Cytokinin-Mediated Translational Control of Protein Synthesis in Cultured Cells of Glycine max

Polyribosome formation was stimulated by cytokinin treatmentof cultured cells of Glycine max cv. Funk Delicious. When suspensioncultures were given 0·5 µM zeatin after 24 h inculture in medium lacking a cytokinin, a nearly 2-fold increasein the polyribosome/monoribosome ratio occurred over the subsequent3 h. The effect of actinomycin D and of 5-fluorouridine on RNAsynthesis and on the polyribosome/monoribosome ratios of thesecells was examined. Actinomycin D at 5 and 20 µg/ml^–1inhibitedtotal RNA synthesis by 39 and 60%, respectively, as measuredby [³H]uridine incorporation into acid-precipitable material.The degree of inhibition of precursor incorporation into polyribosomalRNA was similar. At 0·1 mM, 5-fluorouridine inhibited[³H]uridine incorporation by 76%, and [³H]guanosine incorporationby 66% into polyribosomal RNA after 3 h of treatment. Fractionationof the polyribosomal RNA by oligo(dT)-cellulose chromatographydemonstrated that low concentrations of both actinomycin D (5µg ml^–1) and 5-fluorouridine (0·1 mM) inhibitedthe synthesis of ribosomal RNA to a greater extent than thepoly(A)-containing fraction of the messenger RNA. Synthesisof the poly(A)-containing RNA was inhibited by 24% with 5µgml^–1 actinomycin D and by 30% with 0·1 mM 5-fluorouridine.At the above concentrations, these two inhibitors reduced thepolyribosome/monoribosome ratio of the cytokinin-deprived cellsover a 3 h period, but they did not prevent cytokinin-inducedpolyribosome formation. These results provide further evidencethat cytokinin regulates polyribosome levels through an effecton protein synthesis at the translational level     

Trimming and solubilization of xyloglucan after deposition in the walls of cultured rose cells

A pulse-chase technique involving the in vivo feeding of L-[1-³H]arabinoseto suspension-cultured rose (Rosa) cells at 4 d and 9 d aftersubculture (fast- and slow-growing, respectively) was used tocreate a population of [³H]xyloglucan molecules and to followtheir subsequent fate. The weight-average relative molecu larmass (M_w) of [³H]xyloglucan freshly deposited in the cell wallwas 160 000 and 240 000 in the fast- and slow-growing cells,respectively. The wall-bound [³H]xyloglucan of both culturesunderwent a decrease in M_w of 40 000 during the first 2 d afterthe pulse-labelling. At the same time, 20–30% of the initially-deposited[³H]xyloglucan disappeared from the cell wall, and a similaramount appeared in solution in the culture medium. Its failureto remain bound to the cell wall and its low M_w (39 000) indicatedthat this soluble extracellular ( was derived from partial degradationof segments of wall-bound xyloglucan that were not directlyhydrogen-bonded to microfibrils (‘loose ends’ and‘tethers’). The possible enzymic basis and biologicalroles of the degradation are discussed. Key words: Cell expansion, cell wall, hemicellulose, sloughing, xyloglucan     

Estradiol-17beta stimulates proliferation of mouse embryonic stem cells: involvement of MAPKs and CDKs as well as protooncogenes

