G. R. Akhiyarova

Participation of Plant Hormones in Growth Resumption of Wheat Shoots Following Short-Term NaCl Treatment

The effects of sodium-chloride salinity on the leaf elongation rate, transpiration rate, cell sap osmolality, and phytohormone content in 7-day-old shoots of durum wheat (Triticum durum L.) were studied. Leaf growth was suppressed under the salinity stress and resumed 1 h after NaCl removal. The resumption of leaf growth coincided with a decrease in the transpiration rate due to the rapid ABA accumulation in the differentiation leaf zone. The increased IAA concentration in the growing leaf zone promoted the formation of the attraction signal. The authors concluded that the changes in phytohormonal status in wheat plants occurred already following short-term (up to 1 h) salinity and were directed to the maintenance of plant growth under these conditions.

Accumulation of cytokinins in roots and their export to the shoots of durum wheat plants treated with the protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP)

Cytokinin flow from roots to shoots can serve as a long-distance signal important for root-to-shoot communication. In the past, changes in cytokinin flow from roots to shoots have been mainly attributed to changes in the rate of synthesis or breakdown in the roots. The present research tested the possibility that active uptake of cytokinin by root cells may also influence its export to shoots. To this end, we collapsed the proton gradient across root membranes using the protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP) to inhibit secondary active uptake of exogenous and endogenous cytokinins. We report the impact of CCCP on cytokinin concentrations and delivery in xylem sap and on accumulation in shoots of 7-day-old wheat plants in the presence and absence of exogenous cytokinin applied as zeatin. Zeatin treatment increased the total accumulation of cytokinin in roots and shoots but the effect was smaller for the shoots. Immunohistochemical localization of cytokinins using zeatin-specific antibodies showed an increase in immunostaining of the cells adjacent to xylem in the roots of zeatin-treated plants. Inhibition of secondary active cytokinin uptake by CCCP application decreased cytokinin accumulation in root cells but increased both flow from the roots and accumulation in the shoots. The possible importance of secondary active uptake of cytokinins by root cells for the control of their export to the shoot is discussed.

The Role of Hormones in Fast Growth Responses of Wheat Plants to Osmotic and Cold Shocks

The effects of nutrient-solution cooling and PEG addition to the nutrient solution on the phytohormone content, the rate of leaf growth, leaf extensibility under the influence of external mechanical action, osmotic potential, and transpiration were studied in seven-day-old wheat plants. Leaf growth rapidly ceased, and the transpiration rate was reduced in both treatments. Growth cessation induced by PEG was transient, and growth resumption was preceded by an increase in the leaf extensibility. The functional role of auxin accumulation in plant shoots in the control of extensibility as well as the relationship between the ABA accumulation and a decrease in the cytokinin content, on the one hand, and reduced transpiration, on the other hand, under stress conditions are discussed.

Effects of nitrate and ammonium on growth of Arabidopsis thaliana plants transformed with the ARR5::GUS construct and a role for cytokinins in suppression of disturbances induced by the presence of ammonium

Determination of physiologically active cytokinins in transformed Arabidopsis thaliana (L.) Heynh. plants (ecotype Wassilevskija) by means of the ARR5::GUS gene expression analysis and by the immunohistochemical assay demonstrated an increase of the cytokinin content during leaf growth and its decrease after growth cessation. After leaf growth cessation cytokinins (CKs) were found only in chloroplasts. Nitrate but not ammonium salt increased the cytokinin content in leaves. The replacement of nitrate in nutrient medium with ammonium salts suppressed leaf and root growth and decreased the content of physiologically active CKs in them. In plants cultivated on a media with NH4Cl as a sole nitrogen source, 10–9 M trans-zeatin several-fold activated leaf growth, prevented a decrease in the chlorophyll a and b content, and reduced proline accumulation indicating suppression of the NH4Cl stress action. Thus, negative effects of NH4Cl as a sole nitrogen source depended on the cytokinin shortage.

