Sterol Composition in Relation to Chill-Sensitivity in Phaseolus spp.

Guye, M. G. 1988. Sterol composition in relation to chill-sensitivityin Phaseolus spp.—J. exp. Bot. 39: 1091–1096. The relative composition of the free sterols campesterol, sitosteroland stigmasterol was determined in the primary leaf laminaeof six bean (Phaseolus spp.) cultivars, known to differ in chill-sensitivity,prior to, during and following chilling of whole plants at 5?Cfor 24 h. Campesterol levels showed no significant changes during thechill-warm cycle and no relation was found between campesterollevels and chill-sensitivity. Transfer of plants from chillingto warm conditions resulted in an increase in the mol ratioof sitosterol to stigmasterol (S/S) in one of the most chill-tolerantgenotypes (P. coccineus cv. Prizewinner) and a decrease in thissterol ratio in the most chill-sensitive genotype {P. aureuscv. Berken). Genotypes representing a range of intermediatechill-sensitivities (P. vulgaris cvs 251, 222 and Tendergreen)showed no changes in S/S during a chill-warm cycle. Prior toand during chilling S/S was positively related to chill-tolerance Key words: Chill-sensitivity, campesterol, sitosterol/stigmasterol, physiological age, Phaseolus     

Chilling-Induced Ethylene Production in Relation to Chill-Sensitivity in Phaseolus spp.

Guye, M. G, Vigh, L. and Wilson, J. M. 1987. Chilling-inducedethylene production in relation to chill-sensitivity in Phaseolusspp.—J. exp. Bot. 38: 680–690. Ethylene production from the primary leaves of six bean (Phaseolusspp.) cultivars known to differ in chill-sensitivity, was monitoredat 23 ?C following chilling of whole plants at 5 ?C for 24 h.The more chill-tolerant cultivars produced greater amounts ofchilling-induced ethylene than the chill-sensitive cultivars.The onset of maximum ethylene production rates and the followingdecline in rates was more rapid in chill-tolerant cultivars.This pattern of ethylene production was also similar when chill-tolerancewas chemically enhanced by choline treatment. The low levelsof ethylene production in chill-sensitive genotypes was alsoreflected by their poor ability to convert the exogenously appliedethylene precursor, 1-aminocyclopropane-l-carboxylic acid (ACC),to ethylene. Moderate levels of leaf water deficit induced by chilling chill-tolerantcultivars and choline treated plants appeared to stimulate chilling-inducedethylene production. High levels of leaf wilt, shown by morechill-sensitive cultivars, reduced this stimulatory effect.Ethylene production was slightly greater when warming was carriedout in the light rather than in the dark. Key words: Ethylene, ACC, choline, chill-sensitivity, Phaseolus     

Polyamine Metabolism in the Roots of Phaseolus vulgaris. Interaction of the Inhibitors of Polyamine Biosynthesis with Putrescine in Growth and Polyamine Biosynthesis

The effects of the inhibitors of polyamine biosynthesis, canavanineand -methyl ornithine on growth, the activities of argininedecarboxylase (EC 4.1.1.19 [EC] ) and ornithine decarboxylase (EC4.1.1.17 [EC] ) and on polyamine content were examined in two differentgrowth regions of Phaseolus vulgaris L. cv. Taylor's Horticulturalroots. Separately, in the same manner, in the same bean rootsystem exogenous putrescine effect and the interaction of canavaninewith putrescine were determined. The arginine and ornithine decarboxylase activities found inroot apex were high where cell division activity was highest.Polyamine (putrescine and spermine) content did not correlatewith these activities, but polyamine level was high in the rootbase where cell elongation is the main process. The arginineanalogue, canavanine, inhibited arginine decayboxylase activityand polymine liters. Putrescine partially reversed the canavanineinhibition of root growth as well as arginine decarboxylaseactivity and polyamine content. Similarly -methyl ornithineslightly inhibited the root length and ornithine decarboxylaseactivity in the root apex. Besides, exogenous putrescine didnot effect significantly the endogenous polyamine titers. Theseresults reinforce the growing connection between polyaminesand the rates of cell devision in the roots of bean plants.Separately, arginine decarboxylase is the main enzyme in thebean roots. (Received November 10, 1986; Accepted March 3, 1987)     

