Cytokinin biosynthesis and interconversion

To maintain hormone homeostasis, the rate of cytokinin biosynthesis, interconversion, and degradation is regulated by enzymes in plant cells. Cytokinins can be synthesized via direct (de novo) or indirect (tRNA) pathways. In the de novo pathway, a cytokinin nucleotide is synthesized from 5'-AMP and isopentenyl pyrophosphate; a key enzyme which catalyzes this synthesis has been isolated from plant tissues, slime mold, and some microorganisms. Studies on the in vitro synthesis of the isopentenyl side chain of cytokinin in tRNA demonstrated that the isopentenyl group was derived from mevalonate, and turnover of the cytokinin-containing tRNA may serve as a minor source of free cytokinins in plant cells. The interconversion of cytokinin bases, nucleosides and nucleotides is a major feature of cytokinin metabolism; and enzymes that regulate the interconversion have been identified. The N⁶-side chain and purine moiety of cytokinins are often modified and some of the enzymes involved in the modifications have been isolated. Most of the cytokinin metabolites have been characterized but very few enzymes regulating their metabolism have been purified to homogeneity. It remains a significant challenge to isolate plant genes involved in the regulation of cytokinin biosynthesis, interconversion and degradation.     

Mechanisms of cytokinin action

Cytokinins are involved in the control of numerous and important processes associated with plant growthand development. They take part in the control of cell division, chloroplast development, bud differentiation, shoot initiationand growth or leaf senescence. In contrast to the wide knowledge of cytokinin effects, the mechanisms of cytokinin actionremain largely unknown. Therefore, it is still difficult to explain how a group of molecules can control so many different biological responses. In this review, we propose some arguments in order to solve this question. In a first part, we underline that cytokinins act in concert with other signals for the control of biological responses. Therefore, the effects observed in responses to cytokinins would be not only related to cytokinins but would also be dependent on all other environmental and hormonal signals perceived by plant cells. In a second part, we present the different actors which could be involved in the signalling pathways of cytokinins. It is very likely that these different elements could be implicated in different cytokinin signalling pathways. Therefore, we propose that the diversity of the cytokinin responses could be also correlated with a diversity of cytokinin signalling pathways.     

The interaction of cytokinin with other signals

Cytokinins are important signalling molecules in plants, and recent studies have begun to shed light on the molecular mechanisms underlying their biosynthesis and response pathways. However, from the time of their discovery, it has been clear that cytokinins interact with other signals to regulate plant growth and development. Herein the interaction of cytokinin with three other signals: light, ethylene, and auxin is discussed. The interaction between light and cytokinin signalling, highlighted by recent analysis of cytokinin signalling mutants is reviewed. A discussion of another aspect of cytokinin cross-talk, its induction of ethylene biosynthesis in etiolated Arabidopsis seedlings, and recent studies that have begun to elucidate the mechanism underlying this regulation is also presented. Finally, there is a brief review of the interaction of auxin and cytokinin, and present novel expression profiling data of Arabidopsis seedlings treated with combinations of these two hormones, which provide insight into this interaction.     

Cytokinins: Biosynthesis metabolism and perception

Summary Cytokinins are essential hormones for plant growth and development. They are also of vital importance for in vitro manipulations of plant cells and tissues. The biological activities and chemistry of cytokinins are well defined but very little is known about their mode of action and it is only recently that cytokinin genes have been identified in plants. This review summarizes the current status of knowledge on cytokinin biosynthesis, metabolism and signal transduction, with an emphasis on genes encoding metabolic enzymes and putative receptors, and genes rapidly induced by cytokinins.     

Cytokinin signaling

Although cytokinin plays a central role in plant development, our knowledge of the biosynthesis, distribution, perception and signal transduction of cytokinin is limited. Recent molecular-genetic studies have, however, implicated involvement of a two-component system in cytokinin signal transduction. Furthermore, new mutants with altered cytokinin responses and genes involved in cytokinin signaling have been identified.     

