Isolation and characterization of the cell wall receptor of 14C-malformin in Phaseolus vulgaris L.

¹⁴C-malformin attaches to at least two cell wall receptors inPhaseolus vulgaris. One receptor was extracted with Tris buffer(pH 8.5) and the other with 0.1 N NaOH. In both cases, priortreatment of the walls with wall degrading enzymes (macerase,cellulysin) was required. The two receptors differed with regardto ultrafiltration and gel filtration chromatography. The Tris-extractedreceptor is a protein, probably a glycoprotein, which containshydroxyproline and sulfhydryl groups. Although cuttings nottreated with malformin had Tris-extractable receptor, formationof the receptor appeared to be enhanced by malformin. ¹ Present address: American Cyanamid, P. O. Box 400, Princeton,New Jersey 08540, U. S. A. (Received August 2, 1976; )     

Identification of some major cytokinins in Phaseolus vulgaris and their distribution

The determination of the various endogenous cylokinins and their distribution among organs is important in understanding their role in growth and development in the intact plant. Cytokinins in young plants of Phaseolus vulgaris were purified by immunoaffinity chromatography and HPLC, and characterised by UV spectra. Zeatin nucleotide (zeatin riboside-5'-monophosphate) and isopentenyladenine nucleotide (isopentenyladenosne-5'-monopnosphate) were the most abundant cytokinins in all organs. Their identities were confirmed by GC-MS. The levels of zeatin, zeatin riboside, isopentenyladenine and isopentenyladenosine never exceeded 5% of the nucleotides, as assessed by a methodology that preserves cytokinin nucleotides. Three extraction methods were compared with qualitatively similar results, though differing in their suppression of nucleotidase activity. Cytokinin nucleotide levels were greater in the stems and petioles than in the roots and leaves on a per gram fresh weight basis, and were greater in the stems than in the other organs on a per plant basis. Levels of the zeatin and isopentenyladenine nucleotides were about equal in the stems and leaves, but in the petioles the zeatin nucleotide levels were about twice the level of isopentenyladenine nucleotide, while in the roots they were about half the isopentenyladenine nucleotide level. The importance of considering the cytokinin form is emphasised.     

Apical Dominance in Phaseolus vulgaris L.

Although determinations of the ABA content of lateral buds ofPhaseolus vulgaris revealed no difference between decapitatedand intact control plants in the first 12 h following decapitation,a relative decrease in the ABA content of lateral buds of decapitatedplants was detectable 24 h following decapitation. Shoot decapitationwas also observed to result in a decrease in the ABA contentof stem tissue. The application of IAA to the stem of decapitatedplants prevented these changes and increased the ABA contentof stem tissue relative to that of intact plants. The levelsof IAA and ABA were also determined in the stem tissue fromthe nodes of intact bean plants. The possible interdependenceof these two plant hormones was further investigated by a studyof [2_–14ClABA metabolism. The results are discussed inrelation to the possible role of these hormones in apical dominance. Key words: Apical dominance, Abscisic acid, Indole-3-acetic acid     

Cytokinin metabolism in Phaseolus vulgaris L.

The major cytokinins in stems of decapitated, disbudded bean plants have been identified by enzymic degradation, Sephadex LH20 and reversed phase high performance liquid chromatography, and by combined gas chromatography-mass spectrometry as 6-(4-hydroxy-3-methylbut-trans-2-enylamino)-9--D-ribofuranosylpurine (zeatin riboside), 6-(4-hydroxy-3-methylbutylamino)-9--D-ribofuranosylpurine (dihydrozeatin riboside), and the 5-phosphates of these compounds (zeatin ribotide and dihydrozeatin ribotide). Minor cytokinins in this tissue were tentatively identified as dihydrozeatin-O--D-glucoside and zeatin ribotide-O--D-glucoside. [8-¹⁴C-]Dihydrozeatin appeared to be rapidly metabolized to dihydrozeatin ribotide when supplied to segments of stems from decapitated plants. These results are discussed in relation to the metabolism and distribution of cytokinins in the whole plant.Abbreviations TEAB triethyl ammonium bicarbonate - UV ultra-violet - GCMS gas chromatography-mass spectrometry - HPLC high performance liquid chromatography - TMS trimethyl silyl     

