Showing papers on "Zeatin published in 2000"

A New Family of High-Affinity Transporters for Adenine, Cytosine, and Purine Derivatives in Arabidopsis

Abstract: In many organisms, including plants, nucleic acid bases and derivatives such as caffeine are transported across the plasma membrane. Cytokinins, important hormones structurally related to adenine, are produced mainly in root apices, from where they are translocated to shoots to control a multitude of physiological processes. Complementation of a yeast mutant deficient in adenine uptake (fcy2) with an Arabidopsis cDNA expression library enabled the identification of a gene, AtPUP1 (for Arabidopsis thaliana purine permease1), belonging to a large gene family (AtPUP1 to AtPUP15) encoding a new class of small, integral membrane proteins. AtPUP1 transports adenine and cytosine with high affinity. Uptake is energy dependent, occurs against a concentration gradient, and is sensitive to protonophores, potentially indicating secondary active transport. Competition studies show that purine derivatives (e.g., hypoxanthine), phytohormones (e.g., zeatin and kinetin), and alkaloids (e.g., caffeine) are potent inhibitors of adenine and cytosine uptake. Inhibition by cytokinins is competitive (competitive inhibition constant K(i) = 20 to 35 microM), indicating that cytokinins are transported by this system. AtPUP1 is expressed in all organs except roots, indicating that the gene encodes an uptake system for root-derived nucleic acid base derivatives in shoots or that it exports nucleic acid base analogs from shoots by way of the phloem. The other family members may have different affinities for nucleic acid bases, perhaps functioning as transporters for nucleosides, nucleotides, and their derivatives.

Increased cytokinin levels in transgenic PSAG12–IPT tobacco plants have large direct and indirect effects on leaf senescence, photosynthesis and N partitioning

Abstract: We studied the impact of delayed leaf senescence on the functioning of plants growing under conditions of nitrogen remobilization. Interactions between cytokinin metabolism, Rubisco and protein levels, photosynthesis and plant nitrogen partitioning were studied in transgenic tobacco (Nicotiana tabacum L.) plants showing delayed leaf senescence through a novel type of enhanced cytokinin synthesis, i.e. targeted to senescing leaves and negatively auto-regulated (PSAG12‐IPT), thus preventing developmental abnormalities. Plants were grown with growthlimiting nitrogen supply. Compared to the wild-type, endogenous levels of free zeatin (Z)- and Z riboside (ZR)type cytokinins were increased up to 15-fold (total ZR up to 100-fold) in senescing leaves, and twofold in younger leaves of PSAG12‐IPT. In these plants, the senescenceassociated declines in N, protein and Rubisco levels and photosynthesis rates were delayed. Senescing leaves accumulated more ( 15 N-labelled) N than younger leaves, associated with reduced shoot N accumulation (‐60%) and a partially inverted canopy N profile in PSAG12‐IPT plants. While root N accumulation was not affected, N translocation to non-senescing leaves was progressively reduced. We discuss potential consequences of these modified sink‐source relations, associated with delayed leaf senescence, for plant productivity and the efficiency of utilization of light and minerals.

Inoculation and nitrate alter phytohormone levels in soybean roots: differences between a supernodulating mutant and the wild type.

Abstract: The levels of different cytokinins. indole-3-acetic acid (IAA) and abscisic acid (ABA) in roots of Glycine max [L.] Merr. cv. Bragg and its supermodulating mutant nts382 were compared for the first time. Forty-eight hours after inoculation with Bradyrhizobium, quantitative and qualitative differences were found in the root's endogenous hormone status between cultivar Bragg and the mutant nts382. The six quantified cytokinins, ranking similarly in each genotype, were present at higher concentrations (30-196% on average for isopentenyl adenosine and dihydrozeatin riboside, respectively) in mutant roots. By contrast, the ABA content was 2-fold higher in Bragg, while the basal levels of IAA [0.53 mu mol (g DW)(-1), on average] were similar in both genotypes. In 1 mM NO3--fed Bragg roots 48 h post-inoculation, IAA, ABA and the cytokinins isopentenyl adenine, and isopentenyl adenosine quantitatively increased with respect to uninoculated controls. However, only the two cytokinins increased in the mutant. High NO3- (8 mM) markedly reduced root auxin concentration, and neither genotypic differences nor the inoculation-induced increase in auxin concentration in Bragg was observed under these conditions. Cytokinins and ABA, on the other hand, Were little affected by 8 mM NO3-. Root IAA/cytokinin and ABA/cytokinin ratios were always higher in Bragg relative to the mutant, and responded to inoculation (mainly in Bragg) and nitrate (both genotypes). The overall results are consistent with the auxin-burst-control hypothesis for the explanation of autoregulation and supernodulation in soybean. However, they are still inconclusive with respect to the inhibitory effect of NO3-.