Polyamine Metabolism and Osmotic Stress : I. Relation to Protoplast Viability

Cereal leaves subjected to the osmotica routinely used for protoplast isolation show a rapid increase in arginine decarboxylase activity, a massive accumulation of putrescine, and slow conversion of putrescine to the higher polyamines, spermidine, and spermine (HE Flores, AW Galston 1984 Plant Physiol 75: 102). Mesophyll protoplasts from these leaves, which have a high putrescine:polyamine ratio, do not undergo sustained division. By contrast, in Nicotiana, Capsicum, Datura, Trigonella, andVigna, dicot genera that readily regenerate plants from mesophyll protoplasts, the response of leaves to osmotic stress is opposite to that in cereals. Putrescine titer as well as arginine and ornithine decarboxylase activities decline in these osmotically stressed dicot leaves, while spermidine and spermine titers increase. Thus, the putrescine:polyamine ratio in Vigna protoplasts, which divide readily, is 4-fold lower than in oat protoplasts, which divide poorly. We suggest that this differing response of polyamine metabolism to osmotic stress may account in part for the failure of cereal mesophyll protoplasts to develop readily in vitro.     

Osmotic Stress-Induced Polyamine Accumulation in Cereal Leaves : II. Relation to Amino Acid Pools

Arginine decarboxylase activity increases 2- to 3-fold in osmotically stressed oat leaves in both light and dark, but putrescine accumulation in the dark is only one-third to one-half of that in light-stressed leaves. If arginine or ornithine are supplied to dark-stressed leaves, putrescine rises to levels comparable to those obtained by incubation under light. Thus, precursor amino acid availability is limiting to the stress response. Amino acid levels change rapidly upon osmotic treatment; notably, glutamic acid decreases with a corresponding rise in glutamine. Difluoromethylarginine (0.01-0.1 millimolar), the enzyme-activated irreversible inhibitor of arginine decarboxylase, prevents the stress-induced putrescine rise, as well as the incorporation of label from [¹⁴C]arginine, with the expected accumulation of free arginine, but has no effect on the rest of the amino acid pool. The use of specific inhibitors such as α-difluoromethylarginine is suggested as probes for the physiological significance of stress responses by plant cells.     

Polyamine Content and Action in Roots of Zea mays L. in Relation to Aerenchyma Development

Roots of Zea mays L. developed more aerenchyma (cortical gas-filledspace) when partially deficient in oxygen (3 kPa) than whensupplied with air (20·8 kPa oxygen) in association withfaster production of ethylene (ethene). The possibility wastested that the additional ethylene production resulted fromdecreases in spermidine (spd) and spermine (spm) which share,with ethylene, a common precursor, S-adenosylmethionine. However,no decreases in spd and spm were seen in root tissue up to 4d-old. Removing oxygen completely also had little effect onspd and spm, but strongly suppressed both ethylene productionand aerenchyma formation. Partial oxygen shortage (3 kPa) increased the concentrationof putrescine (put), the precursor of spd and spm. This increasewas not a response to the extra ethylene formed by such rootssince ethylene treatment did no reproduce the effect. Applicationof inhibitors of put biosynthesis, difluoromethylarginine anddifluoromethylornithine, led to increased aerenchyma formation.Exogenous put inhibited the development of aerenchyma whilestimulating rather than inhibiting ethylene production, whentested in either air or 3 kPa oxygen. Thus, put appears to limitaerenchyma formation by suppressing ethylene action rather thanits production.Copyright 1993, 1999 Academic Press Ethylene, ethene, roots, aerenchyma, polyamines, oxygen shortage, anaerobiosis, environmental stress, Zea mays     

Polyamine Levels in Relation to Growth and Somatic Embryogenesis in Tissue 6Medicago Sativa L.