Regulation of cytokinin biosynthesis, compartmentalization and translocation

Cytokinins, a group of mobile phytohormones, play an important role in plant growth and development, and their activity is finely controlled by environmental factors in the control of morphogenic and metabolic adaptations. Inorganic nitrogen sources, such as nitrate, are a major factor regulating gene expression of adenosine phosphate-isopentenyltransferase (IPT), a key enzyme of cytokinin biosynthesis. Modulation of IPT and macronutrient transporter gene expression in response to nitrate, sulphate and phosphate, and cytokinin-dependent repression of the transporter genes suggest that cytokinins play a critical role in balancing acquisition and distribution of macronutrients. Biased distribution of trans-zeatin (tZ)-type cytokinins in xylem and N(6)-(Delta(2)-isopentenyl)adenine (iP)-type cytokinins in phloem saps suggest that, in addition to acting as local signals, cytokinins communicate acropetal and systemic long-distance signals, and that structural side chain variations mediate different biological messages. The compartmentalization of tZ- and iP-type cytokinins implies the involvement of a selective transport system. Recent studies have raised the possibility of subsets of the purine permease family as a transporter of cytokinin nucleobases and equilibrative nucleoside transporters (ENT) for cytokinin nucleosides. These biochemical and transgenic data suggest that AtENT6, an Arabidopsis ENT, could also participate in cytokinin nucleoside transport with a preference for iP riboside in vascular tissue.     

Do cytokinins function as two‐way signals between plants and animals?

Cytokinins are plant hormones that have, among many other functions, senescence‐modulatory effects in plant tissue. This is evident not only from biochemical data, but is vividly illustrated in the “green island” phenotype in plant leaves caused by cytokinins released for example by leaf mining insects or microbial pathogens. It is beyond doubt that, in addition to their roles in plants, cytokinins also provoke physiological and developmental effects in animals. It is hypothesized that the recently much discussed modification of plant metabolism by insects and associated microbes via cytokinin signals has a counterpart in direct cytokinin signalling that interferes with the animals’ hormonal systems and impacts their population dynamics.     

Environmental perception avenues: the interaction of cytokinin and environmental response pathways

Cytokinins were discovered in the 1950s by their ability to promote cell division in cultured plant cells. Recently, there have been significant breakthroughs in our understanding of the biosynthesis, metabolism, perception and signal transduction of this phytohormone. These advances, coupled with physiological and other approaches, have enabled remarkable progress to be made in our understanding of the interactions between cytokinin function and environmental inputs. In this review, we first highlight the most recent advances in our understanding of cytokinin biosynthesis, metabolism and signalling. We then discuss how various environmental signals interact with these pathways to modulate plant growth, development and physiology.     

The Arabidopsis AtIPT8/PGA22 gene encodes an isopentenyl transferase that is involved in de novo cytokinin biosynthesis

Cytokinin plays a critical role in plant growth and development by stimulating cell division and cell differentiation. Despite many years' research efforts, our current understanding of this hormone is still limited regarding both its biosynthesis and signaling. To genetically dissect the cytokinin pathway, we have used a functional screen to identify Arabidopsis gain-of-function mutations that enable shoot formation in the absence of exogenous cytokinins. By using a chemical-inducible activation tagging system, we have identified over 40 putative mutants, designated as pga (plant growth activators), which presumably were affected in key components of cytokinin biosynthesis and signaling pathway. Here, we report a detailed characterization of pga22, a representative mutant from this collection. A gain-of-function mutation in the PGA22 locus resulted in typical cytokinin responses. Molecular and genetic analyses indicated that PGA22 encodes an isopentenyl transferase (IPT) previously identified as AtIPT8. Plants of the pga22 mutant accumulated at remarkably higher levels of isopentenyladenosine-5'-monophosphate and isopentenyladenosine when analyzed by mass spectrometry, suggesting that AtIPT8/PGA22 is a functional IPT that may direct the biosynthesis of cytokinins in planta via an isopentenyladenosine-5'-monophosphate-dependent pathway.     

The role of local biosynthesis of auxin and cytokinin in plant development

Plant hormones are tightly regulated in response to environmental and developmental signals. It has long been speculated that biosynthesis of hormones occurs broadly in plant organs and that intricate, spatiotemporal regulation of hormones in developing organ primordia is achieved through transport and signal perception. However, recent identification of genes crucial for biosynthesis of auxin and cytokinin reveals that localized hormone biosynthesis also plays an important role in organ growth and patterning.     