Temperature Effects on Mitochondrial Respiration in Phaseolus acutifolius A. Gray and Phaseolus vulgaris L

Electron transport, using succinate as a substrate, was measured polarographically in mitochondria isolated from Phaseolus vulgaris and P. acutifolius plants at 25°C and 32°C. Mitochondria isolated from P. vulgaris plants grown at 32°C had reduced electron transport and were substantially uncoupled. Growth at 32°C had no effect on electron transport or oxidative phosphorylation in P. acutifolius compared to 25°C grown plants. Mitochondria isolated from 25°C grown P. vulgaris plants measured at 42°C were completely uncoupled. Similarly treated P. acutifolius mitochondria remained coupled. The uncoupling of P. vulgaris was due to increased proton permeability of inner mitochondrial membrane. The alternative pathway was more sensitive to heat than the regular cytochrome pathway. At 42°C, no alternative pathway activity was detected. The substantially greater heat tolerance of P. acutifollus compared to P. vulgaris mitochondrial electron transport suggests that mitochondrial sensitivity to elevated temperatures is a major limitation to growth of P. vulgaris at high temperatures and is an important characteristic conveying tolerance in P. acutifolius.     

Partitioning of 14C-photoassimllates within seeds of Phaseolus coccineus (Lam.) and Phaseolus coccineus x Phaseolus vulgaris L.

The objective of this study was to determine partitioning within seeds of ¹⁴C-photoassimilates at three stages of seed development in two Phaseolus crosses — P. coccineus Lam. selfed, and P. coccineus x P. vulgaris L. Abortion of the interspecific embryos occurred when the seed reached 10 mm seed length. When expressed as sink strength (% dpm) or sink activity (% dmp/d.wt.) there were no differences in partitioning of ¹⁴C-photoassimilates when whole seeds were analyzed. If the seed was divided into seed coat, liquid endosperm, and embryo, the sink activity of the interspecific embryo was higher than that of the embryo in the selfed seed. Therefore, abortion of these interspecific Phaselus embryos appeared not to be caused by a lack of photoassimilates.Assistant Professor, Professors, respectively.Contribution from the Agr. Expt. Station, University of Minnesota, St. Paul, MN 55108. Paper No. 13,548, Scientific Journal Series. This research was supported in part by the Science and Education Administration of the United States Department of Agriculture under Grant 59-2271-9-2-020-0 and in part by a grant from the Minnesota Soybean Research and Promotion Council.     

Tissue Cultures of Phaseolus vulgaris L.

Abstract

Callus production and plant regeneration from different explants of Phaseolus vulgaris L. cv. Giza are reported. Calli cultures were induced from leaf, hypocotyl, embryo and root explants. Rapid growth of callus was achieved by leaf explants cultured on MS salts, B5 vitamins and supplemented with 2,4— dichlorophenoxyacetic acid (2, 4—D)+0.5 mg/l kinetin (kin). Addition of casein hydrolysate at 2 g/l to maintenance medium enhanced callus growth and hindered the early appearance of necrotic parts. This report also provides a detailed method for production of multiple shoots directly from the wounded edges of immature cotyledon explant via organogenesis on 1 mg/l benzyladenine (BA) or indirectly on 0.5 mg/l naphthaleneacetic acid (NAA)+2 mg/l BA. The regeneration of bean plants through the two ways described here (direct or indirect) may be of use in genetic improvement of bean.     

Ureide Metabolism in Non-nodulated Phaseolus vulgaris L.

