Single and combined effects of Cd and Pb on the growth, medium pH, membrane potential and metal contents in maize (Zea mays L.) coleoptile segments.

The mechanisms of the toxic effects of Cd and Pb on plant cell growth are still poorly understood. In particular, little is known about their interactive effects, which usually occur in the environment. Moreover, the data that do exist in the literature are controversial. This study describes experiments that were performed with maize (Zea mays) coleoptile segments, which is a classical model system for studies of plant cell elongation growth. Cadmium and lead, which were added at 0.1 mM, reduced the endogenous and IAA-induced elongation growth of maize coleoptile cells. When both metals were added together or in sequence, their effect on IAA-induced growth was more toxic. The medium pH changes, which were measured simultaneously with growth, indicated that while Pb stopped IAA-induced proton extrusion, Cd only partially diminished it. Although Cd was generally more accumulated than Pb in the maize coleoptile segments, when IAA was added together with Pb, it significantly increased the accumulation of the metal. The short-term electrophysiological experiments showed that the addition of Cd caused the depolarisation of the membrane potential (Em), whereas Pb caused membrane hyperpolarisation. In the long-term electrophysiological experiments, it was found that the Cd-induced Em changes are complex. In conclusion, these results suggest that the effects of Cd and Pb as well as their combination on the elongation growth of maize coleoptile cells and the accumulation of the metals result, among others, from different ionic mechanisms by which each metal change the membrane potential of the cells.

Role of auxin (IAA) in the regulation of slow vacuolar (SV) channels and the volume of red beet taproot vacuoles.

BACKGROUND: Auxin (IAA) is a central player in plant cell growth. In contrast to the well-established function of the plasma membrane in plant cell expansion, little is known about the role of the vacuolar membrane (tonoplast) in this process. RESULTS: It was found that under symmetrical 100 mM K+ and 100 µM cytoplasmic Ca2+ the macroscopic currents showed a typical slow activation and a strong outward rectification of the steady-state currents. The addition of IAA at a final concentration of 1 µM to the bath medium stimulated the SV currents, whereas at 0.1 and 10 µM slight inhibition of SV currents was observed. The time constant, τ, decreased in the presence of this hormone. When single channels were analyzed, an increase in their activity was recorded with IAA compared to the control. The single-channel recordings that were obtained in the presence of IAA showed that auxin increased the amplitude of the single-channel currents. Interestingly, the addition of IAA to the bath medium with the same composition as the one that was used in the patch-clamp experiments showed that auxin decreased the volume of the vacuoles. CONCLUSIONS: It is suggested that the SV channels and the volume of red beet taproot vacuoles are modulated by auxin (IAA).

Effects of juglone and lawsone on oxidative stress in maize coleoptile cells treated with IAA.

Naphthoquinones are secondary metabolites widely distributed in nature and produced by bacteria, fungi and higher plants. Their biological activity may result from induction of oxidative stress, caused by redox cycling or direct interaction with cellular macromolecules, in which quinones act as electrophiles. The redox homeostasis is known as one of factors involved in auxin-mediated plant growth regulation. To date, however, little is known about the crosstalk between reactive oxygen species (ROS) produced by quinones and the plant growth hormone auxin (IAA). In this study, redox cycling properties of two naphthoquinones, juglone (5-hydroxy-1,4-naphthoquinone) and lawsone (2-hydroxy-1,4-naphthoquinone), were compared in experiments performed on maize coleoptile segments incubated with or without the addition of IAA. It was found that lawsone was much more effective than juglone in increasing both H2O2 production and the activity of antioxidative enzymes (SOD, POX and CAT) in coleoptile cells, regardless of the presence of IAA. An increase in the activity of Cu/Zn-SOD isoenzymes induced by both naphthoquinones suggests that juglone- and lawsone-generated H2O2 was primarily produced in the cytosolic and cell wall spaces. The cell potential to neutralize hydrogen peroxide, determined by POX and CAT activity, pointed to activity of catalase as the main enzymatic mechanism responsible for degradation of H2O2 Therefore, we assumed that generation of H2O2, induced more efficiently by LW than JG, was the major factor accounting for differences in the toxicity of naphthoquinones in maize coleoptiles. The role of auxin in the process appeared negligible. Moreover, the results suggested that oxidative stress imposed by JG and LW was one of mechanisms of allelopathic action of the studied quinones in plants.

Role of chloride ions in the promotion of auxin-induced growth of maize coleoptile segments.

BACKGROUND AND AIMS: The mechanism of auxin action on ion transport in growing cells has not been determined in detail. In particular, little is known about the role of chloride in the auxin-induced growth of coleoptile cells. Moreover, the data that do exist in the literature are controversial. This study describes experiments that were carried out with maize (Zea mays) coleoptile segments, this being a classical model system for studies of plant cell elongation growth. METHODS: Growth kinetics or growth and pH changes were recorded in maize coleoptiles using two independent measuring systems. The growth rate of the segments was measured simultaneously with medium pH changes. Membrane potential changes in parenchymal cells of the segments were also determined for chosen variants. The question of whether anion transport is involved in auxin-induced growth of maize coleoptile segments was primarily studied using anion channel blockers [anthracene-9-carboxylic acid (A-9-C) and 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS)]. In addition, experiments in which KCl was replaced by KNO3 were also performed. KEY RESULTS: Both anion channel blockers, added at 0·1 mm, diminished indole-3-acetic acid (IAA)-induced elongation growth by ~30 %. Medium pH changes measured simultaneously with growth indicated that while DIDS stopped IAA-induced proton extrusion, A-9-C diminished it by only 50 %. Addition of A-9-C to medium containing 1 mm KCl did not affect the characteristic kinetics of IAA-induced membrane potential changes, while in the presence of 10 mm KCl the channel blocker stopped IAA-induced membrane hyperpolarization. Replacement of KCl with KNO3 significantly decreased IAA-induced growth and inhibited proton extrusion. In contrast to the KCl concentration, the concentration of KNO3 did not affect the growth-stimulatory effect of IAA. For comparison, the effects of the cation channel blocker tetraethylammonium chloride (TEA-Cl) on IAA-induced growth and proton extrusion were also determined. TEA-Cl, added 1 h before IAA, caused reduction of growth by 49·9 % and inhibition of proton extrusion. CONCLUSIONS: These results suggest that Cl(-) plays a role in the IAA-induced growth of maize coleoptile segments. A possible mechanism for Cl(-) uptake during IAA-induced growth is proposed in which uptake of K(+) and Cl(-) ions in concert with IAA-induced plasma membrane H(+)-ATPase activity changes the membrane potential to a value needed for turgor adjustment during the growth of maize coleoptile cells.

