Cytokinins modulate a number of important developmental processes, including the last phase of leaf development, known as senescence, which is associated with chlorophyll breakdown, photosynthetic apparatus disintegration and oxidative damage. There is ample evidence that cytokinins can slow down all these senescence-accompanying changes. Here, we review relationships between the various mechanisms of action of these regulatory molecules. We highlight their connection to photosynthesis, the pivotal process that generates assimilates, however may also lead to oxidative damage. Thus, we also focus on cytokinin induction of protective responses against oxidative damage. Activation of antioxidative enzymes in senescing tissues is described as well as changes in the levels of naturally occurring antioxidative compounds, such as phenolic acids and flavonoids, in plant explants. The main goal of this review is to show how the biological activities of cytokinins may be related to their chemical structure. New links between molecular aspects of natural cytokinins and their synthetic derivatives with antisenescent properties are described. Structural motifs in cytokinin molecules that may explain why these molecules play such a significant regulatory role are outlined.

Plant hormone cytokinins are perceived by a subfamily of sensor histidine kinases (HKs), which via a two-component phosphorelay cascade activate transcriptional responses in the nucleus. Subcellular localization of the receptors proposed the endoplasmic reticulum (ER) membrane as a principal cytokinin perception site, while study of cytokinin transport pointed to the plasma membrane (PM)-mediated cytokinin signalling. Here, by detailed monitoring of subcellular localizations of the fluorescently labelled natural cytokinin probe and the receptor ARABIDOPSIS HISTIDINE KINASE 4 (CRE1/AHK4) fused to GFP reporter, we show that pools of the ER-located cytokinin receptors can enter the secretory pathway and reach the PM in cells of the root apical meristem, and the cell plate of dividing meristematic cells. Brefeldin A (BFA) experiments revealed vesicular recycling of the receptor and its accumulation in BFA compartments. We provide a revised view on cytokinin signalling and the possibility of multiple sites of perception at PM and ER.

Increasing crop productivity is our major challenge if we are to meet global needs for food, feed and fuel. Controlling the level of the plant hormones cytokinins is a mean of improving plant productivity. Cytokinin oxidase/dehydrogenase (CKO/CKX) is a major target in this regard because it degrades cytokinins. Here, we describe the synthesis and biological activities of new CKX inhibitors derived mainly from diphenylurea. They were tested on four CKX isoforms from maize and Arabidopsis, where the best compounds showed IC₅₀ values in the 10 ^-8 M concentration range. The binding mode of the most efficient inhibitors was characterized from high-resolution crystal complexed structures. Although these compounds do not possess intrinsic cytokinin activity, we have demonstrated their tremendous potential for use in the plant tissue culture industry as well as in agriculture. We have identified a key substance, compound 19, which increases stress resistance and seed yield in Arabidopsis, but also improves the yield of wheat, barley and rapeseed grains under field conditions. Thus our findings reveal that subtle modulation of cytokinin levels via CKX inhibition can positively affect plant growth, development and yield, and underline the fact that CKX inhibitors truly deserve to attract interest in plant biotechnology and agriculture

Three cytokinin derivatives (CKd) designated as RR-G, RR-O, and RR-V applied by foliar spraying at tillering, and one compound previously described as a cytokinin antagonist (CKa) designated as RR-P applied as a seed coating were tested in winter wheat and spring barley in field trial experiments. The aim of the study was to examine the influence of the compounds that were tested on the number of productive tillers, grain yield, and endogenous CK content. With the exception of the compound RR-V, the measured parameters clearly showed the stimulatory effects of CKd on tillering and grain yield in spring barley and winter wheat. The RR-V showed a stimulatory effect on the number of productive tillers and yield in spring barley, but not in winter wheat. Although in winter wheat CKa stimulated both the number of productive tillers and the grain yield, there was an inhibitory effect in terms of the number of productive tillers observed in spring barley. The results of the endogenous cytokinin analysis suggested, among others, the importance of the role of isopentenyl-adenine types of cytokinins in the tillering of spring barley. In conclusion, the cytokinin derivative compounds with an agonistic or antagonistic role showed strong potential for application in the future development of plant growth regulators.

We have developed and validated a novel LC-MS/MS method for simultaneously analyzing amino acids, biogenic amines, and their acetylated and methylated derivatives in plants. This method involves a one-step extraction of 2–5 mg of lyophilized plant material followed by fractionation of different biogenic amine forms, and exploits an efficient combination of hydrophilic interaction liquid chromatography (HILIC), reversed phase (RP) chromatography with pre-column derivatization, and tandem mass spectrometry (MS). This approach enables high-throughput processing of plant samples, significantly reducing the time needed for analysis and its cost. We also present a new synthetic route for deuterium-labeled polyamines. The LC-MS/MS method was rigorously validated by quantifying levels of nitrogen-related metabolites in seedlings of seven plant species, including Arabidopsis, maize, and barley, all of which are commonly used model organisms in plant science research. Our results revealed substantial variations in the abundance of these metabolites between species, developmental stages, and growth conditions, particularly for the acetylated and methylated derivatives and the various polyamine fractions. However, the biological relevance of these plant metabolites is currently unclear. Overall, this work contributes significantly to plant science by providing a powerful analytical tool and setting the stage for future investigations into the functions of these nitrogen-related metabolites in plants.