The distribution of ureide-N was studied throughout vegetativeand reproductive growth of non-nodulated Phaseolus vulgarisL. (bushbean) grown in nitrate nutrient solution. Largest increasesin ureide-N per plant were correlated with flowering and earlypod formation and with seed filling. Highest amounts of ureidesper organ were measured in stems and axillary trifoliates. Highestconcentrations (µmol ureide-N g^–1 fr. wt.) weremeasured in young developing organs and stems. Seeds did notaccumulate ureides until the ureide content of pods had reacheda maximum. Results obtained using the inhibitor of xanthine oxidase, allopurinol,are consistent with the origin of ureides via purine degradationbut alternative pathways cannot be discounted. Leaves and stems were shown to have the ability to degrade allantoatevia an enzymic process.     

The influence of aluminium availability on phosphate uptake in Phaseolus vulgaris L. and Phaseolus lunatus L.

Aluminium toxicity is one of the major limiting factors of crop productivity on acid soils. High levels of available aluminium in soil may induce phosphorus deficiency in plants. This study investigates the influence of Aluminium (Al) on the phosphate (P_i) uptake of two Phaseolus species, Phaseolus vulgaris L. var. Red Kidney and Phaseolus lunatus L. The two bean species were treated first with solutions of Al at different concentrations (0, 25, 50 and 100 μM, pH 4.50) and second with solutions of P_i (150 μM) at pH 4.50. The higher the Al concentration the higher the Al concentration sorbed but P. vulgaris L var. Red Kidney adsorbed significantly more Al than P. lunatus L. Both species released organic acids: P. vulgaris L var. Red Kidney released fumaric acid and P. lunatus L. fumaric and oxalic acids which could have hindered further Al uptake.The two bean species showed a sigmoid P_i uptake trend but with two different mechanisms. P. vulgaris L var. Red Kidney showed a starting point of 3 h whereas P. lunatus L. adsorbed P_i immediately within the first minutes. In addition, P. vulgaris L var. Red Kidney presented significantly higher P_i uptake (higher uptake rate ‘k’ and higher maximum adsorption ‘a’ of the kinetic uptake model). The Al treatments did not significantly influence P_i uptake. Results suggest that P. lunatus L. might adopt an external Al detoxification mechanism by the release of oxalic acid. P. vulgaris L var. Red Kidney on the other hand seemed to adopt an internal detoxification mechanism even if the Al sorbed is poorly translocated into the shoots. More detailed studies will be necessary to better define Al tolerance and/or resistance of Phaseolus spp.     

Cytokinin Effects on Leaf Architecture in Phaseolus vulgaris L.

Treatment of expanding primary leaves of bean plants (Phaseolnsvulgaris L. cv. Limburgse vroege) with benzyladenine (BA) orkinetin at 0.5 mM for five consecutive days resulted in thickerleaves showing a significant decrease in intercellular air spacevolume. Compared with control plants, exposed mesophyll cellsurface area was lower per unit tissue volume, but unchangedwhen expressed per unit leaf surface area. Stomata of treatedplants were not fully closed in the dark and they did not openas wide as controls in the middle of the light period, suggestingthat the treatment resulted in impaired stomatal action. Allthe effects mentioned were more pronounced after treatment withBA, compared to kinetin. In spite of their magnitude, the observedchanges in leaf structure and function did not seem to havean important effect on total leaf diffusion resistance to carbondioxide during the course of the light period. Key words: Cytokinins, Leaf architecture     

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.     

Isozymes of Glutamine Synthetase in Phaseolus vulgaris L. and Phaseolus lunatus L. Root Nodules

The glutamine synthetase (GS) isozymes in the plant fraction of nodule extracts from 62 cultivars of Phaseolus vulgaris L. and one cultivar of Phaseolus lunatus L. were analyzed by polyacrylamide gel electrophoresis. All P. vulgaris nodule extracts displayed two GS activity bands: a nodule-specific band (GS_n1) and a band (GS_n2) similar to the single band (GS_r) present in root extracts. In nodule extracts of P. lunatus, the GS_n1 band was detected, but the GS_n2 band was barely detectable. In contrast to P. vulgaris, the GS_n2 band and the GS_r band of P. lunatus appeared to be different. The electrophoretic mobility of the GS_n1 band in P. vulgaris was governed by both the plant cultivar and the development stage of the nodule. In nodule extracts of P. vulgaris and P. lunatus, the zone of GS_n1 activity coincided with six to nine distinct protein bands as revealed after treatment of gels, which had previously been stained for GS activity, with Coomassie blue. All these protein bands were shown to consist of polypeptides of identical molecular weight (approximately 47,000 daltons) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Our results indicate that P. vulgaris continuously generates isozymes of GS_n1 of increasing electrophoretic mobility during the course of nodule development.     