Dependence of cytokinin distribution in plants on their physical and chemical properties and transpiration rate

The effect of transpiration on cytokinin accumulation and distribution in 7-day-old wheat (Triticum durum Desf.) seedlings grown on nutrient medium supplemented with zeatin or its riboside was studied. The content of cytokinins in plants and nutrient medium was measured by the immunoenzyme analysis; cytokinin distribution between root cells was assessed immunohistochemically using antibodies against zeatin derivatives. The rate of transpiration was reduced 20-fold by plant placing in humid chamber. At normal transpiration, after 6 h of plant incubation on the solution of zeatin, the level of cytokinins in plant tissues increased stronger than after incubation on the solution of zeatin riboside (by 7.3 and 3.5 times, respectively, as compared with control), although the rates of both cytokinin uptake were equal. Most portions of cytokinins were retained in the roots, which was stronger expressed in the case of free zeatin uptake. A decrease in the rate of transpiration did not affect substantially the zeatin absorption from nutrient medium and the total level of cytokinin accumulation in plants, but these indices were sharply decreased in the case of zeatin riboside. In the zone of absorption of both control roots and roots treated with cytokinins, more intense cytokinin immunostaining was observed in the cells of the central cylinder. The interrelation between cytokinin distribution between the cells and apoplast, their inactivation, and transport over the plant and their form (zeatin or zeatin riboside) used for treatment is discussed.

The influence of local IPT gene induction in roots on content of cytokinins in cells of tobacco leaves

Little attention has been paid to detection of cytokinins in differentiated leave cells. We carried out immunochemical staining of cytokinins in transgenic tobacco plants, in which the content of cytokinins increased as a result of induction of the ipt gene expression (which controls the synthesis of cytokinins) in roots. Staining on cytokinins (based on using antibodies against zeatin riboside) was typical for mesophyll cells. The tag was localized in cytoplasm adjacent to the cell wall and was almost absent in vacuoles. Chloroplasts were clearly visible due to tag concentration around them in cytoplasm. Immunohistochemical staining also detected the presence of cytokinins in stomatal cells. Induction of the synthesis of cytokinins increased immunochemical staining of both mesophyll and stomatal cells, which was accompanied by an increase in the stomatal conductance. The possibility of a direct influence on stomatal conductance of cytokinins (accumulating directly in stomatal cells) and mediated influence through photosynthesis in mesophyll cells is discussed.

Water relations and growth of original barley plants and its ABA-deficient mutants at increased air temperature

Data on the effects of air temperature increase by 4°C on leaf growth and water relation parameters in barley (Hordeum vulgare L.) plants in original cv. Steptoe and its ABA-deficient mutant (AZ24) are presented. An increase in temperature firstly resulted in the cessation of leaf elongation in both genotypes; however, later in cv. Steptoe plants, as distinct from mutants, the rate of leaf length increment was completely restored. Before air warming, transpiration was more intense in mutant plants; at increased temperature, transpiration was activated in both genotypes. After growth resumption, the water potential in cv. Steptoe plants somewhat increased as compared with initial level (before warming). In AZ34 leaves, in contrast, the water potential, which was initially below that in cv. Steptoe leaves, reduced after temperature increase. The calculation of total hydraulic conductivity of the plants and osmotic hydraulic conductivity in the roots showed that these parameters increased in cv. Steptoe and were not changed in AZ34 mutants. At temperature increase, the level of ABA was not changed in AZ34 mutants, whereas in Steptoe plants it increased in the roots and decreased in the shoots. It was concluded that a capability of ABA synthesis is required for the control of total hydraulic conductivity under changing environmental conditions.

Study of cytokinin transport from shoots to roots of wheat plants is informed by a novel method of differential localization of free cytokinin bases or their ribosylated forms by means of their specific fixation

The aim of the present report was to demonstrate how a novel approach for immunohistochemical localization of cytokinins in the leaf and particularly in the phloem may complement to the study of their long-distance transport. Different procedures of fixation were used to conjugate either cytokinin bases or their ribosides to proteins of cytoplasm to enable visualization and differential localization of these cytokinins in the leaf cells of wheat plants. In parallel to immunolocalization of cytokinins in the leaf cells, we immunoassayed distribution of free bases of cytokinins, their nucleotides and ribosides between roots and shoots of wheat plants as well as their presence in phloem sap after incubation of leaves in a solution supplemented with either trans-zeatin or isopentenyladenine. The obtained data show ribosylation of the zeatin applied to the leaves and its elevated level in the phloem sap supported by in vivo localization showing the presence of ribosylated forms of zeatin in leaf vessels. This suggests that conversion of zeatin to its riboside is important for the shoot-to-root transport of zeatin-type cytokinins in wheat. Exogenous isopentenyladenine was not modified, but diffused from the leaves as free base. These metabolic differences may not be universal and may depend on the plant species and age. Although the measurements of cytokinins in the phloem sap and root tissue is the most defining for determining cytokinin transport, study of immunolocalization of either free cytokinin bases or their ribosylated forms may be a valuable source of information for predicting their transport in the phloem and to the roots.