Although the importance of estradiol-17 (E₂) in many physiological processes has been reported, to date no researchers have investigated the effects of E₂ on embryonic stem (ES) cell proliferation. Therefore, in the present study, we have examined the effect of E₂ on the DNA synthesis of murine ES (ES-E14TG2a) cells and its related signaling pathways. The results of this study show that E₂ (10^–9 M) significantly increased [³H]thymidine incorporation at >4 h and that E₂ (>10^–12 M) induced an increase of [³H]thymidine incorporation after 8-h incubation. Moreover, E₂ (>10^–12 M) also increased 5'-bromo-2'-deoxyuridine (BrdU) incorporation and cell number. Indeed, E₂ stimulated estrogen receptor (ER)- and - protein levels and increased mRNA expression levels of protooncogenes (c-fos, c-jun, and c-myc). Tamoxifen (antiestrogen) completely inhibited E₂-induced increases in [³H]thymidine incorporation. In addition, estradiol-6-O-carboxymethyl oxime-BSA (E₂-BSA; 10^–9 M) increased [³H]thymidine incorporation at >1 h, and E₂-BSA (>10^–12 M) increased [³H]thymidine incorporation after 1-h incubation. E₂-BSA-induced increase in BrdU incorporation also occurred in a dose-dependent manner. Tamoxifen had no effect on E₂-BSA-induced increase of [³H]thymidine incorporation. Also, E₂ and E₂-BSA displayed maximal phosphorylation of p44/42 MAPKs at 10 and 5 min, respectively. E₂ increased cyclins D1 and E as well as cyclin-dependent kinase (CDK)2 and CDK4. In contrast, E₂ decreased the levels of p21^cip1 and p27^kip1 (CDK-inhibitory proteins). Increases of these cell cycle regulators were blocked by 10^–5 M PD-98059 (MEK inhibitor). Moreover, E₂-induced increase of [³H]thymidine incorporation was inhibited by PD-98059 or butyrolactone I (CDK2 inhibitor). In conclusion, estradiol-17 stimulates the proliferation of murine ES cells, and this action is mediated by MAPKs, CDKs, or protooncogenes. cyclin-dependent kinase; mitogen-activated protein kinase     

Xyloglucan−pectin linkages are formed intra-protoplasmically,contribute to wall-assembly,and remain stable in the cell wall

We tested two hypotheses for the mechanism by which xyloglucan–pectin covalent bonds are formed in Arabidopsis cell cultures. Hypothesis 1 proposed hetero-transglycosylation, with xyloglucan as donor substrate and a rhamnogalacturonan-I (RG-I) side-chain as acceptor. We looked for enzyme activities that catalyse this reaction using α-(1→5)-l-[³H]arabino- or β-(1→4)-d-[³H]galacto-oligosaccharides as model acceptor substrates. The ³H-oligosaccharides were supplied (with or without added xyloglucans) to living Arabidopsis cell-cultures, permeabilised cells, cell-free extracts, or four authentic XTHs. No hetero-transglycosylation occurred. Therefore, we cannot support hypothesis 1. Hypothesis 2 proposed that some xyloglucan is manufactured de novo as a side-chain on RG-I. To test this, we pulse-labelled Arabidopsis cell-cultures with [³H]arabinose and monitored the radiolabelling of anionic (pectin-bonded) xyloglucan, which was resolved from free xyloglucan by ion-exchange chromatography. [³H]Xyloglucan–pectin complexes were detectable <4 min after [³H]arabinose feeding, which is shorter than the transit-time for polysaccharide secretion, indicating that xyloglucan–pectin bonds were formed intra-protoplasmically. Thereafter, the proportion of the wall-bound [³H]xyloglucan that was anionic remained almost constant at ∼50% for ≥6 days, showing that the xyloglucan–pectin bond was stable in vivo. Some [³H]xyloglucan was rapidly sloughed into the medium instead of becoming wall-bound. Only ∼30% of the sloughed [³H]xyloglucan was anionic, indicating that bonding to pectin promoted the integration of xyloglucan into the wall. We conclude that ∼50% of xyloglucan in cultured Arabidopsis cells is synthesised on a pectic primer, then secreted into the apoplast, where the xyloglucan–pectin bonds are stable and the pectic moiety aids wall-assembly.     