Studies on ethylene binding by cell free preparations from cotyledons of Phaseolus vulgaris L.

Various protein reactive agents such as dithioerythritol, dithiothreitol, mercaptoethanol and p-chloromercuribenzoate inhibit binding of ethylene to cell free preparations of Phaseolus vulgaris L. The effect of the thiols is partially reversed by treatment with diamide; occupation of the binding site by ligand diminishes the inhibition caused by p-chloromercuribenzoate but not that caused by thiols. Growth regulators other than ethylene do not affect binding. Physiologically active structural analogues of ethylene competitively inhibit binding of the growth regulator and their relative effectiveness in the cell free system closely resembles that in developmental processes controlled ethylene.Abbreviations DTE dithioerythritol - DTT dithiothreitol - EPPS N-2-hydroxyethyl-piperazine propane sulphonic acid - ME mercaptoethanol - PCMB p-chloromercuribenzoate     

A major stress-inducible Mr-42000 wall glycoprotein of French bean (Phaseolus vulgaris L.)

A major wall protein of suspension-cultured cells of French bean has been isolated and characterised. It can be prepared from walls or the culture filtrate and in composition it is particularly rich in proline, valine and glutamic acid/glutamine and contains appreciable amounts of hydroxyproline. The N-terminus shows some glycosylation, while following chemical deglycosylation the first 38 residues were found to be identical to those of proline-rich proteins from soybean. However, the composition of the highly purified M_r-42000 bean protein differs considerably from the soybean proteins and must contain its own specific domains. An antibody was raised and used to demonstrate the inducibility of the M_r-42000 bean protein in response to elicitor action. The protein was found to be mainly localised in the intercellular spaces of the cortical cells of bean hypocotyls and at the wall-plasmalemma interface of xylem vessels, another potentially accessible compartment for pathogens. Following wounding, the protein was found to be generally distributed in the wall of epidermal and cortical cells of the hypocotyls. The M_r-42000 protein is cross reactive with antibodies raised to glycoproteins of the Rhizobium infection thread and the chitin-binding hydroxyproline-rich glycoprotein, potato lectin. These common epitopes together with the previously demonstrated chitin-binding properties of the bean protein indicate a role in host-microbial interactions. Furthermore, the M_r-42000 protein itself bound to the growing hyphal tips of the bean pathogen, Colletotrichum lindemuthianum.Abbreviations FITC fluorescein isothiocyanate - IgG immunoglobulin G - PAL phenylalanine ammonia-lyase We thank Dr Nick Brewin for advice on interpretation of immunolocalisations and for the gift of MCA 265. We thank Dudley Fernandino for carrying out the confocal microscopy. GPB thanks the Science and Engineering Research Council for funding.     

Effect of malformin on the major constituents of Phaseolus vulgaris

Malformin inhibits wet and dry weight, nitrogen accumulation,and cell wall, RNA, DNA and protein synthesis in Phaseolus vulgaris.The relative proportion of dry matter and nitrogen in malformedtissues is increased in the ethanol soluble fraction and decreasedin the residue remaining after hydrolysis with 0.5 N HCl. Inhibitionof cell wall and protein synthesis was generally greater thaninhibition of nitrogen accumulation and RNA and DNA synthesis.The effects of malformin on the composition of P. vulgaris aresimilar to alterations in composition reported for ethylene,and opposite to those reported for gibberellic acid. ¹This research was supported by grant GB-7158 from the NationalScience Foundation and grant E-146-F from the American CancerSociety. ²Journal Paper No. 3509 of the Purdue Agricultural ExperimentStation. (Received October 23, 1968; )     

Compensation among root classes in Phaseolus vulgaris L.