Effects of Elevated [CO2] and Nitrogen Nutrition on Cytokinins in the Xylem Sap and Leaves of Cotton

Abstract: We measured the level of xylem-derived cytokinins (CKs) entering a cotton leaf, and the CK levels in the same leaf, thus enabling xylem sap and foliar CKs to be compared concurrently. Although zeatin was the dominant CK in xylem sap, zeatin, dihydrozeatin, and N6-(2-isopentenyl) adenine were present in approximately equimolar levels in leaves. Elevated [CO2] (EC) has an effect on the levels of cytokinins in sap and leaf tissues. This effect was modulated by the two levels of root nitrogen nutrition (2 and 12 mm nitrate). Growth enhancement (70%) in EC over plants in ambient [CO2] (AC) was observed for both nitrogen nutrition treatments. Low-nitrogen leaves growing in EC exhibited photosynthetic acclimation, whereas there was no sign of photosynthetic acclimation in high-nitrogen grown leaves. Under these prevailing conditions, xylem sap and leaf tissues were obtained for CK analysis. Higher nitrogen nutrition increased the delivery per unit leaf area of CKs to the leaf at AC. EC caused a greater increase in CK delivery to the leaf at low nitrogen conditions (106%) than at high nitrogen conditions (17%). EC induced a significant increase in CK content in low-nitrogen leaves, whereas CK content in leaf tissues was similar for high-nitrogen leaves growing in AC and EC.

Cytokinins: biosynthesis, metabolism and perception.

Abstract: 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.

Facultative and obligate aerobic methylobacteria synthesize cytokinins

Abstract: The presence and expression of genes controlling the synthesis and secretion of cytokinins by the pink-pigmented facultative methylotroph Methylobacterium mesophilicum VKM B-2143 with the serine pathway and nonpigmented obligate methylotroph Methylovorus mays VKM B-2221 with the ribulose monophosphate pathway of C1 metabolism were shown using the polymerase chain reaction (PCR) and reverse transcription–PCR methods. The presence of the corresponding mRNA in M. mesophilicum cells grown on methanol or succinate suggests that the expression of these genes is constitutive. The cytokinin activity of culture liquid and its fractions was determined by a biotest with Amarantus caudatus L. seedlings. Using enzyme-linked immunosorbent analysis, we detected zeatin (riboside) in the culture liquid of both bacteria studied. The data obtained show that the aerobic methylobacteria are phytosymbionts that are able to utilize the single- and polycarbon compounds secreted by symbiotic plants and to synthesize cytokinins.

Transgenic tobacco plants co‐expressing Agrobacterium iaa and ipt genes have wild‐type hormone levels but display both auxin‐ and cytokinin‐overproducing phenotypes

Abstract: Transgenic tobacco lines simultaneously expressing the Agrobacterium iaaM, iaaH and ipt genes, obtained by crossing lines expressing ipt with lines expressing iaaM and iaaH, were used to study in planta interactions between auxin and cytokinins. All phenotypic traits of the respective parental lines characteristic of cytokinin and auxin overproduction were present in the cross. Indole-3-acetic acid (IAA) and combined zeatin riboside (ZR) and zeatin riboside-5'-monophosphate (ZRMP) contents were analysed by mass spectrometry in young, developing leaves from the cross, the parental lines and the wild type. Unexpectedly, hormone levels in the cross were very similar to wild-type levels. Thus IAA levels in the cross were much lower throughout vegetative development than in the parental IAA overproducing line, although expression of the bacterial IAA biosynthesis genes was not reduced. The results suggest that effects on apical dominance, adventitious root formation, leaf morphology and other traits commonly +/- associated with IAA and cytokinin overproduction, and observed in the iaa E ipt cross, cannot be explained solely by analysis of auxin and cytokinin contents in individual organs. As traits associated with both hormones are expressed in close spatial and temporal proximity, it is likely that cellular resolution of hormone contents is essential to explain physiological responses to auxins and cytokinins.