Specific Labeling of Cell Wall Polysaccharides with myo-[2-3H] Inositol during Germination and Growth of Phaseolus vulgaris L.

myo-[2-³H]Inositol was fed to bean seeds by imbibition and itsmetabolic fate was studied during germination and seedling growth.The largest amount of myo-inositol was taken up from a 500 HIMsupply (8 mg/seed) and the highest percentage was from 1 HIM(29%). myo-Inositol was incorporated to new cell wall polysaccharidesof hypocotyl and roots, mostly as uronic acid and pentose residues.In the 80% ethanolinsoluble cell walls of hypocotyls at 3, 4and 5 days after imbibition, 47 to 52% of ³H was detected asuronic acids, 20 to 24% as arabinose and 11 to 19% as xylose.Glucogenesis from myo-inositol was low: less than 6% was recoveredas hexoses. The ³H in uronic acid and arabinose residues decreasedwith increasing age (i.e. 0 to 6 cm from cotyledons) and increasedin older segments (further than 6 cm from cotyledons). In theoldest segment of 5-day-old hypocotyl (> 10 cm), ³H in thesugar residues was more than that in the youngest part (0–2cm). On the other hand, ³H in xylose residues increased steadilyin the older part, but did not exceed that in arabinose. The results show that the myo-inositol oxidation pathway functionsin growing hypocotyls and roots of bean seedlings to provideexclusively uronic acid and pentose units for cell wall synthesis.Results also show that incorporation of arabinose and uronicacids derived from myo-[2-³H]inositol to cell wall polysaccharidesis active in two regions of the hypocotyl; first, for the constructionof the primary walls in the young, growing region of the hypocotyl,and second, for thickening of the walls after completion ofelongation growth. ¹Supported by NSERC of Canada. (Received April 10, 1984; Accepted June 12, 1984)     

Effect of {alpha}-Tomatine on the Response of Wheat Coleoptile Segments to Exogenous Indole-3-acetic Acid

A concentration of 10^–5 M tomatine had no effect on leakagefrom, or elongation of, wheat coleoptile segments, but consistentlyreduced IAA-enhanced extension growth by c. 50 per cent. Therewas no evidence of chemical interaction between the alkaloidand the auxin in solution, and IAA action was not affected bypre-treatment for up to 3 h with 10^–5 M tomatine. Studieswith [2-¹⁴C]IAA revealed that 10^–5 M tomatine did notinhibit uptake of auxin into segments. The effect of pre-treatingsegments for up to 3 h with IAA could be virtually nullifiedby 10^–5 M tomatine, as could also IAA-induced changesin properties of coleoptile cell walls. Results are discussedin relation to the ability of tomatine to disrupt membrane functionand to current hypotheses implicating membranes in the primaryaction of auxin.     

Inhibition of Cell Division and DNA Synthesis by Gibberellin in Isolated Zinnia Mesophyll Cells

A culture system of isolated mesophyll cells of Zinnia eleganswas used to examine the action of gibberellic acid (GA) on celldivision. Isolated Zinnia mesophyll cells cultured in a mediumcontaining auxin and cytokinin reinitiated cell division ina partly synchronized manner. When mesophyll cells isolatedfrom 21-day-old seedlings were used, GA added to the culturemedium at concentrations of 1 x 10^–6 M or higher suppressedthe initial rise in the number of divided cells. Tracer experimentswith [³H]-dThd revealed that GA treatment inhibited the incorporationof [³H]-dThd into DNA in the nucleus without inhibiting theuptake of [³H]-dThd into the cells, indicating that GA inhibitedDNA synthesis. GA applied at 48 h inhibited the incorporationof [³H]-dThd into DNA during the following 24 h, but GA appliedat 72 h did not inhibit the incorporation during the subsequent24 h. This suggests that GA affects the process of reinitiationof DNA synthesis, but does not affect DNA synthesis once cellshave become proliferative. (Received January 14, 1986; Accepted March 31, 1986)     