Polyamine levels in petiole-derived tissue cultures of two highlyregenerable Medicago sativa L. genotypes were determined duringthe embryo induction and embryo differentiation phases of somaticembryogenesis. Putrescine levels increased 27–32-foldduring the 10 d on 2, 4-D- and kinetin-containing embryo inductionmedium and fell sharply following transfer to growth substance-freeembryo differentiation medium. The rapid increase in putrescinecontent occurred during a period of relatively little growthwhile the decline coincided with the initiation of rapid tissuegrowth on embryo differentiation medium. The addition of putativeinhibitors of putrescine or spermidine biosynthesis to the embryoinduction medium led to reduced levels of polyamines, particularlyof putrescine, in cultures of both genotypes but subsequentsomatic embryo formation was inhibited in cultures of one genotypeonly. It was concluded that the pronounced change in putrescinemetabolism was not specifically associated with embryogenesis,but appeared to be related generally to re-programming of cellsinto a new pattern of in vitro development. Key words: Medicago sativa, polyamines, somatic embryogenesis     

Dynamics of Imbibition in Phaseolus vulgaris L. in Relation to Initial Seed Moisture Content

The seed moisture level marking the onset of imbibitional injury (breakpoint) was determined for two cultivars of Phaseolus vulgaris L. cvs `Tendercrop' (TC) and `Kinghorn Wax' (KW). At 20°C the breakpoints were 0.15 gram H₂O/gram dry weight (gram per gram) for TC and 0.11 gram per gram for KW. When seeds were imbibed at 5°C, the breakpoints were 0.19 gram per gram (TC) and 0.16 gram per gram (KW). Below the breakpoint germination changed 4.6%/0.01 gram per gram for all treatments. Imbibition rates were maximal at 0.07 gram per gram and 0.33 gram per gram after 20 minutes imbibition. Rates of electrolyte leakage were correlated with the imbibition rate maximum at 0.07 gram per gram but were unaffected by the maximum at 0.33 gram per gram. The transition from tightly bound to semibound water occurred at 0.09 gram per gram and 0.11 gram per gram for KW and TC, respectively. T1 values increased exponentially as seed moisture decreased from 0.47 gram per gram to 0.05 gram per gram. ¹³C-NMR sugar signals increased at moisture levels above 0.14 gram per gram and plateaued at approximately 0.33 gram per gram seed moisture. These results suggest that the breakpoint moisture level for imbibitional damage is a function of temperature while the injury process is similar at both 5 and 20°C. Imbibition and leakage rate maxima reflect transitions in the states of seed water. NMR data support the application of the Water Replacement Hypothesis to seeds. Thus, imbibitional injury may be related to specific, temperature dependent moisture levels that are determined by water binding characteristics in the seed tissue.     

Polyamine metabolism in barley reacting hypersensitively to the powdery mildew fungus Blumeria graminis f. sp. hordei

Polyamine levels and activities of enzymes of polyamine biosynthesis and catabolism were examined in the barley cultivar Delibes (Ml1al + Ml(Ab)) reacting hypersensitively to the powdery mildew fungus, Blumeria graminis f. sp. hordei (race CC220). Levels of free putrescine and spermine and of conjugated forms of putrescine, spermidine and spermine were greatly increased 1–4 d following inoculation of barley with the powdery mildew. These changes in polyamine levels were accompanied by elevated activities of the polyamine biosynthetic enzymes ornithine decarboxylase (ODC), arginine decarboxylase (ADC) and S‐adenosylmethionine decarboxylase (AdoMetDC) and the polyamine catabolic enzymes diamine oxidase (DAO) and polyamine oxidase (PAO). Activities of two enzymes involved in conjugating polyamines to hydroxycinnamic acids, putrescine hydroxycinnamoyl transferase (PHT) and tyramine feruloyl‐CoA transferase (TFT) were also examined and were found to increase significantly 1–4 d after inoculation. The possibility that the increased levels of free spermine, increased polyamine conjugates, and increased DAO and PAO activities are involved in development of the hypersensitive response of Delibes to powdery mildew infection is discussed.     

