pH In Plant Cells Research Articles

Genome-wide characterization of the CPA gene family in potato and a preliminary functional analysis of its role in NaCl tolerance.

The cation/proton antiporter (CPA) superfamily plays a crucial role in regulating ion homeostasis and pH in plant cells, contributing to stress resistance. However, in potato (Solanum tuberosum L.), systematic identification and analysis of CPA genes are lacking. A total of 33 StCPA members were identified and classified into StNHX (n = 7), StKEA (n = 6), and StCHX (n = 20) subfamilies. StCHX owned the highest number of conserved motifs, followed by StKEA and StNHX. The StNHX and StKEA subfamilies owned more exons than StCHX. NaCl stress induced the differentially expression of 19 genes in roots or leaves, among which StCHX14 and StCHX16 were specifically induced in leaves, while StCHX2 and StCHX19 were specifically expressed in the roots. A total of 11 strongly responded genes were further verified by qPCR. Six CPA family members, StNHX1, StNHX2, StNHX3, StNHX5, StNHX6 and StCHX19, were proved to transport Na+ through yeast complementation experiments. This study provides comprehensive insights into StCPAs and their response to NaCl stress, facilitating further functional characterization.

Role of Ionic Strength and pH in Modulating Thermodynamic Profiles Associated with CO Escape from Rice Nonsymbiotic Hemoglobin 1.

Type 1 nonsymbiotic hemoglobins are found in a wide variety of land plants and exhibit very high affinities for exogenous gaseous ligands. These proteins are presumed to have a role in protecting plant cells from oxidative stress under etiolated/hypoxic conditions through NO dioxygenase activity. In this study we have employed photoacoustic calorimetry, time-resolved absorption spectroscopy, and classical molecular dynamics simulations in order to elucidate thermodynamics, kinetics, and ligand migration pathways upon CO photodissociation from WT and a H73L mutant of type 1 nonsymbiotic hemoglobin from Oryza sativa (rice). We observe a temperature dependence of the resolved thermodynamic parameters for CO photodissociation from CO-rHb1 which we attribute to temperature dependent formation of a network of electrostatic interactions in the vicinity of the heme propionate groups. We also observe slower ligand escape from the protein matrix under mildly acidic conditions in both the WT and H73L mutant (τ = 134 ± 19 and 90 ± 15 ns). Visualization of transient hydrophobic channels within our classical molecular dynamics trajectories allows us to attribute this phenomenon to a change in the ligand migration pathway which occurs upon protonation of the distal His73, His117, and His152. Protonation of these residues may be relevant to the functioning of the protein in vivo given that etiolation/hypoxia can cause a decrease in intracellular pH in plant cells.

Improved cytoplasmic pH measurements in SNARF-1 stained plant cells by image processing

Cytoplasmic pH has long been considered to act as a secondary messenger of various cellular responses by affecting the ionization state of proteins.1 In plant biology, cytoplasmic pH has traditionally been measured, especially in guard cells and as a response to plant microorganism interactions, with pH-sensitive microelectrodes.2 More recently, the development of fluorescent pH markers, such as BCECF and SNARF-1, has allowed us to monitor cytoplasmic pH without the need for electrophysiological equipment. However, because of vacuolar structures that occupy a large volume of plant cells, simple measurements of fluorescent intensities are insufficient to provide precise cytoplasmic pH values. In this addendum, we describe our improved method to monitor cytoplasmic pH in plant cells stained by SNARF-1 by image processing using a noise-reducing filter after determination of an optimal ROI size. In addition, further developments for automated region extraction are proposed.

Activity of biochemical pH-stat enzymes in cereal root tips under oxygen deficiency

The activity of the enzymes of alcoholic and lactic-acid fermentation: pyruvate decarboxylase (PDC, EC 4.1.1.1), alcohol dehydrogenase (ADH, EC 1.1.1.1), lactate dehydrogenase (LDH, EC 1.1.1.27) and the enzymes of malic acid metabolism: phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.23), NAD-dependent malate dehydrogenase (NAD-MDH, EC 1.1.1.37), and NADP-dependent malic enzyme (NADP-ME, EC 1.1.1.40) involved in the operation of biochemical pH-stat was investigated in the root tips of wheat (Triticum aestivum L.) and rice (Oryza sativa L.) under hypoxia and anoxia. Exposures lasted for 6, 12, and 18 h. The most pronounced response was detected for the enzymes of alcoholic fermentation. The activation of ADH and PDC in wheat occurred only under hypoxia, whereas in rice it was detected both under hypoxia and anoxia. The activation of LDH in wheat occurred under hypoxia, and in rice, the activity of this enzyme was slightly enhanced. The activity of the enzymes of malic acid metabolism did not change except in wheat root tips under hypoxia when PEPC activity decreased and NADP-ME activity simultaneously rose. The role of biochemical pH-stat in the regulation of cytoplasmic pH in plant cells under oxygen deficit and the mechanisms for regulating the activities of enzymes involved in biochemical pH-stat are discussed as well as the interaction between biochemical pH-stat and other mechanisms maintaining pH of plant cells. The results are analyzed within a context of intracellular pH regulation.