O-Benzylhydroxylamine: An Inhibitor of Phenylpropanoid Metabolism in Plants

O-Benzylhydroxylamine (OBHA) is a potent inhibitor of phenylalanineammonialyase (PAL, EC 4.3.1.5 [EC] ) and phenylpropanoid metabolismas evidenced by its effects on three plant species [soybean(Glycine max (L.) Merr.), buckwheat (Fagopyrum esculentum Moench.),and mung bean (Vigna radiata L.)]. When supplied to roots, OBHA(10^–5 M) did not significantly inhibit light- or dark-growthof soybean seedlings, but reduced (25%) soluble hydroxyphenoliccompound accumulation in light-grown axes. Higher concentrations(510^–5 M) of OBHA caused reductions (25%) in axis freshweight of light-grown seedlings (72 h), but did not lower axisweight of dark-grown seedlings. Anthocyanin accumulation inhypocotyls of intact mung bean seedlings was reduced by 25%after 3 days light growth after treatment with OBHA (10^–5M) via root feeding. Anthocyanin content of excised, etiolatedbuckwheat hypocotyls floated on solutions of OBHA (10^–5M) and incubated in the light for 24 h was reduced by 40%. L-Phenylalanineand t-cinnamic acid, intermediates of phenylpropanoid metabolism,were able to partially reverse this inhibition in buckwheat.Extractable PAL activity (specific activity basis) in soybeanaxes was substantially reduced (20% in dark, 40% in light) asearly as 24 h after root feeding with OBHA (10^–5 M). Reductionof PAL activity (specific activity or per axis basis) by OBHAcompared to control levels, continued throughout a time courseof 96 h. Kinetic studies on soybean PAL revealed a K_m of 1.1mM for L-phenylalanine and an apparent K_i of 3.5 µM forOBHA. (Received May 31, 1985; Accepted August 6, 1985)     

Azetidine-2-carboxylic Acid and Lily Pollen Tube Elongation

Azetidine-2-carboxylic acid (AZC), which occurs naturally inLiliaceous plants, is reported to be a proline (pro) analoguePlant cell walls contain ‘extensin’, which is richin hydroxyproline (hyp). Peptidyl hyp arises through hydroxylationof peptidyl pro followed by glycosylation (arabinose attachment)of hyp Because AZC replaces peptidyl prolyl residues, it maybe a useful tool for evaluating the significance of hyp-o-arabinoselinkages in cell elongation. Therefore, we determined the effectof AZC on [¹⁴C]pro uptake, incorporation and conversion to wall-bound[¹⁴C]hyp in relation to elongation of lily pollen tubes whosewalls consist, in part, of hyp-containing glycopeptides TheAZC suppressed pollen germination 9–42 per cent (1–10mM) and subsequent tube elongation 40–54 per cent (0·1–1mM without affecting respiration In contrast, similar hyp concentrationswere without effect on tube elongation Whereas uptake of [¹⁴C]prowas 16·5–6·2 per cent of the control at0·1–1 mM AZC, [¹⁴C]leucine uptake was 85–25per cent of the control. Light microscope radioautography revealedfewer silver grains over tubes elongated in 0·1–1mM AZC than in its absence. Incorporation of [¹⁴C]pro into tnchloroaceticacid (TCA)-precipitable cytoplasm was reduced by only 10 percent at 0·01–1 mM but 43 per cent at 10 mM AZCGel filtration of cytoplasm from pollen germinated without AZCbut with [¹⁴C]pro resulted in labelled void volume (V) and threeretarded peaks (RI–III) Incorporation into V and RI wasinhibited at both 0·01 and 1 mM AZC These AZC concentrationsreduced conversion of [¹⁴C]pro to wall-bound hyp by 20 percent However, total incorporation of [¹⁴C]pro into salt-water-purifiedwall fractions was suppressed 47–53 per cent (0·1–1mM AZC). Lilium longiflorum, lily, hydroxyproline, proline, azetidine-2-carboxylic acid, pollen, pollen tube elongation     

Measurement of the Rates of Oxindole-3-Acetic Acid Turnover, and Indole-3-Acetic Acid Oxidation in Zea mays Seedlings

