The addition of 100 mM NaCl to the root medium of barley plants caused the rapid cessation of elongation of the growing leaf three, followed by a sudden resumption of growth during the following hour. The idea that resumption of growth is preceded and mediated by rapid and tissue-specific changes in ABA concentration and by changes in transpiration was tested. Leaf elongation velocity was recorded continuously using linear variable displacement transducers (LVDT), ABA was determined by immunoassay, and transpiration and stomatal conductivity were measured gravimetrically and by porometry, respectively. Within 10 min following addition of salt, ABA increased 6-fold in the distal portion of the leaf elongation zone; in the proximal portion, ABA accumulated with a delay. In the portion of the growing blade that had emerged ABA increased 3-fold and remained elevated during the following 20 min. This preceded a decrease in transpiration and stomatal conductivity, which, in turn, coincided with growth resumption. Twenty hours following the addition of salt, the ABA concentrations had returned to the level before stress. Leaf elongation velocity was still reduced. It is concluded that NaCl causes a rapid increase in ABA in the transpiring portion of the growing leaf. This leads to a decrease in transpiration. As a result, xylem water potential is expected to rise. The moment that the water potential gradient between the xylem and the peripheral cells in the growth zone favours water uptake again into the latter, leaf elongation resumes. The results suggest that ABA causes different responses in different leaf regions, all aimed at promoting the resumption of leaf growth.

Recent results concerning the short-term growth response to salinity of the developing barley leaf are reviewed. Plants were grown hydroponically and the growth response of leaf 3 was studied between 10 min and 5 d following addition of 100 mM NaCl to the root medium. The aim of the experiments was to relate changes in variables that are likely to affect cell elongation to changes in leaf growth. Changes in hormone content (ABA, cytokinins), water and solute relationships (osmolality, turgor, water potential, solute concentrations), gene expression (water channel), cuticle deposition, membrane potential, and transpiration were followed, while leaf elongation velocity was monitored. Leaf elongation decreased close to zero within seconds following addition of NaCl. Between 20 and 30 min after exposure to salt, elongation velocity recovered rather abruptly, to about 46% of the pre-stress level, and remained at the reduced rate for the following 5 d, when it reached about 70% of the level in non-stressed plants. Biophysical and physiological analyses led to three major conclusions. (i) The immediate reduction and sudden recovery in elongation velocity is due to changes in the water potential gradient between leaf xylem and peripheral elongating cells. Changes in transpiration, ABA and cytokinin content, water channel expression, and plasma membrane potential are involved in this response. (ii) Significant solute accumulation, which aids growth recovery, is detectable from 1 h onwards; growing and non-growing leaf regions and mesophyll and epidermis differ in their solute response. (iii) Cuticular wax density is not affected by short-term exposure to salt; transpirational changes are due to stomatal control.

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 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.

Lipid transfer proteins (LTPs) participate in many important physiological processes in plants, including adaptation to stressors, e.g., salinity. Here we address the mechanism of this protective action of LTPs by studying the interaction between LTPs and abscisic acid (ABA, a “stress” hormone) and their mutual participation in suberin deposition in root endodermis of salt-stressed pea plants. Using immunohistochemistry we show for the first time NaCl induced accumulation of LTPs and ABA in the cell walls of phloem paralleled by suberin deposition in the endoderm region of pea roots. Unlike LTPs which were found localized around phloem cells, ABA was also present within phloem cells. In addition, ABA treatment resulted in both LTP and ABA accumulation in phloem cells and promoted root suberization. These results suggested the importance of NaCl-induced accumulation of ABA in increasing the abundance of LTPs and of suberin. Using molecular modeling and fluorescence spectroscopy we confirmed the ability of different plant LTPs, including pea Ps-LTP1, to bind ABA. We therefore hypothesize an involvement of plant LTPs in ABA transport (unloading from phloem) as part of the salinity adaptation mechanism.