Cytokinin receptors play a key role in cytokinin-dependent processes regulating plant growth, development, and adaptation; therefore, the functional properties of these receptors are of great importance. Previously the properties of cytokinin receptors were investigated in heterologous assay systems using unicellular microorganisms, mainly bacteria, expressing receptor proteins. However, within microorganisms receptors reside in an alien environment that might distort the receptor properties. Therefore, a new assay system has been developed allowing studies of individual receptors within plant membranes (i.e. closer to their natural environment). The main ligand-binding characteristics of receptors from Arabidopsis [AHK2, AHK3, and AHK4] and maize (ZmHK1) were refined in this new system, and the properties of full-length Arabidopsis receptor AHK2 were characterized for the first time. Ligand specificity profiles of receptors towards cytokinin bases were comparable with the profiles retrieved in bacterial assay systems. In contrast, cytokinin-9-ribosides displayed a strongly reduced affinity for receptors in the plant assay system, indicating that ribosides as the common transport form of cytokinins have no or very weak cytokinin activity. This underpins the central role of free bases as the sole biologically active cytokinin compounds. According to molecular modelling and docking studies, N⁹-ribosylation alters the bonding pattern in cytokinin–receptor interaction and prevents β6–β7 loop movement important for tight hormone binding. A common feature of all receptors was a greatly reduced ligand binding at low (5.0–5.5) pH. The particularly high sensitivity of ZmHK1 to pH changes leads to the suggestion that some cytokinin receptors may play an additional role as pH sensors in the lumen of the endoplasmic reticulum.

The ligand-binding properties of the maize (Zea mays L.) cytokinin receptors ZmHK1, ZmHK2, and ZmHK3a have been characterized using cytokinin binding assays with living cells or membrane fractions. According to affinity measurements, ZmHK1 preferred N⁶-(Δ²-isopentenyl)adenine (iP) and had nearly equal affinities to trans-zeatin (tZ) and cis-zeatin (cZ). ZmHK2 preferred tZ and iP to cZ, while ZmHK3a preferred iP. Only ZmHK2 had a high affinity to dihydrozeatin (DZ). Analysis of subcellular fractions from leaves and roots of maize seedlings revealed specific binding of tZ in the microsome fraction but not in chloroplasts or mitochondria. In competitive binding assays with microsomes, tZ and iP were potent competitors of [³H]tZ while cZ demonstrated significantly lower affinity; adenine was almost ineffective. The binding specificities of microsomes from leaf and root cells for cytokinins were consistent with the expression pattern of the ZmHKs and our results on individual receptor properties. Aqueous two-phase partitioning and sucrose density-gradient centrifugation followed by immunological detection with monoclonal antibody showed that ZmHK1 was associated with the endoplasmic reticulum (ER). This was corroborated by observations of the subcellular localization of ZmHK1 fusions with green fluorescent protein in maize protoplasts. All these data strongly suggest that at least a part of cytokinin perception occurs in the ER.

Tuber formation is a complex process comprising several stages: stolon formation and growth, induction of tuberization, tuber initiation, and tuber growth. This review considers successive stages of tuber formation and their hormonal regulation. Special attention is paid to the effects on tuber formation of such phytohormones as gibberellins, cytokinins, jasmonic acid, and auxins. Physiological and some molecular-genetic aspects of their action on tuber photoperiodic induction and initiation are discussed.

The lecture presents modern knowledge of the mechanisms of cytokinin perception and signal transduction to the genes of primary and secondary responses. It also demonstrates the relations between the rapid cytokinin-induced processes and cytokinin-induced physiological effects. The characteristics of the cytokinin regulatory system and its role in the control of plant growth and development are discussed.