•Five IPT genes were isolated from Streptocarpus rexii.•Realtime-PCR and in-situ hybridization revealed differential expression of IPT genes.•IPT5 and IPT9 express in the macrocotyledon and phyllomorph, and also in the groove meristem.•IPT5 and IPT9 reacted differently to the external hormone treatment.The enzyme ISOPENTENYLTRANSFERASE (IPT) is responsible for the rate limiting step of cytokinin biosynthesis, an important plant hormone with key roles in meristem maintenance and organ development. In this study, we isolated IPT genes from the acaulescent Streptocarpus rexii, a plant that shows an unorthodox development starting with post-germination anisocotyly, in which cytokinins play an integral role. Three adenosine phosphate-IPTs and two tRNA-IPTs were isolated from S. rexii. Their expression levels and patterns in different tissues were compared by means of realtime-PCR and mRNA in-situ hybridization. We found that each SrIPT had a distinctive expression pattern. Interestingly, in vegetative tissues as well as in meristems only the adenosine phosphate-IPT SrIPT5 and the tRNA-IPT SrIPT9 were found. In addition, they were differentially affected by external hormone application, suggesting their different regulation and expression during meristem formation and maintenance and lamina growth. Our results indicate that SrIPTs are involved in shaping the architecture of S. rexii, working differentially and redundantly, and show that differentially expressed IPT genes regulate plant form.

Cytokinins (CKs) are a group of phytohormones that play a crucial role in the regulation of plant growth and development. Identification of the enzymes and the corresponding genes that are involved in CK metabolism allowed us to understand how plants synthesize CKs and adjust CK activity to optimal levels. A major accomplishment toward these goals was the identification of genes for the first enzyme in the CK biosynthetic pathway, adenosine phosphate-isopentenyltransferase (IPT). In Arabidopsis thaliana and Agrobacterium tumefaciens, detailed analyses of IPTs were conducted through not only enzymatic characterization but also molecular structural approaches. These studies revealed the molecular basis for the Agrobacterium-origin of IPT used for the efficient biosynthesis of trans-zeatin that promotes tumorigenesis in host plants. Another landmark in CK research was the identification of CYP735A as an enzyme that converts iP-nucleotide to tZ-nucleotide. Furthermore, the identification of a CK-activating enzyme, LOG, which catalyzes a novel activation pathway, is a remarkable recent achievement in CK research. Collectively, these advances have revealed the complexity of the entire metabolic scheme for CK biosynthesis.Identification of the enzymes and the corresponding genes that are involved in cytokinin metabolism allowed us to understand how plants synthesize cytokinins and adjust their activity to optimal levels. These advances also have revealed the complexity of the entire metabolic scheme for CK biosynthesis.

The growth of plants depends on continuous function of the meristems. Shoot meristems are responsible for all the post-embryonic aerial organs, such as leaves, stems and flowers1. It has been assumed that the phytohormone cytokinin has a positive role in shoot meristem function2, 3, 4. A severe reduction in the size of meristems in a mutant that is defective in all of its cytokinin receptors has provided compelling evidence that cytokinin is required for meristem activity5, 6. Here, we report a novel regulation of meristem activity, which is executed by the meristem-specific activation of cytokinins. The LONELY GUY (LOG) gene of rice is required to maintain meristem activity and its loss of function causes premature termination of the shoot meristem. LOG encodes a novel cytokinin-activating enzyme that works in the final step of bioactive cytokinin synthesis. Revising the long-held idea of multistep reactions, LOG directly converts inactive cytokinin nucleotides to the free-base forms, which are biologically active, by its cytokinin-specific phosphoribohydrolase activity. LOG messenger RNA is specifically localized in shoot meristem tips, indicating the activation of cytokinins in a specific developmental domain. We propose the fine-tuning of concentrations and the spatial distribution of bioactive cytokinins by a cytokinin-activating enzyme as a mechanism that regulates meristem activity.

•The trans-hydroxylation of CK is catalyzed by CYP735A1/A2 in Arabidopsis•trans-hydroxylated CKs are required for shoot growth but dispensable for root growth•trans-hydroxylation of CKs affects their efficacy but not total CK levelsCytokinins (CKs), a class of plant hormones, are central regulators of plant growth and development. Based on numerous physiological and genetic studies, the quantitative regulation of cytokinin levels is the major mechanism regulating cytokinin action in diverse developmental processes. Here, we identified a different mechanism with which the physiological function of CK is modulated through side-chain modification (trans-hydroxylation). The trans-hydroxylation that forms trans-zeatin (tZ)-type CK from N6-(Δ2-isopentenyl)adenine (iP)-type CK is catalyzed by the cytochrome P450 enzymes CYP735A1 and CYP735A2 in Arabidopsis. Deficiency in trans-hydroxylation activity results in dramatic retardation of shoot growth without affecting total CK quantity, while augmentation of the activity enhances shoot growth. Application of exogenous tZ but not iP recovers the wild-type phenotype in the mutants, indicating that trans-hydroxylation modifies the physiological function of CK. We propose that the control of cytokinin function by side-chain modification is crucial for shoot growth regulation in plants.