Numerous biotechnological applications require a fast and efficient clonal propagation of whole plants under controlled laboratory conditions. For most plant species, the de novo regeneration of shoots from the cuttings of various plant organs can be obtained on nutrient media supplemented with plant hormones, auxin and cytokinin. While auxin is needed during the early stages of the process that include the establishment of pluripotent primordia and the subsequent acquisition of organogenic competence, cytokinin-supplemented media are required to induce these primordia to differentiate into developing shoots. The perception of cytokinin through the receptor ARABIDOPSIS HISTIDINE KINASE4 (AHK4) is crucial for the activation of the two main regulators of the establishment and maintenance of shoot apical meristems (SAMs): SHOOTMERISTEMLESS (STM) and the WUSCHEL-CLAVATA3 (WUS-CLV3) regulatory circuit. In this review, we summarize the current knowledge of the roles of the cytokinin signaling cascade in the perception and transduction of signals that are crucial for the de novo establishment of SAMs and lead to the desired biotechnological output—adventitious shoot multiplication. We highlight the functional differences between individual members of the multigene families involved in cytokinin signal transduction, and demonstrate how complex genetic regulation can be achieved through functional specialization of individual gene family members.

Cytokinins (CKs) are a class of phytohormones that regulate a wide range of developmental processes in plants. Different structural forms of these phytohormones have been widely used in plant tissue culture, although comparative studies on their temporal effects on gene expression remain limited. In this study, we compared the effects of trans-zeatin (tZ), meta-topolin (mT), and thidiazuron (TDZ) on the expression dynamics of ten CK-responsive genes associated with shoot growth and development in Arabidopsis thaliana. Ten-day-old Arabidopsis seedlings were exposed to liquid nutrient media containing 5 μM tZ, mT, or TDZ, or a control medium, for 15 min and 1, 2, 4, 8, and 24 h. Gene expression was analyzed by RT-qPCR using three independent biological replicates. All tested genes were strongly upregulated at 4 h following mT treatment, whereas most genes showed the highest expression at 8 h in response to TDZ. In contrast, tZ generally induced weaker and slower transcriptional responses. Expression levels of all analyzed genes returned to control-like or lower levels within 24 h. Overall, the three structurally distinct cytokinins induced markedly different temporal transcriptional responses, with mT producing the fastest and most pronounced effects on the expression of the tested genes under the applied experimental conditions. These findings improve our understanding of CK-dependent transcriptional dynamics during shoot development and provide a basis for future studies on CK-regulated morphogenesis and plant regeneration.