Effects of Anoxia on Amino Acid Levels in Rice Coleoptiles

Effects of anoxia on the levels of free-amino acids were investigated in the coleoptiles of rice (Oryza sativa L.) seedlings. Rice coleoptiles are able to grow in extremely low oxygen conditions. Anoxic stress increased the concentration of total free-amino acids in the coleoptiles. Alanine (Ala) and γ-aminobutyric acid (Gaba) were the main amino acids accumulated. After 48 h, Ala and Gaba concentrations in anoxic coleoptiles were 3.4- and 11.2-fold greater than those in non-stressed coleoptiles, respectively. Ala and Gaba represented 19 and 23 % of the amino acid pool in anoxic coleoptiles, respectively. Submergence stress also increased Ala and Gaba concentrations in rice coleoptiles. Since Ala and Gaba are bio-compatible solutes and their accumulation is known to stabilize osmotic potential and/or cytoplasmic pH in plant cells, these stress-induced amino acids may allow rice coleoptiles to make biochemical adjustment that enable them to cope with the stress conditions. Therefore, the ability to increase the concentrations of Ala and Gaba may be important for anoxic and submergence stress tolerance of rice seedlings.

Metabolic regulation of pH in plant cells: Role of cytoplasmic pH in defense reaction and secondary metabolism

A new biochemical pH-stat hypothesis that revised the classic hypothesis is presented to understand the metabolic regulation of intracellular pH in plant cells. Alternative pathway glycolysis, alternative pathway respiration and malate-derived lactic and alcoholic fermentation (alternative pathway fermentation), all unique to plants, are integrated into a regulatory mechanism of pH in the cytoplasm. Its uniqueness to plant kingdom is discussed from the evolutionary viewpoint: it is suggested that when the ancestors of extant terrestrial plants expanded their habitat from oceans to freshwater, they abandoned a "sodium system" and adopted a "proton system" for nutrient uptake. Validity of the new hypothesis is examined with available data on a secondary active transport, anoxia and other experimental evidence. The hypothesis predicts that biotic and abiotic stress-induced cytoplasmic acidification triggers synthesis of phytoalexins and other secondary metabolites. Possible roles of cyanide-resistant alternative pathway respiration in the secondary metabolite production, metabolic switching between primary and secondary metabolisms, and defense reactions are proposed.

Effects of auxin and abscisic acid on cytosolic calcium and pH in plant cells.

Dark-grown corn coleoptiles and parsley hypocotyls and their roots were loaded with acetoxymethyl esterified forms of the Ca2+ indicator fluo-3, and the pH indicator 2',7'-bis (2-carboxyethyl)-5(and-6)-carboxyfluorescein. These tissues were treated with the plant growth regulator 2,4-dichlorophenoxyacetic acid (2,4-D), an auxin analogue, or abscisic acid (ABA), and the cytosolic pH (pHcyt) and cytosolic Ca2+ ([Ca2+]cyt) changes were monitored by confocal scanning optical microscopy. Over a period of 4 min pHcyt decreased 0.1-0.2 pH unit and [Ca2+]cyt increased from 280 to 380 nM in response to 2,4-D. ABS, on the other hand, induced cytosolic alkalinization of 0.05-0.1 pH unit with a concomitant increase in [Ca2+]cyt from 240 to 320 nM over a 4-min period. Responses similar to these were observed in all the tissues tested. We suggest that pHcyt profoundly influences signaling by[Ca2+]cyt, possibly by regulating Ca2+-protein binding, and that the divergent effects of auxin and ABA on pHcyt underlie their mutual antagonism.

A hydrogen ion-selective liquid-membrane microelectrode for measurement of the vacuolar pH of plant cells in suspension culture

H +-selective microelectrodes were made according to the method described by D. Ammann, F. Lanter, R. A. Steiner, P. Schulthess, Y. Shijo, and W. Simon (1981, Anal. Chem. 53, 2267–2269). Some practical aspects of the preparation and use of these microelectrodes for in vitro and in vivo pH measurements in plant vacuoles have been examined. The trials indicate that the microelectrodes can be kept for up to 48 h without modification of their slope (mV/pH unit) and resistance. The H +-selective liquid can be used more than 4 months after being prepared. The vacuole is known to be the storage compartment of plant cells where solutes are accumulated; as an example, sucrose and a model protein, bovine serum albumin (BSA), were chosen to study the effect of solutes on the response of the microelectrodes. The results indicate that the slope of the regression line is not modified by sucrose (20–100 m m) added to citrate buffer solution, but it is slightly decreased when the microelectrodes are tested in the presence of BSA or in plant juice, indicating that some components of the cell sap are able to interfere with and to modify the response of the microelectrodes. More experiments are needed in order to determini it proteins, ions, or another substance are the factors causing this effect. Microelectrodes with tip diameters in the range 0.3–0.6 μm and electrical resistance in the range 2 × 10 12 ohms are shown to be suitable for the measurement of vacuolar pH in plant cells. Their short response time (several seconds) when inserted into vacuoles makes them appropriate for follwing vacuolar pH modifications in situ.

Regulation of internal pH in plant cells: The relationship between cytosol and vacuole

Regulation of internal pH in plant cells: The relationship between cytosol and vacuole

Regulation of Cytoplasmic and Vacuolar pH in Maize Root Tips under Different Experimental Conditions

(31)P-Nuclear magnetic resonance spectra of perfused maize (Zea mays L., hybrid WW x Br 38) root tips, obtained at 10-minute intervals over 12 hours or longer, indicate that no cytoplasmic or vacuolar pH changes occur in these cells in the presence of 25 millimolar K(2)SO(4), which induces extrusion of 4 to 5 microequivalents H(+) per gram per hour. In contrast, hypoxia causes cytoplasmic acidification (0.3-0.6 pH unit) without a detectable change in vacuolar pH. The cytoplasm quickly returns to its original pH on reoxygenation. Dilute NH(4)OH increases the vacuolar pH more than it does the cytoplasmic pH; after NH(4)OH is removed, the vacuole recovers its original pH more slowly than does the cytoplasm. The results indicate that regulation of cytoplasmic pH and that of vacuolar pH in plant cells are separate processes.