Polyamine Content in Relation to Ploidy Level and to Different Organs of Nicotiana plumbaginifolia

Levels of free and bound polyamines (PAs) have been analysedin different organs of diploid, haploid and hypohaploid phenotypesof Nicotiana plumbaginifolia. In leaves and stems there is apositive correlation between the content of the PAs bound tomacromolecules and ploidy level. In contrast, in flowers a negativecorrelation is found with bound spermidine (Spd) levels. Besidesthe ploidy degree, the high levels of Spd in the flowers ofthe haploid and hypohaploid phenotypes might be also due tothe abnormal floral development and thus supports the view thatthis triamine could play a role in floral differentiation. (Received January 16, 1990; Accepted June 6, 1990)     

Swelling of Phaseolus Mitochondria in Relation to Free Fatty Acid Levels

Freshly isolated Phaseolus vulgaris mitochondria contain, 1.8 micromoles of long chain, primarily unsaturated, free fatty acids per milligram of nitrogen. Although there is no measurable increase in free fatty acid content as a result of spontaneous swelling in buffered KCl, bovine serum albumin strongly inhibits the degree of swelling. The mitochondria only swell slowly in sucrose, but rapid swelling can be induced by the addition of oleic acid in a process inhibited by bovine serum albumin. The possible participation of the endogenous free fatty acids of Phaseolus mitochondria in spontaneous swelling is discussed.     

Fontibacillus phaseoli sp. nov. isolated from Phaseolus vulgaris nodules

A bacterial strain, designated BAPVE7B^T, was isolated from root nodules of Phaseolus vulgaris in Spain. Phylogenetic analysis based on its 16S rRNA gene sequence placed the isolate into the genus Fontibacillus with Fontibacillus panacisegetis KCTC 13564^T its closest relative with 97.1 % identity. The isolate was observed to be a Gram-positive, motile and sporulating rod. The catalase test was negative and oxidase was weak. The strain was found to reduce nitrate to nitrite and to produce β-galactosidase but the production of gelatinase, caseinase, urease, arginine dehydrolase, ornithine or lysine decarboxylase was negative. Acetoin production and aesculin hydrolysis were found to be positive. Growth was observed to be supported by many carbohydrates and organic acids as carbon source. MK-7 was identified as the predominant menaquinone and the major fatty acid (43.7 %) as anteiso-C_15:0, as occurs in the other species of the genus Fontibacillus. Strain BAPVE7B^T displayed a complex lipid profile consisting of diphosphatidylglycerol, phosphatidylglycerol, four glycolipids, four phospholipids, two lipids, two aminolipids and an aminophospholipid. Mesodiaminopimelic acid was detected in the peptidoglycan. The G+C content was determined to be 45.6 mol% (Tm). Phylogenetic, chemotaxonomic and phenotypic analyses showed that strain BAPVE7B^T should be considered a new species of genus Fontibacillus, for which the name Fontibacillus phaseoli sp. nov. is proposed (type strain, LMG 27589^T, CECT 8333^T).     

Transport of 14C-Assimilates in Seedlings of Phaseolus vulgaris L. in Relation to Vascular Anatomy

Patterns of ¹⁴C-photosynthate translocation in bean (Phaseolusvulgaris L.) seedlings have been investigated in relation tovascular-bundle continuity between exporting and importing organsby use of radioassay and tissue-clearing techniques. Assimilatefrom the primary leaves reaches the first trifoliate leaf byan indirect route. There is no direct vascular connection betweenthe primary leaves and distal tissues. Bundles of the primaryleaf petiole connect with an anastomosis at the node, and allbundles which originate from this structure descend the stem.Assimilate from primary leaves moves down the stem, and is thentransferred to an ascending pathway, the bundles of which traversethe anastomosis at the second node. The lateral leaflets ofthe first trifoliate leaf are served differentially by primaryleaves with respect to assimilate supply. Differences are relatedto position, and may be accounted for by differences in vascularcontinuity.     

Polyamine biosynthesis and titer during various developmental stages of Phaseolus vulgaris

The activities of arginine decarboxylase (ADC; EC 4.1.1.19) and ornithine decarboxylase (ODC; EC 4.1.1.17) as well as polyamine content were examined in Phaseolus vulgaris L. cv. Taylor's Horticultural before and during anthesis, during fruit development and throughout vegetative growth. The specific activities of polyamine biosynthetic enzymes were highest in all rapidly growing tissues, e.g., root apices, hypocotyls, young internodes, young leaves, flower buds, young pods and pericarps. They were lowest in mature, non-growing tissues. Similarly, the content of the major polyamines (putrescine, spermidine, spermine) is highest in rapidly growing tissues, and lowest in mature tissue. These correlations reinforce the growing connection between polyamines and rates of cell division and metabolic activity during both vegetative and reproductive development.     