Strigolactone biosynthesis,transport and perception

Strigolactones (SLs) are plant hormones that regulate diverse developmental processes and environmental responses. They are also known to be root-derived chemical signals that regulate symbiotic and parasitic interactions with arbuscular mycorrhizal fungi and root parasitic plants, respectively. Since the discovery of the hormonal function of SLs in 2008, there has been much progress in the SL research field. In particular, a number of breakthroughs have been achieved in our understanding of SL biosynthesis, transport and perception. The discovery of the hormonal function of SL was quite valuable not only as the identification of a new class of plant hormones, but also as the discovery of the long-sought-after SL biosynthetic and response mutants. These mutants in several plant species provided us the genetic resources to address fundamental questions regarding SL biosynthesis and perception. Such mutants were further characterized later, and biochemical analyses of these genetically identified factors have uncovered the outline of SL biosynthesis and perception so far. Moreover, new genes involved in SL transport have been discovered through reverse genetic analyses. In this review, we summarize recent advances in SL research with a focus on biosynthesis, transport and perception.     

Cytokinins and plant immunity: old foes or new friends?

Cytokinins are plant growth promoting hormones involved in the specification of embryonic cells, maintenance of meristematic cells, shoot formation and development of vasculature. Cytokinins have also emerged as a major factor in plant-microbe interactions during nodule organogenesis and pathogenesis. Microbe-originated cytokinins confer abnormal hypersensitivity of cytokinins to plants, augmenting the sink activity of infected regions. However, recent findings have shed light on a distinct role of cytokinins in plant immune responses. Plant-borne cytokinins systemically induce resistance against pathogen infection. This resistance is orchestrated by endogenous cytokinin and salicylic acid signaling. Here, we discuss how plant- and pathogen-derived cytokinins inversely affect the plant defense response. In addition, we consider the molecular mechanisms underlying plant-derived cytokinin action in plant immunity.     

细胞分裂素对植物衰老的延缓作用

细胞分裂素是一类重要的植物激素,它可在一定程度上延缓植物的衰老。主要从3个方面综述了细胞分裂素与植物衰老之间的关系,即:(1)植物衰老过程中内源细胞分裂素含量变化;(2)外源细胞分裂素的影响;(3)转入与细胞分裂素的合成、降解相关的基因对植物衰老产生的影响。此外,还从细胞分裂素与糖、与脂质氧化反应以及与其它植物激素的关系方面探讨了细胞分裂素在延缓植物衰老中的作用机理。     

Moss biology and phytohormones--cytokinins in Physcomitrella

Mosses present several advantages for the analysis of phytohormone physiology. Their enormous regeneration capacity, the possibility of controlling their whole life cycle under in vitro culture conditions, as well as the small number of cell types facilitate studies of hormone homeostasis. This review focuses on the metabolism and biosynthesis of cytokinins, mostly summarising data obtained using the moss Physcomitrella patens (Hedw.) B.S.G. which has served as a model system for cytokinin research for many years. A comparison of metabolic differences with respect to seed plants is presented, pointing out an important role of adenosine kinase for the formation of nucleotides during cytokinin interconversion in Physcomitrella. Results on cytokinin biosynthesis in Physcomitrella are summarised with respect to the OVE mutants, which can be considered unique in the plant kingdom due to their strong overproduction of cytokinins. The OVE phenotype is correlated with both increased activity in early stages of cytokinin biosynthesis as well as increased conversion of cytokinin riboside to the base. Cytokinin interconverting reactions can contribute to the increased levels of cytokinins in OVE mutants. Further studies on hormone physiology in moss will help to complete our understanding of hormonal homeostasis by elucidating the situation in an evolutionary early embryophyte.     

Regulation of cytokinin content in plant cells

Cytokinin levels in plant cells are dependent on cytokinin biosynthesis and/or uptake from extracellular sources, metabolic interconversions, inactivation and degradation. Cytokinin conversion to compounds differing in polarity seems to be decisive for their entrapment within the cell and intracellular compartmentation, which affects their metabolic stability. Increase in cytokinin levels, resulting either from their uptake or intracellular biosynthesis, may promote further autoinductive accumulation of cytokinins which may function in the induction of cytokinin-initiated physiological processes. Accumulated cytokinins are capable of inducing cytokinin oxidase which consequently decreases cytokinin levels. This seems to be the mechanism of re-establishment and maintenance of cytokinin homeostasis required for further development of physiological events induced by transient cytokinin accumulation. Auxin may influence cytokinin levels by down regulation of cytokinin biosynthesis and/or by promotion of cytokinin degradation. A model of the regulation of cytokinin levels in plant cells based on these phenomena is presented and its physiological role(s) is discussed.     

