation incubated in the presence of Mg-ATP. The vacuoles were isolated and collected in a BTP Mes buffer (25 mm, pH 7.5) as described (22). The NMR tube contained 3 x 107 vacuoles in a 12 mL suspension (pHe = 7.03).…”

Section: Introductionmentioning

confidence: 99%

“…as described in the first paper of this series (22). In some experiments, as indicated in the text and figure legends, the isolation procedure was modified as follows.…”

mentioning

confidence: 99%

“…The conditions for 31P NMR measurements have been described in a preceding paper (22). 23Na NMR spectra were obtained at 105.84 MHz with a 20 mm diameter broadband probe.…”

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confidence: 99%

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Regulation of Vacuolar pH of Plant Cells

Guern

Mathieu²,

Kurkdjian³

et al. 1989

Plant Physiol.

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The vacuolar pH and the trans-tonoplast ApH modifications induced by the activity of the two proton pumps H+-ATPase and H -PPase and by the proton exchanges catalyzed by the Na+/H+ and Ca2+/H+ antiports at the tonoplast of isolated intact vacuoles prepared from Catharanthus roseus cells enriched in inorganic phosphate (Y Mathieu et al 1988 Plant Physiol [in press]) were measured using the 31P NMR technique. The H+-ATPase induced an intravacuolar acidification as large as 0.8 pH unit, building a trans-tonoplast ApH up to 2.2 pH units. The hydrolysis of the phosphorylated substrate and the vacuolar acidification were monitored simultaneously to estimate kinetically the apparent stoichiometry between the vectorial proton pumping and the hydrolytic activity of the H+-ATPase. A ratio of H+ translocated/ ATP hydrolyzed of 1.97 ± 0.06 (mean ± standard error) was calculated. Pyrophosphate-treated vacuoles were also acidified to a significant extent. The H+-PPase at 2 millimolar PPi displayed hydrolytic and vectorial activities comparable to those of the HATPase, building a steady state ApH of 2.1 pH units. Vacuoles incubated in the presence of 10 millimolar Na+ were alkalinized by 0.4 to 0.8 pH unit. It has been shown by using 23Na NMR that sodium uptake was coupled to the H+ efflux and occurred against rather large concentration gradients. For the first time, the activity of the Ca2+/H+ antiport has been measured on isolated intact vacuoles. Ca2+ uptake was strongly inhibited by NH4CI or gramicidin. mulation and anion retrieval (6). However, aside the relatively well studied tonoplast proton pumping ATPase (26, 28 and references therein), our knowledge of the other ionic exchanges involving fluxes of protons or proton-equivalents is more limited. A second proton pump, the H+-PPase, present on tonoplast-enriched microsomal vesicles, has been described several times as distinct from the H+-ATPase (12,25,31). Its hydrolytic activity has also been measured on isolated intact vacuoles of beet roots (32) and tulipa petals (31) but, in these few cases, the possible effect of PPi hydrolysis on the vacuolar pH has not been determined.The study of H+-coupled secondary transport of cations through the functioning of Na+/H+ or Ca2+/H' antiports has been restricted to tonoplast vesicles isolated from beet (8, 9) carrot (1 1), or oat (27) Figure 1 illustrates the evolution with time of 31P NMR spectra of a vacuolar suspension incubated with Mg-ATP. The extravacuolar and intravacuolar inorganic phosphate peaks were clearly distinct from the peaks corresponding to the three phosphate groups of ATP. As ATP was hydrolyzed, the two ADP peaks appeared along with a weak AMP signal. The main interest of using 31P NMR was that it allowed simultaneously (a) the measurement of ATP hydrolysis, (b) the measurement ofvacuolar pH modifications linked to ATP hydrolysis, and (c) the estimation of the apparent stoichiometry between vectorial H+-pumping and the hydrolytic activities of the H+-ATPase.The rate of ATP hydrolysis was measured from...

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supporting

confidence: 78%

Section: Introductionmentioning

confidence: 99%

“…as described in the first paper of this series (22). In some experiments, as indicated in the text and figure legends, the isolation procedure was modified as follows.…”

mentioning

confidence: 99%

“…The conditions for 31P NMR measurements have been described in a preceding paper (22). 23Na NMR spectra were obtained at 105.84 MHz with a 20 mm diameter broadband probe.…”

mentioning

confidence: 99%

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Regulation of Vacuolar pH of Plant Cells

Guern

Mathieu²,

Kurkdjian³

et al. 1989

Plant Physiol.

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“…Nevertheless, the NMR spectrum of these cells (Fig. 5A) indicates that the cytoplasmic pH was around 7.5, and the profile of the spectrum for the regions of sugar phosphates and nucleotide phosphates including ATP was normal and quite similar to those reported for other plant materials (9,11,17) as well as to that of the cells in buffered medium (Fig. SB).…”

Section: Possible Disturbing Factorsmentioning

confidence: 84%

Cytoplasmic Acidification Induced by Inorganic Phosphate Uptake in Suspension Cultured Catharanthus roseus Cells

Sakano¹,

Yazaki²,

Mimura³

1992

Plant Physiol.

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Cytoplasmic acidification during inorganic phosphate (Pi) absorption by Catharanthus roseus cells were studied by means of a fluorescent pH indicator, 2',7'-bis-(2-carboxyethyl)-5 carboxyfluorescein (acetomethylester) (BCECF), and 31P-nuclear magnetic resonance spectroscopy. Cytoplasmic acidification measured by decrease in the fluorescence intensity started immediately after Pi application. Within a minute or so, a stable state was attained and no further acidification occurred, whereas Pi absorption was still proceeding. As soon as Pi in the medium was exhausted, cytoplasmic pH started to recover. Coincidentally, the medium pH started to recover toward the original acidic pH. The Pi-induced changes in the cytoplasmic pH were confirmed by 31P-nuclear magnetic resonance study. Maximum acidification of the cytoplasm induced by 1.7 millimolar Pi was 0.2 pH units. Vacuolar pH was also affected by Pi. In some experiments, but not all, pH decreased reversibly by 0.2 to 0.3 pH units during Pi absorption. Results suggest that the cytoplasmic pH is regulated by proton pumps in the plasma membrane and in the tonoplast. In addition, other mechanisms that could consume extra protons in the cytoplasm are suggested.

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Cell Suspensions

Leibfritz¹,

Altenburger²

2007

Encyclopedia of Magnetic Resonance

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Regulation of Vacuolar pH of Plant Cells

Cited by 42 publications

References 40 publications

Regulation of Vacuolar pH of Plant Cells

Regulation of Vacuolar pH of Plant Cells

Cytoplasmic Acidification Induced by Inorganic Phosphate Uptake in Suspension Cultured Catharanthus roseus Cells

Cell Suspensions

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