Influence of the antibiotic timentin on plant regeneration of tomato (Lycopersicon esculentum Mill.) cultivars.

Abstract: Cotyledon explants of tomato (Lycopersicon esculentum Mill. cvs 'Santa Clara', 'Firme' mutant, 'IPA-5' and 'IPA-6') were excised from 8- to 10-day-old in vitro-grown seedlings. Four different shoot induction media supplemented with timentin (300 mg l–1) were screened. When cotyledon explants were cultured on MS-based medium with 1.0 mg l–1 zeatin plus 0.1 mg l–1 IAA and supplemented with timentin, higher regeneration frequencies and a greater number of elongated shoots were obtained. It was observed that timentin caused an increase in the morphogenesis of in vitro cotyledon explants of tomato cultivars. In two of three cultivars tested, rooting of shoots was positively influenced, both in the presence and absence of timentin in the rooting medium, among shoots regenerated from explants derived from timentin-supplemented medium. The results confirm those of a previous investigation on the beneficial effects of this class of antibiotics on tomato regeneration and, consequently, its reliability for use in the transformation of this species.

Changes in cis‐zeatin and cis‐zeatin riboside levels and biological activity during potato tuber dormancy

Abstract: The effects of postharvest storage duration and temperature on endogenous cis-zeatin (cis-Z) and cis-zeatin riboside (cis-ZR) levels in potato (Solanum tuberosum L.) tubers were determined in relation to tuber bud dormancy. The tubers used in these studies were completely dormant for at least 81 days of storage. Thereafter, tuber bud dormancy diminished gradually and after 165 days of postharvest storage, the tubers were completely non-dormant. Immediately after harvest, endogenous levels of cis-Z and cis-ZR were approximately 25 pmol (g fresh weight)−1 and 8 pmol (g fresh weight)−1, respectively. In tubers exiting dormancy but stored at a growth-inhibiting temperature (3°C), endogenous levels of cis-Z rose over threefold after 25 days of storage and remained elevated for the duration of the study. Levels of cis-ZR remained essentially constant during this same period. In tubers transferred to a growth permissive temperature (20°C) prior to use, the rise in endogenous cis-Z was less dramatic and more protracted; increasing twofold after 53 days of storage. No change in cis-Z riboside content was observed in these tubers during this period. Dose-response studies using either cis-Z or trans-Z demonstrated a time-dependent increase in cytokinin sensitivity during postharvest storage. Immediately after harvest, dormant tubers were insensitive to both zeatin isomers. Thereafter, tubers exhibited a dose-dependent increase in premature sprouting following injection with either cytokinin isomer. After injection into dormant tubers, cis-[8-14C]-zeatin was metabolized primarily to adenine/adenosine and cis-Z riboside. Seven days after injection, less than 10% of the recovered radioactivity was associated with trans-ZR. These results are consistent with a role for endogenous cis-Z (and its derivatives) in the regulation of potato tuber dormancy.

Direct somatic embryogenesis, plant regeneration and in vitro flowering in rapid-cycling Brassica napus.

Abstract: A simple method to induce somatic embryogenesis from seeds of rapid-cycling Brassica napus is described. Seedlings cultured on Murashige and Skoog (MS) basal medium produced somatic embryos directly on hypocotyls and cotyledons after 2 to 3 subcultures onto the same medium. A low pH of the medium (3.5–5) was more conducive to somatic embryogenesis than a higher pH (6 and 7). Embryogenic potential of the seeds was inversely correlated to seed age: about 41–68% of immature seeds between the ages of 14 and 28 days after pollination (DAP) formed somatic embryos compared to 0–11% of the seeds obtained 29–37 DAP. About 54% of the somatic embryos produced secondary embryos after subculturing onto the same medium. The embryogenic potential of the cultures has been maintained on MS basal medium for 2 years (12 generations) without diminution. Up to 75% of the secondary embryos developed into plantlets on MS medium enriched with 10–6 M zeatin, and 40% of these produced flowers when transferred to an optimised flower-induction medium. Viable seeds were produced in self-pollinated in vitro flowers.