Co-inoculation with Bacillus sp. CECT 450 improves nodulation in Phaseolus vulgaris L.

The strain Bacillus sp. CECT 450 increased nodulation on bean (Phaseolus vulgaris L.) when co-inoculated with Rhizobium tropici CIAT 899. This positive effect occured under controlled conditions on perlite-vermiculite, sand, or in a mixture of soil and sand. This increase was also observed in a field assay. Nodulation kinetic studies suggested that the synergistic effect is pronounced during the latter stages of cultivation. In contrast, the same bacteria co-inoculated with Bradyrhizobium japonicum USDA 110 reduced nodulation on soybean (Glycine max (L.) Merr.). Inoculation with Bacillus sp. CECT 450 alone had no effect on bean plants, but reduced root growth in soybean. The survival of Bacillus sp. CECT 450 on inoculated seeds was high, even when inoculated seeds were maintained for several months at room temperature.     

Polyamine Metabolism in Ripening Tomato Fruit : II. Polyamine Metabolism and Synthesis in Relation to Enhanced Putrescine Content and Storage Life of a/c Tomato Fruit

The fruit of the Alcobaca landrace of tomato (Lycopersicon esculentum Mill.) have prolonged keeping qualities (determined by the allele a/c) and contain three times as much putrescine as the standard Rutgers variety (A/c) at the ripe stage (ARG Dibble, PJ Davies, MA Mutschler [1988] Plant Physiol 86: 338-340). Polyamine metabolism and biosynthesis were compared in fruit from Rutgers and Rutgers-a/c—a near isogenic line possessing the allele a/c, at four different stages of ripening. The levels of soluble polyamine conjugates as well as wall bound polyamines in the pericarp tissue and jelly were very low or nondetectable in both genotypes. The increase in putrescine content in a/c pericarp is not related to normal ripening as it occurred with time and whether or not the fruit ripened. Pericarp discs of both normal and a/c fruit showed a decrease in the metabolism of [1,4-¹⁴C]putrescine and [terminal labeled-³H]spermidine with ripening, but there were no significant differences between the two genotypes. The activity of ornithine decarboxylase was similar in the fruit pericarp of the two lines. Arginine decarboxylase activity decreased during ripening in Rutgers but decreased and rose again in Rutgers-a/c fruit, and as a result it was significantly higher in a/c fruit than in the normal fruit at the ripe stage. The elevated putrescine levels in a/c fruit appear, therefore, to be due to an increase in the activity of arginine decarboxylase.     

A Study of the Aminoacyl-sRNA Synthetases of Phaseolus vulgaris in Relation to Germination

Optimum conditions for the extraction and assay by ATP-pyrophosphate exchange of the aminoacyl-sRNA synthetases of the various tissues of french bean (Phaseolus vulgaris) seeds and seedlings are described. Extracts of plumules, after passage through Sephadex G-25, were assayed for synthetase activity using an amino acid mixture as substrate, when a 30 to 100-fold stimulation of exchange above the endogenous level was obtained. This marked enhancement of exchange by added amino acids is largely attributed to the use of dilute extracts.     

Polyamine metabolism and lipoxygenase activity during Fusarium oxysporum f. sp. ricini -Castor interaction

Polyamine oxidase and lipoxygenase enzymes are key players for hyper sensitive reaction (HR) during incompatible interaction of host-pathogen. Thus, the role of lipoxygenase and polyamines was studied in the wilt pathogen infected and non infected tissues of resistant and susceptible genotypes of castor at 0 days after infection (DAI), 5 DAI and 10 DAI (30 days after sowing). The lipoxygenase (LOX) and polyamine oxidase (PAO) activities were higher in the incompatible interaction at all the stages of analysis. The constitutive level of malondyaldehyde (MDA) content, a product of lipid peroxidation was higher in susceptible genotypes (VP-1 and VI-9), while induced level was higher in resistant genotypes (48–1 and SKP-84) at 5 DAI and 10 DAI . Polyamine profiling using HPTLC showed higher spermidine and spermine content in resistant genotypes at 10 DAI. Furthermore, spermidine was detected only in the roots of resistant genotypes at 10 DAI. These results suggest the role of high titers of polyamines, LOX and PAO in disease resistance possibly through HR induction.