Nonhcbcl, H. M. 1986. Measurement of the rates of oxindole-3-aceticacid turnover and indole-3-acetic acid oxidation in Zea maysseedlings.—J. exp. Bat. 37: 1691–1697. Oxindole-3-acetic acid is the pnncipal catabolite of indole-3-aceticacid in Zea mays seedlings. In this paper measurements of theturnover of oxindole-3-acetic acid are presented and used tocalculate the rate of indole-3-acetic acid oxidation. [³H]Oxindolc-3-acetic acid was applied to the endosperm of Zeamays seedlings and allowed to equilibrate for 24 h before thestart of the experiment. The subsequent decrease in its specificactivity was used to calculate the turnover rate. The averagehalf-life of oxindole-3-acetic acid in the shoots was foundto be 30 h while that in the kernels had an average half-lifeof 35 h. Using previously published values of the pool sizesof oxindole-3-acetic acid in shoots and kernels from seedlingsof the same age and variety, and grown under the same conditions,the rate of indole-3-acetic acid oxidation was calculated tobe I-I pmol plant^–1 h^–1 in the shoots and 7·1pmol plant^–1 h^–1 in the kernels. Key words: Oxindole-3-acetic acid, indole-3-acetic acid, turnover, Zea mays     

Effect of cellulose synthesis inhibition on growth and the integration of xyloglucan into pea internode cell walls

2,6-Dichlorobenzonitrile (DCB, 100 μm) inhibited by 80 to 85% the incorporation of [³H]glucose into cellulose in stem segments of etiolated pea (Pisum sativum) seedlings. The inhibition lasted for at least 24 h. In the period 1 to 4 h after the excision of the segments, DCB did not influence elongation in the presence or absence of 2,4-dichlorophenoxyacetic acid (2,4-D). However, during the period 1 to 24 h after excision, DCB enhanced endogenous and 2,4-D-stimulated elongation by 65 and 34%, respectively. DCB did not affect the incorporation of ³H from [³H]arabinose into xyloglucan, and did not change the ability of the [³H]xyloglucan formed in vivo to bind strongly to the cell wall. Therefore, at least 80 to 85% of newly synthesized cellulose was excess to the requirements for tight wall binding of newly synthesized xyloglucan. This conflicts with the hypothesis that xyloglucan is held in the cell wall solely by direct hydrogen bonding to the surfaces of cellulosic microfibrils.     

Biosynthesis of Xyloglucan in Suspension-cultured Soybean Cells. Evidence that the Enzyme System of Xyloglucan Synthesis does not Contain {beta}-1,4-Glucan 4-{beta}-D-Glucosyltransferase Activity (EC 2.4.1.12)

Xyloglucan 4-ß-D-glucosyltransferase, an enzyme responsiblefor the formation of the xyloglucan backbone, in a particulatepreparation of soybean cells has been compared with ß-1,4-glucan4-ß-D-glucosyltransferase of the same origin. Thefollowing observations indicate that the enzyme system of xyloglucansynthesis does not contain ß-1,4-glucan 4-ß-D-glucosyltransferaseactivity, although both enzymes transfer the glucosyl residuefrom UDP-glucose to form the ß-1,4-glucosidic linkage:1. The incorporation of [¹⁴C]glucose into xyloglucan dependedon the presence of UDP-xylose in the incubation mixture. 2.No measurable amount of radioactivity was incorporated fromUDP-[¹⁴C]xylose into the cello-oligosaccharides, although theincorporation of [¹⁴C]xylose into xyloglucan depended on thepresence of UDP-glucose in the incubation mixture (Hayashi andMatsuda 1981b). 3. The activity of xyloglucan 4-ß-D-glucosyltransferasewas stimulated more strongly by Mn²⁺ than by Mg²⁺, whereas Mg²⁺was the most active stimulator for the activity of ß-1,4-glucan4-ß-D-glucosyltransferase. 4. An addition of GDP-glucose(100 µM) to the incubation mixture inhibited the activityof xyloglucan 4-ß-D-glucosyltransferase by 17%, whereasthe activity of ß-1,4-glucan 4-ß-D-glucosyltransferasewas inhibited 56% under the same conditions. 5. Irpex exo-cellulasedid not hydrolyze the xyloglucan synthesized in vitro. 6. Theß-1,4-glucan synthesized in vitro was not a branchedxyloglucan because it gave no 2,3-di-O-methyl glucose derivativeon methylation analysis. 7. Pulse-chase experiments indicatedthat the ß-1,4-glucan was not transformed into thexyloglucan. The subcellular distribution of the xyloglucan synthase, however,was similar to that of the ß-1,4-glucan synthase (Golgi-located1,4-ß-D-glucan 4-ß-D-glucosyltransferase).Thus, it appears that the latter enzyme is located at a siteclose to xyloglucan synthase and is set aside for the assemblyof these polysaccharides into the plant cell surface. (Received May 21, 1981; Accepted October 13, 1981)     