Somatic embryogenesis in the endemic black iris

Abstract: Somatic embryogenesis was achieved from callus, cell suspension and protoplast culture systems in the endemic black iris (Iris nigricans). Subculture of friable callus fragments on embryogenesis induction medium (EIM) containing 4.5 μM 2,4-dichlorophenoxyacetic acid (2,4-D), 0.5 μM kinetin, 4.5 μM 1-naphthaleneacetic acid (NAA) and 300 mg l-1 proline in the dark was necessary before transfer to regeneration medium (RM). Regeneration was studied by transferring friable callus fragments from EIM to RM containing (0.0, 4.5, 9.0, 13.5 μM) of either 6-benzyladenine (BA), 2-isopentenyladenine (2iP), zeatin or thidiazuron (TDZ) in combination with 0.49 μM indole-3-butyric acid (IBA), 0.45 μM 2,4-D. Maximum embryogenesis was obtained at 4.5 μM BA while zeatin and TDZ were not effective and embryogenesis did not occur with these treatments. Sucrose at 0.2 M was more effective for embryogenesis when compared to glucose or fructose. Growing cells in suspension culture on EIM containing 4.5 μM 2,4-D in combination with 0.2 M sucrose for four weeks and transferring cells to RM (containing 4.5 μM BA) gave significant embryogenesis with maximum number of embryos (3568 embryos/g cells). Using 4.5 μM 2,4-D in protoplast culture was necessary for the best protoplast division and colony formation. In all experiments, embryos developed on RM were transferred to hormone-free medium (HFM) and 90% converted to rooted plantlets. Produced plantlets gave 95% survival ex vitro. Plantlets developed to whole plants in the greenhouse and flowered.

In vitro adventitious bud regeneration from internode segments of beech

Abstract: Internode explants collected from in vitro grown shoots of two clones of Fagus sylvatica L. (European beech) and five clones of F. orientalis Lipski (Oriental beech) were used to evaluate their bud regeneration capacity. Adventitious shoot-buds formed on callus, which developed from internode segments cultured in a Woody Plant Medium supplemented with different concentrations of either thidiazuron (TDZ) or benzyladenine (BA). After 4 weeks of culture on induction media, the explants were transferred to a proliferation medium supplemented with 2.2 μM BA, 9.1 μM zeatin and 2.9 μM indole-3-acetic acid (IAA) for another 8 weeks. Medium containing TDZ was much more efficient than medium containing BA in inducing adventitious buds, the optimal TDZ concentration being 4.5 μM and the optimal BA concentration 17.8 μM. Genotypic variation in shoot regeneration capacity was observed among the two Fagus species and between clones within each species, with a significant interaction between TDZ concentration and genotype regarding mean bud number. Thidiazuron induction medium supplemented with a range of individual auxins was investigated, and it was found that IAA or indole-3-butyric acid at 2.9 μM enhanced the bud forming capacity of explants. Morphogenic response varied significantly with the position of the internode along the stem. The highest regeneration potential was obtained from apical internodes, while those distal to the apex were the least productive. Elongated shoots of adventitious origin can be readily proliferated by axillary branching.

Cytokinin synthesis is higher in the Arabidopsis amp1 mutant

Abstract: 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.