Cytokinin synthesis is higher in the Arabidopsis amp1 mutant

Cytokinins are involved in plant cell proliferation leading to plant growth and morphogenesis. Earlier we described a mutant of Arabidopsis thaliana, amp1, that had five times higher levels of cytokinin and had a number of pleiotropic phenotypes, including increased cell proliferation and de-etiolated growth in the dark. While these phenotypes were correlated with higher levels of cytokinin, the actual mechanism of how cytokinin is elevated was not elucidated before. In order to understand if the increased cytokinin is a result of increased biosynthesis or decreased degradation we have compared the synthesis of cytokinins from radiolabelled adenine and the degradation of zeatin ribosides and other cytokinins between amp1 and wild type plants. The degradation of the hormone is not affected in the mutant but there is a 4 to 6 fold increase in cytokinin synthesis compared to the wild type. Because the amp1 mutant is recessive we hypothesise that the AMP1 product negatively regulates cytokinin production.     

Identification of plant cytokinin biosynthetic enzymes as dimethylallyl diphosphate:ATP/ADP isopentenyltransferases

It has been believed that the key step in cytokinin biosynthesis is the addition of a 5-carbon chain to the N(6) of AMP. To identify cytokinin biosynthesis enzymes that catalyze the formation of the isopentenyl side chain of cytokinins, the Arabidopsis genomic sequence was searched for genes that could code for isopentenyltransferases. This resulted in the identification of nine putative genes for isopentenyltransferases. One of these, AtIPT4, was subjected to detailed analysis. Overexpression of AtIPT4 caused cytokinin-independent shoot formation on calli. As shoot formation on calli normally occurs only when cytokinins are applied, it suggested that this gene product catalyzed cytokinin biosynthesis in plants. Recombinant AtIPT4 catalyzed the transfer of an isopentenyl group from dimethylallyl diphosphate to the N(6) of ATP and ADP, but not to that of AMP. AtIPT4 did not exhibit the DMAPP:tRNA isopentenyltransferase activity. These results indicate that cytokinins are, at least in part, synthesized from ATP and ADP in plants.     

Evolution of cytokinin biosynthesis and degradation

Cytokinin hormones are important regulators of development and environmental responses of plants that execute their action via the molecular machinery of signal perception and transduction. The limiting step of the whole process is the availability of the hormone in suitable concentrations in the right place and at the right time to interact with the specific receptor. Hence, the hormone concentrations in individual tissues, cells, and organelles must be properly maintained by biosynthetic and metabolic enzymes. Although there are merely two active cytokinins, isopentenyladenine and its hydroxylated derivative zeatin, a variety of conjugates they may form and the number of enzymes/isozymes with varying substrate specificity involved in their biosynthesis and conversion gives the plant a variety of tools for fine tuning of the hormone level. Recent genome-wide studies revealed the existence of the respective coding genes and gene families in plants and in some bacteria. This review summarizes present knowledge on the enzymes that synthesize cytokinins, form cytokinin conjugates, and carry out irreversible elimination of the hormones, including their phylogenetic analysis and possible variations in different organisms.     

Role of cytokinins in stress resistance of plants

The facts of both positive and negative influences of cytokinins on stress resistance of plants are known today. Without pretending to a final choice between these points of view, we have made an attempt to analyze the details of the experiments that gave rise to conclusions about the nature of the effect of cytokinins on the resistance to stress-causing influences with a focus on their intensity and duration. The review deals with the data concerning the influence of different adverse factors on the content of endogenous cytokinins and transduction of cytokinin signals, examines the influence on plant resistance of treatment with exogenous hormone, and the effects of genetic modifications causing changes in cytokinin content and signaling. Resistance is considered not only as a mean of plant survival under severe stress but also as an instrument of maintaining growth rate in plants exposed to moderate stress. Literature data and our own results make it possible to conclude that cytokinins play an important role in formation of plant resistance to adverse influences; however, the effect of these hormones depends on stress intensity. Under moderate stress, cytokinins ensure maintenance of plant growth, whereas a drop in cytokinins hampers growth under a strong influence of adverse factors, which is a prerequisite for mobilization of limited resources characteristic of severe stress and ensures preservation of plant viability.