Transport, degradation and cell wall-integration of XXFGol, a growth-regulating nonasaccharide of xyloglucan, in pea stems

When [glucitol-³H]XXFGol (a NaB³H₄-reduced xyloglucan nonasaccharide) was applied to excised shoots of pea (Pisum sativum L. cv. Progress) at the base of the epicotyl, it inhibited growth in the elongation zone, 4–5 cm distal. Experiments were conducted to discover whether such ³H-oligosaccharides are translocated intact over this distance, or whether an intercellular second messenger would have to be postulated. After 24 h, ³H from [glucitol-³H]XXFGol and [glucitol-³H]XXXGol showed U-shaped distributions, with most ³H at the base and apex of the stem. Radioactivity from [fucosyl-³H]XXFG and [xylosyl-³H]XXFG also moved acropetally, but did not concentrate at the apex, possibly owing to removal from the transpiration stream of fucose and xylose formed by partial hydrolysis of XXFG en route. When 10⁻⁷ M [glucitol-³H]XXFGol was supplied, about 14 fmol ·  seedling^–1 of apparently intact [³H]XXFGol was extractable from the elongation zone after 24 h. Larger amounts of degradation products were extractable (including free [³H]glucitol) and some wall-bound ³H-hemicellulose was present (presumably formed by the oligosaccharides acting as acceptor substrates for transglycosylation). We conclude that biologically active oligosaccharides of xyloglucan can move through the stem acropetally and that they are maintained at low steady-state concentrations by both hydrolysis and transglycosylation. Received: 1 April 1997 / Accepted: 28 May 1997     

Promotion of Plant Cell Division by an Epidermal Growth Factor

In some specified treatments, an epidermal growth factor (EGF)promoted adventitious root formation in epicotyl cuttings ofVigna angularis. The number of the roots induced in cuttingstreated with 0.1 mg liter^-1 EGF during the first 24 h and with210^-4 M IAA during the second 24 h was 15% greater than thatof the roots in cuttings treated without EGF and with IAA. Analysisof the optimum timing of EGF application was performed by dividingthe first 24 h period into three sequential 8 h periods (0–8h, 8–16 h and 16–24 h). The most effective timeperiods in terms of the root formation were 8–16 h and16–24 h. The 0–8 h period was ineffective with respectto the formation. When carrot suspension cells were culturedfor 15 days at a very low cell density (1,000 cells/3 ml Murashigeand Skoog's medium) with more than 0.1 mg liter^-1 EGF, cellnumbers were 72% higher than those cultured without EGF. Theseresults suggest that EGF promotes cell division of plants. (Received October 5, 1992; Accepted May 24, 1993)     

Diurnal change in temperature sensitivity gibba G3 induced by acetylcholine in continuous light

When Lemna gibba G3 was in contact with 10^–5 M Ach oreserine, floral response to chilling changed diurnally under[24(0)], as was the case for control cultures exposed to [16(8)].On the other hand, with a rise in temperature, the min-[24(0)]decreased discontinuously in 24-hr units in Ach cultures subjectedto [24(0)], as did the min-[16(8)] in control cultures. Thusin the presence of Ach or eserine, this long-day plant tookthe continuous light regime, i.e. [24(0)], for another typeof long-day regime consisting of a long light period and a shortdark period, e.g. [16(8)]. In Ach cultures, the lower limitof the min-[24(0)] (72 hr) was attained at 22.5?C and remainedunaltered at the higher temperatures examined. The min-[16(8)]for control cultures, however, known to reach its lower limit(48 hr) at 26?C. Sodium lauryl sulphate (10^–5 M) and ouabain(10^–6 M) also caused a similar diurnal change in temperaturesensitivity of 24(0)] cultures. We surmised that exogenous Ach or inserted dark period modifiesthe relative rates chemical and physical component reactionsinvolved in the floral evocation processes, resulting in therhythmical floral response to chilling. (Received August 1, 1974; )     