Inhibition of carnation petal inrolling by growth retardants and cytokinins

Abstract: Excised carnation petals induced to senescence by ethrel (an ethylene-releasing compound) exhibited morphological changes that closely resembled those of senescing petalsin situ in cut flowers. The sensitivity of the excised petals to ethylene was reduced by exogenous cytokinin and this type of hormonal interaction in the control of plant development is discussed. Using the excised petals, a number of known and potential growth inhibitors were compared for ability to prevent petal inrolling induced by ethrel. Cycloheximide and 6-methylpurine were the most effective and inhibited inrolling almost completely, but purine, purine riboside, lauric acid, L-azetidine-2-carboxylic acid and n-decyl alcohol were also very effective. All these compounds were considerably more effective than any cytokinin tested. When supplied through the transpiration stream to short-stemmed carnations, cycloheximide, 6-methylpurine and purine inhibited inrolling nearly completely and the flowers finally senesced by water loss. 6-Methylpurine inhibited ethylene production in cut flowers and RNA synthesis in excised petals very markedly. Degradation of exogenous zeatin riboside by cytokinin oxidase, and the level of activity of the enzyme in petals, were reduced by 6-methylpurine. These biochemical changes probably account for the strong inhibition of inrolling induced by this compound.

Regeneration of plantlets from leaf and petiole explants of Pelargonium x hortorum

Abstract: The in vitro plant regeneration potential of vegetatively propagated geraniums (Pelargonium x hortorum) has been investigated. Using various combinations of growth regulators and a choice of different explants, a regeneration protocol has been developed to raise in vitro plantlets from young petiole and leaf explants from three different cultivars of geraniums. In all three cultivars, very young petiole explants exhibited a higher regeneration potential as compared with leaf explants. Regeneration efficiencies were found to be highly dependent on the cultivar, with cv. Samba showing the highest regeneration potential, followed by cvs. Yours Truly and then Sincerity. Samba also showed the highest number of shoots from both the petiole [57 shoot buds per petiole explant in the presence of 3 μM zeatin and 1 μM indole-3-acetic acid (IAA) and leaf explants (43 shoots per leaf explant with 10 μM zeatin and 2 μM IAA). Shoot buds transferred to Murashige and Skoog (MS) medium supplemented with 0.44 μM N6-benzyladenine and 0.11 μM IAA grew vigorously and attained 1–2 cm in length in 3–4 wk. These shoots rooted with 100% efficiency on MS basal medium, and plants developed that showed normal growth and flowering under greenhouse conditions.

Light Mediates Endogenous Plant Growth Substances in Thidiazuron-induced Somatic Embryogenesis in Geranium Hypocotyl Cultures

Abstract: Thidiazuron-induced somatic embryogenesis in geranium (Pelargonium x hortorum Bailey) was readily achieved under both light and darkness. The hypocotyl explants treated with TDZ formed well differentiated embryos when incubated under conditions of continuous light, complete darkness or 16-h photoperiod. However, embryogenesis was significantly impaired by light at both 16-h and continuous light exposure. Maintaining the cultures in the dark increased the frequency of the somatic embryos formed with a corresponding moderate elevation of the endogenous plant growth substances measured. In the complete absence of TDZ, all of the hypocotyl explants cultured in the dark remained thin but formed etiolated roots. The number of explants that formed roots decreased with increasing light exposure time; in addition the roots formed were shorter in length. The levels of endogenous adenine, adenosine, DHZ, zeatin, isopentenyladenine (2iP), tryptamine, IAA and ABA remained elevated for the first 2 days of culture in explants maintained on MSO and under continuous light, while they decreased on day 3 of culture. We provide evidence that the interaction between TDZ and light treatments modulated the endogenous plant growth substances, which in turn affected the embryogenic process in geranium hypocotyl explants.

Genes encoding zeatin O-glycosyltransferases.

Abstract: Zeatin is a highly active and ubiquitous cytokinin. O-Glycosylation of zeatin occurs widely in plant tissues and is likely to be important in regulating the level of active cytokinins. Enzymes mediating the conversion of zeatin to O-glucosylzeatin and O-xylosylzeatin have been isolated from Phaseolus seeds. The corresponding genes have been cloned recently. The current status of molecular and biochemical studies of these genes and enzymes is summarised in this paper.

