Cytokinins are structurally diverse and biologically versatile. The chemistry and physiology of cytokinin have been studied extensively, but the regulation of cytokinin biosynthesis, metabolism, and signal transduction is still largely undefined. Recent advances in cloning metabolic genes and identifying putative receptors portend more rapid progress based on molecular techniques. This review centers on cytokinin metabolism with connecting discussions on biosynthesis and signal transduction. Important findings are summarized with emphasis on metabolic enzymes and genes. Based on the information generated to date, implications and future research directions are presented.

trans-Zeatin is a major and ubiquitous cytokinin in higher plants. cis-Zeatin has traditionally been viewed as an adjunct with low activity and rare occurrence. Recent reports of cis-zeatin and its derivatives as the predominant cytokinin components in some plant tissues may call for a different perspective on cis-isomers. The existence of a maize (Zea mays) gene (cisZOG1) encoding an O-glucosyltransferase specific to cis-zeatin (R.C. Martin, M.C. Mok, J.E. Habben, D.W.S. Proc Natl Acad Sci USA 98: 5922-5926) lends further support to this view. Results described here include the isolation of a second maize cisZOG gene, differential expression of cisZOG1 and cisZOG2, and identification of substantial amounts of cis-isomers in maize tissues. The open reading frame of cisZOG2 has 98.3% identity to cisZOG1 at the nucleotide level and 97.8% at the amino acid level. The upstream regions contain common and unique segments. The recombinant enzymes have similar properties, K m values of 46 and 96 m, respectively, for cis-zeatin and a pH optimum of 7.5. Other cytokinins, including N 6 -(⌬ 2 -isopentenyl)adenine, trans-zeatin, benzyladenine, kinetin, and thidiazuron inhibited the reaction. Expression of cisZOG1 was high in maize roots and kernels, whereas cisZOG2 expression was high in roots but low in kernels. cis-Zeatin, cis-zeatin riboside, and their O-glucosides were detected in all maize tissues, with immature kernels containing very high levels of the O-glucoside of cis-zeatin riboside. The results are a clear indication that O-glucosylation of cis-zeatin is a natural metabolic process in maize. Whether cis-zeatin serves as a precursor to the active trans-isomer or has any other unique function remains to be demonstrated.Cytokinins are plant hormones regulating cell division and a range of developmental events such as bud formation, leaf expansion, senescence, seed germination, and chloroplast formation (Mok, 1994). trans-Zeatin is a major and ubiquitous cytokinin in higher plants. Earlier cytokinin analyses detected ciszeatin and its derivatives in trace amounts in some plants, but due to their low activity (Schmitz et al., 1972), cis-isomers were viewed as adjunct to transisomers. Recent analyses, however, showed that the cis-isomers can be the dominant cytokinins at particular stages of development in plants such as chickpea (Cicer arietinum) and lupine (Lupinus albus; Emery et al., 1998Emery et al., , 2000. Moreover, the presence of cis-isomers was associated with male sterility in Mercurialis spp. flowers (Louis et al., 1990;Durand and Durand, 1994). These are indications that cis-isomers may have unique physiological functions. The ability to regulate the levels of cis-zeatin is evidenced by the maize (Zea mays) cisZOG1 gene, encoding an O-glucosyltransferase with specificity to cis-zeatin (Martin et al., 2001).O-Glucosylation is a major step in the metabolism of trans-zeatin ). The resulting O-glucosides seem to serve as storage compounds and are resistant to degradation by cytokinin oxidases (Armstrong, 1994...

Zeatin is the most active and ubiquitous of the naturally occurring cytokinins. The O-glucoside of zeatin, found in all plants examined, is considered to be important in cytokinin transport, storage, and protection against cytokinin

Zeatin is a naturally occurring cytokinin. Biosynthesis and metabolism studies of zeatin have been directed mostly at the trans isomer, although cis-zeatin and its riboside occur as major components in some plant species. It is not known whether parallel regulatory pathways exist for the two isomers. Based on the sequence of the gene ZOG1 encoding a trans-zeatin O-glucosyltransferase from Phaseolus (EC 2.4.1.203), a cis-zeatin-specific Oglucosyltransferase was isolated from maize. This gene, cisZOG1, contains an ORF of 1,401 nucleotides encoding a protein of 51.1 kDa with 41% identity to the Phaseolus ZOG1 protein. Unexpectedly, the maize enzyme recognizes as substrates cis-zeatin and UDP-glucose but not cis-ribosylzeatin, trans-zeatin, or trans-ribosylzeatin. This finding indicates the existence of cis-specific regulatory elements in plants and suggests that cis-zeatin and derivatives may be more important in cytokinin homeostasis than currently recognized. C ytokinins are essential hormones for plant growth. In addition to promoting cell division and differentiation in tissue culture, cytokinins regulate a host of developmental events in whole plants such as bud formation, leaf expansion, delay of senescence, promotion of seed germination, and chloroplast formation (reviewed in ref. 1). Naturally occurring cytokinins are adenine derivatives with a side chain at the N 6 -position; cytokinins with a hydroxylated isoprenoid side chain such as trans-zeatin, first identified in maize (2), are major constituents in plants. Studies on the biosynthesis and metabolism of cytokinins have centered on trans-zeatin, although cis-zeatin and its derivatives have been isolated from a number of species including potato (3, 4), Mercurialis (5), hops (6), rice (7-9), wheat (10), oats (10), and chickpeas (11). This emphasis on the trans isomers may be attributed to their higher biological activity.Two pathways for cytokinin biosynthesis have been proposed. The first is the direct pathway, involving formation of N 6 -isopentenyladenosine monophosphate from AMP and dimethylallyl pyrophosphate, followed by hydroxylation of the side chain to form trans-zeatin-type cytokinins. In support of this pathway, isopentenyltransferase activity in tobacco (12) and hydroxylation activity in cauliflower (13) have been reported. The discovery of the Agrobacterium ipt gene (14, 15) lends support to the existence of a similar biosynthetic pathway in plants. However, plant genes encoding such enzymes have not been identified, and indirect evidence suggests the existence of a biosynthetic pathway for zeatin without isopentenyl intermediates (16, 17). The second, indirect pathway involves release of cytokinins by turnover of tRNAs containing cis-zeatin. Because cis-zeatin is much less active in bioassays, conversion to the trans isomer by enzymes such as the cis-trans isomerase of zeatin (18) Here we report the isolation and characterization of a gene encoding a cis-zeatin specific O-glucosyltransferase from maize. The finding of a gene and enzyme sp...

lnvestigation of the conversion of exogenous cis-zeatin to transzeatin in immature seeds of Phaseolus vulgaris 1. led to the isolation of a cis-trans-isomerase from the endosperm. lhe enzyme was purified more than 2000-fold by chromatography on a series of fast protein liquid chromatography (anion exchange, gel filtration, and hydrophobic interaction) and concanavalin A columns. l h e enzymic reaction favors conversion from the cis to the trans form and requires flavin, light, and dithiothreitol. ris-Zeatin riboside is also a substrate for the enzyme. Retention on the concanavalin A column indicated that the enzyme is a glycoprotein. lhe enzyme was stable for at least 8 weeks when stored at -80°C. l h e occurrence of ris-trans-isomerization suggests that cis-zeatin and ciszeatin riboside formed by tRNA degradation could be precursors of biologically adive cytokinins.

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SummaryCytokinins 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.