Radiolabelling Studies of Lipids in the Marine Cryptomonad Chroomonas salina in Relation to Fatty Acid Desaturation

Cells of Chroomonas salina were exposed to [¹⁴C]acetate, [^l4C]16:0,[¹⁴C]18:0, [¹⁴C]18:1(n-9), [¹⁴C]18:2(n–6) or [¹⁴C]18:3(n–3)for 1 h and then incubated for 24 h in non-radioactive medium.At the end of the pulse period, non-glycolipid polar lipidscontained the highest proportions of radioactivity incorporatedfrom [¹⁴C]acetate and [¹⁴C]18:3(n–3) whereas with [¹⁴C]16:0,[¹⁴C]18:1 and [¹⁴C]18:2(n–6), triacylglycerols were mosthighly labelled. ¹⁴C-18:0 was recovered mainly as non-esterifiedfatty acid. Monogalactosyldiacylglycerol initially contained17% of label incorporated from [¹⁴C]acetate but less than 3%of that from [¹⁴C]fatty acids. With all substrates, excluding[¹⁴C]18:0, a gradual transfer of label from polar lipids totriacylglycerols was observed during the chase period. Saturatesand monoenes synthesised from [¹⁴C]acetate were mostly transferedfrom phospholipids and glycolipids to neutral lipid withoutfurther desaturation. Most of the incorporated ¹⁴C-fatty acidsremained unchanged and only with [¹⁴C]18:3(n–3) was substantialamounts of label recovered in penta- and hexaenoic fatty acids.The results indicate that, under the conditions of the study,lipid synthesis in the algae was heavily dominated by triacylglycerolformation and that the mechanisms of fatty acid desaturationin this species may differ from those in higher plants. (Received December 10, 1991; Accepted March 6, 1992)     

Effect of Inorganic Phosphate on the Biosynthesis of Purine and Pyrimidine Nucleotides in Suspension-Cultured Cells of Catharanthus roseus

The levels of purine and pyrimidine nucleotides in suspensioncultures of Catharanthus roseus were determined 24 h after stationary-phasecells were transferred to fresh complete (‘+Pi’)or phosphate-deficient (‘–Pi’) Murashige-Skoogmedium. The levels of ATP, GTP, UTP and CTP were from approx.3 to 5-fold greater in the cells grown in ‘+Pi’medium than in the cells grown in ‘–Pi’ medium.The levels of almost all other nucleotides were slightly higherin the cells in ‘+Pi’ medium. The rates of de novoand salvage biosynthesis of purine and pyrimidine nucleotideswere estimated from the rates of incorporation of radioactivityfrom [¹⁴C]formate, [2–¹⁴C]glycine, NaH¹⁴CO₃, [6–¹⁴C]orotate,[8–¹⁴C]adenine, [8–¹⁴C]adenosine, [2–¹⁴C]uraciland [2–¹⁴C]uridine. The results indicated that the activityof both the de novo and the salvage pathway was higher in thecells in ‘+Pi’ medium than in the cells in ‘–Pi’medium. The rate of degradation estimated from the rate of releaseof ¹⁴CO₂ from labelled purines and pyrimidines indicated thatdegradation of uridine was significantly reduced in the cellsin ‘+Pi’ medium, but no significant difference wasfound in the degradation of adenine, adenosine and uracil. Thepossible role of Pi in the control of the biosynthesis of nucleotidesand in the degradation of uridine is discussed. Catharanthus roseus, Madagascar periwinkle, suspension culture, inorganic phosphate, nucleotides, purines, pyrimidines, biosynthesis, degradation     

Involvement of cellulose synthesis in actions of gibberellin and kinetin on cell expansion. Gibberellin-coumarin and kinetin-coumarin interactions on stem elongation