Accumulation of zeatin O-glycosyltransferase in Phaseolus vulgaris and Zea mays following cold stress

Abstract: Zeatin O-glycosides have been reported as inactive and stable storage forms of cytokinins whose concentrations increase in cold stressed plants. Zeatin O-glycosides accumulation in developing bean seeds has been correlated with an increase of zeatin O-glycosyltransferase , which is specific to trans-zeatin, and catalyzes the conjugation of zeatin O-glycosides. When Phaseolus vulgaris and Zea mays seedlings were grown for 3 days at 25 and then incubated at 4 or 10 for 6 days no further growth was observed in roots. Hypertrophy was observed in the root tips of both species. In shoot-hypocotyl complexes, in contrast, growth occurred when seedlings were incubated at 10 . Western analysis, with Mabs specific to zeatin O-glycosyltransferase, detected antigenically related proteins in roots, shoot tips and cotyledons after seedlings were cold stressed for 1–6 days at 4 or 10 . Immunolocalization, of both maize and bean root sections grown at 25 revealed antigenically related proteins that were detected at low levels in cortical cells. The signal intensified upon cold stress. The localization of zeatin O-glycosyltransferase in Z. mays root tips was directly comparable to the distribution of the zeatin O-glycosides. The enzyme was detected in the nucleus, cytoplasm, and closely associated with the plasma membrane and in the cell wall of Z. mays root cells. Southern analysis suggested that more than one gene in Z. mays that were homologous to zeatin O-glycosyltransferase in P. vulgaris. Zeatin O-glycosyltransferase may be involved in modulation of cytokinins under cold stress.

Deuterium in vivo labelling of cytokinins in Arabidopsis thaliana analysed by capillary liquid chromatography/frit-fast atom bombardment mass spectrometry

Abstract: A method was developed for analysing the biosynthetic rate of the cytokinin class of plant hormones. Transgenic, cytokinin-overproducing Arabidopsis thaliana plants were incubated in liquid culture media enriched with 30% deuterium oxide, and incorporation into the different parts of the cytokinin molecule was analysed by capillary liquid chromatography/frit-fast atom bombardment mass spectrometry after precolumn propionylation. The sugar moieties of the cytokinins generally showed a high and independent incorporation, so the analysis in this study focused on the cytokinin base moieties. It was observed that during a 24 h incubation period almost all labelling was incorporated into the side-chain, rather than the adenine moiety. The incorporation dynamics of isopentenyladenosine-5'-monophosphate, zeatinriboside-5'-monophosphate (ZRMP) and zeatin-9-glucoside were investigated through analysis of the cytokinin base fragments in high-resolution selective ion monitoring mode. Using a fractional synthetic rate approach, the biosynthetic rate of ZRMP was determined to be 18 ng h(-1) g(-1) fresh weight, giving a turnover time of 25 h. A method for the mass isotopomer abundance analysis of the cytokinins in the zeatin family, based on selective reaction monitoring, was also developed to gain further sensitivity. Use of this technique showed that there was a higher level of enrichment in zeatin nucleotide than in the corresponding nucleoside, in agreement with the hypothesis that cytokinin nucleotides are primary products in this pathway.

Identification and quantification of the major endogenous cytokinins in pistachio seedlings

Abstract: The major endogenous cytokinins, Z, ZR, DHZ, DHZR, iP and iPR in pistachio seedlings (Pistacia vera L. cv. Ohadi) were purified by HPLC and their identities confirmed using GC-MS. The aerial parts of two-year old pistachio seedlings including mature leaves, young leaves, lateral buds, debarked stems and bark were subjected to analysis. All of the above mentioned cytokinins were identified in the aerial parts except DHZ which was only present in mature leaves. Z-type cytokinins contributed almost 43% of the total cytokinins. ZR and DHZR were identified as the major ribosides and iP as the main base. The greatest concentration of ZR was detected in the bark, amounting to about 48%. DHZR and ZR constituted the major portion of the total cytokinins detected in both young and mature leaves while Z was detected as a minor cytokinin in leaves. The sharp increase of iP concentration during leaf maturation indicates that mature leaves are probably capable of de novo biosynthesis of cytokinins. The absence of DHZ (except in mature leaves) and the presence of considerable concentrations of DHZR in pistachio stems suggest that these tissues are able to metabolize DHZ to DHZR. The large amount of ZR in pistachio leaves suggests that root-derived ZR is transported into the leaves after loading into the xylem. The presence of high amounts of iP in pistachio lateral buds indicates that iP has been accumulated in these parts. The occurrence of a totally different cytokinin distribution pattern in buds, as compared with the other aerial parts, possibly results from their different metabolism.