Interactive effects of temperature and heavy metals (Cd, Pb) on the elongation growth in maize coleoptiles.

The effect of Cd and Pb on endogenous and IAA-induced elongation growth and medium pH of maize coleoptile segments incubated at 20, 25 and 30 °C was studied. It was found that the elongation of coleoptile segments and proton extrusion increased with the temperature and reached its maximum at 30 °C. For Cd, the maximal inhibition of endogenous and IAA-induced growth as well as medium acidification of coleoptile segments was observed at 25 °C. Meanwhile, Pb, irrespective of the temperature, diminished the growth of the segments by ca. 20%, increasing the acidification of the incubation medium. It was also found that in contrast to Cd, Pb accumulation in maize coleoptile segments did not correlate with temperature. The results suggest that the toxic effect of Cd on elongation growth of coleoptile segments is connected with the decrease of the PM H(+)-ATPase activity and probably with Cd-induced high acivity of IAA oxidase, whereas the effect of Pb did not depend on activity of any of the enzymes.

Effect of cadmium and lead on the membrane potential and photoelectric reaction of Nitellopsis obtusa cells.

The effects of Cd and Pb on membrane potential (E(m)) and photoelectric reaction of Nitellopsis obtusa cells were investigated. It was found that Cd and Pb at 1.0 mM caused a depolarization of the E(m), whereas both metals at lower concentrations changed the E(m) in a different way. Pb at 0.1 mM and 0.01 mM hyperpolarized the E(m), whereas Cd at the same concentrations depolarized and did not change the E(m), respectively. In the presence of 0.01 mM Pb, the light-induced hyperpolarization of the E(m) was by 18% higher as compared to the control, whereas at 1.0 mM Pb it was by 40% lower. Pb at 0.1 mM and Cd at 0.01 mM or 5 × 0.01 mM did not change the light-induced membrane hyperpolarization. However, in the presence of Cd at 0.1 mM and 1.0 mM this hyperpolarization was 2-fold lower or was completely abolished, respectively. These results suggest that at high Cd and Pb concentrations both depolarization of the E(m) and decrease of light-induced membrane hyperpolarization in Nitellopsis obtusa cells are probably due to inhibition of the plasma membrane H(+)-ATPase activity, whereas both metals at lower concentrations differ in mechanism of membrane potential changes.

Fusicoccin counteracts the toxic effect of cadmium on the growth of maize coleoptile segments.

The effects of cadmium (Cd; 0.1-1000 µM) and fusicoccin (FC) on growth, Cd(2+) content, and membrane potential (E(m)) in maize coleoptile segments were studied. In addition, the E(m) changes and accumulation of Cd and calcium (Ca) in coleoptile segments treated with Cd(2+) combined with 1 µM FC or 30 mM tetraethylammonium (TEA) chloride (K(+)-channel blocker) were also determined. In this study, the effects of Ca(2+)-channel blockers [lanthanum (La) and verapamil (Ver)] on growth and content of Cd(2+) and Ca(2+) in coleoptile segments were also investigated. It was found that Cd at high concentrations (100 and 1000 µM) significantly inhibited endogenous growth of coleoptile segments and simultaneously measured proton extrusion. FC combined with Cd(2+) counteracted the toxic effect of Cd(2+) on endogenous growth and significantly decreased Cd(2+) content (not the case for Cd(2+) at the highest concentration) in coleoptile segments. Addition of Cd to the control medium caused depolarization of E (m), the extent of which was dependent on Cd concentration and time of treatment with Cd(2+). Hyperpolarization of E(m) induced by FC was suppressed in the presence of Cd(2+) at 1000 µM but not Cd(2+) at 100 µM. It was also found that treatment of maize coleoptile segments with 30 mM TEA chloride caused hyperpolarization of E (m) and decreased Cd(2+) content in coleoptile segments, suggesting that, in the same way as for FC, accumulation of Cd(2+) was dependent on plasma membrane (PM) hyperpolarization. Similar to FC, TEA chloride also decreased Ca(2+) content in coleoptile segments. La and Ver combined with Cd(2+) (100 µM) significantly decreased Cd content in maize coleoptile segments, but only La completely abolished the toxic effect of Cd(2+) on endogenous growth and growth in the presence of FC. Taken together, these results suggest that the mechanism by which FC counteracts the toxic effect of Cd(2+) (except at 1000 µM Cd(2+)) on the growth of maize coleoptile segments involves both stimulation of PM H(+)-ATPase activity by FC as well as Cd(2+)-permeable, voltage-dependent Ca channels, which are blocked by FC and TEA chloride-induced PM hyperpolarization.