In azuki bean (Azukia angularis = Vignia angularis) epicotylsections, 5 ? 10^–4 M coumarin inhibited the incorporationof radioactivity from [U^–14C]glucose into the cellulosefraction by 35% in the absence of indole-3-acetic acid (IAA)and by 40% in the presence of 1 ? 10^–4 M IAA. There wasno inhibitory effect on the incorporation of radioactivity intothe other fractions. Coumarin at 5 ? 10^–4 M reversed thepromoting effect of 1 ? 10^–5 M gibberellin A₃ (GA) andthe inhibitory effect of 1 ? 10^–5 M kinetin on IAA-inducedelongation of sections with no significant effects on IAA-inducedelongation. Neither GA nor kinetin had any appreciable effectson cellulose synthesis. No inhibition of cellulose syntheiswas observed with 1 ? 10^–3 M colchichine, which has beenreported to have effects similar to those of coumarin on GA-or kinetin-affected stem elongation. Coumarin at 5 ? 10^–4M was ineffectual in breaking up wall microtubules, while adisrupting effect on wall microtubules was clearly demonstratedwith 3 ? 10^–4M colchicine. From these results, the possible involvement of cellulose synthesisin cell expansion controlled by GA or kinetin was suggested. (Received August 3, 1973; )     

Turgor, Growth and Rheological Gradients of Wheat Roots Following Osmotic Stress

The growth rate of hydroponically grown wheat roots was reducedby mannitol solutions of various osmotic pressures. For example,following 24 h exposure to 0·96 MPa mannitol root elongationwas reduced from 1· mm h^–1 to 0·1 mm h^–1 Mature cell length was reduced from 290 µm in unstressedroots to 100 µm in 0·96 MPa mannitol. This indicatesa reduction in cell production rate from about 4 per h in theunstressed roots to 1 per h in the highest stress treatment. The growing zone extended over the apical 4·5 mm in unstressedroots but became shorter as growth ceased in the proximal regionsat higher levels of osmotic stress. The turgor pressure along the apical 5·0 mm of unstressedroots was between 0·5 and 0·6 MPa but declinedto 0·41 MPa over the next 50 mm. Following 24 h in 0·48(200 mol m^–3) or 0·72 MPa (300 mol m^–) mannitol,turgor along the apical 50 mm was indistinguishable from thatof unstressed roots but turgor declined more steeply in theregion 5·10 mm from the tip. At the highest level ofstress (0·96 MPa or 400 mol m^–3 mannitol) turgordeclined steeply within the apical 20 mm. Key words: Growth, turgor pressure, wall rheology, osmotic stress, osmotic adjustment     

Some Early Responses of Helianthus annuus L. to Flooding: I. THE EFFECTS OF FLOODING ON THE UPTAKE AND LEAKAGE OF 'NON-ELECTROLYTES' BY ROOTS

Drakeford, D. R., Mukherjee, I. and Reid, D. M. 1985. Some earlyresponses of Helianthus annuus L. to flooding. I. The effectsof flooding on the uptake and leakage of ‘non-electrolytes’by roots.—J. exp. Bot. 36: 1705–1715. The object of this work was to examine some of the early effectsof flooding on roots. A hydroponic system was developed thatgave good control over watering, degree of oxygenation of thebathing medium and allowed measurement of short term changesin the composition of the bathing medium. Excised roots, floodedfor 24 h, were shown to take up less [³H) ß-alaninethan non-flooded roots and also leaked more [³H] ß-alanineinto a distilled water bathing medium. Further, flooded excisedroots lost more protein to the bathing medium, with ‘young’(5–7 d) roots showing greater losses than ‘old’(11–14 d) roots. However, young roots had more proteinin the tissue even after greater loss. Young roots remainedhealthier and lost less fresh weight than old roots. Abscisicacid was shown to have a small role in protecting ‘young’roots from the effects of flooding. Key words: ABA, abscisic acid, anaerobic, flooding, leakage, roots, uptake, waterlogging