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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.

Facultative plant hemiparasites exhibit optimal growth only when attached to a suitable host. After attachment, stomata of the parasite remain continuously open, thus, optimising the extraction of host xylem sap. When the host shoot was removed from the hemiparasitic Rhinanthus/barley association ∼14 days after attachment, the resulting host-free attached Rhinanthus continued to grow and develop similarly well as the attached parasites. These plants, however, showed altered stomatal behaviour: their stomata were open at daytime and closed at night, whereas parasitising Rhinanthus has continuously open stomata all day and night and unattached single Rhinanthus has practically closed stomata throughout day and night. After removal of the host the root growth was strongly increased, thereby increasing the root-to-shoot ratio. Abscisic acid and cytokinin relationships became more 'normal' with the Rhinanthus roots becoming able to synthesise zeatin nucleotides and zeatin ribosides, thus, behaving much as non-parasitic plants in general. It is suggested that the degrading root system of the host plant produces signals that trigger this conversion. Two explanations for these changes are discussed, the supply of dissolved organic nitrogen by the degrading host root system and a possible strong growth of growth promoting soil microorganisms using the degrading host root system as a substrate.Additional keywords: abscisic acid, cytokinins, stomatal behaviour, zeatin.

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