The activity of soil pathogens, competition for assimilates, and the changing availability of below-ground resources make root systems subject to a continuous and dynamic process of formation and loss of both fine and coarse roots. As hypocotyl borne roots appear later than other root classes, they may serve to functionally replace basal and primary roots lost to biotic and abiotic stress. Using common bean (Phaseolus vulgaris L.), we conducted experiments in solution and solid media culture with treatments involving the removal of part of the root system (basal, hypocotyl borne or primary roots), phosphorus availability, and depth of seeding to test the hypothesis that there are compensation mechanisms among basal, hypocotyl borne and primary roots to cope with the loss of part of the root system. The root system was highly plastic in response to root excision, which resulted in the maintenance of below-ground biomass accumulation. In most cases, this compensation among root classes was enough to maintain plant performance in both phosphorus sufficient and phosphorus stressed plants. Removal of a specific root class induced an increase in the growth of the remaining root classes. All root classes, but especially the primary root, contributed to the compensation mechanism in some way. Primary roots represented around 10% of the root system in control plants and this proportion increased dramatically (up to 50%) when other root classes were removed. In contrast, negligible compensatory re-growth was observed following removal of the primary root. Greater planting depth increased the production of hypocotyl borne roots at the expense of basal roots. The proportion of hypocotyl borne roots increased from 25% of the whole root system when seeds were placed at a depth of 2 cm to 30% when they were placed at 5 cm and to 38% when placed at 8 cm, with corresponding decreases in the proportion represented by basal roots. The common feature of our observations is the innate ability of the root system for its own regeneration. Total root biomass maintained strict allometric relationships with total shoot biomass in all treatments. Re-stabilization of root to shoot balance after partial root loss is governed by overall plant size following allometric relationships similar to undisturbed plants. However, the pattern of this root regeneration was not uniform since the way the three root classes compensated each other after the removal of any one of them varied among the different growth media and phosphorus supply conditions. The resulting changes in root architecture could have functional significance for soil resource acquisition.     

Developmental Features of the Primary Phloem in Phaseolus vulgaris L.

Certain developmental features of the primary phloem were examinedin Phaseolus vulgaris L., chiefly by the use of the pulvinusat the base of the petiole. The cells included in the studywere the sieve element, the companion cell, and the tannin cell.In the sieve element, the sieve plate shows the usual sequenceof conversion of plasmodesmatal canals into pores. The endoplasmicreticulum, which appears as flat cisternae associated with ribosomesin younger cells, later becomes in part stacked and in partaligned parallel with the walls as a network. The stacked ERprecedes the anastornosing parietal ER in time of development,but the parietal ER persists longer. Of the two forms of P-proteincharacteristic of a number of Fabaceae, the crystalline bodyappears considerably in advance of the body composed of tubules.Neither form of P-protein disperses completely in the maturecell, although the crystalline protein may spread out into aggregatesof fine fibrils. The companion cells show the typical denseprotoplasts and branched plasrnodesmatal connections with thesieve elements. The vacuome of these cells is dispersed intonumerous small vacuoles, many of which appear to be concernedwith autophagic digestion of protoplasmic material. The tannincells have large vacuoles in which the tannin material is located.The cells form vertical series in which the end walls becomeperforated.     

Interspecific hybridization of Phaseolus vulgaris L. and Phaseolus angustissimus A. Gray using in vitro embryo culture

The introgression of new desirable characters is very necessary in common bean (Phaseolus vulgaris L.)Interspecific hybridization of P. vulgaris L. with P. angustissimus A. Gray, a wild species with narrow leaves, could be successfully achieved for the first time using embryo culture. Sixteen to twenty-three-day old embryos could grow to plantlets when they were cultivated on modified Monnier's medium.The leaf shape of the hybrid plants obtained showed intermediary traits between the two parents.A chromosome stock doubling could be achieved for one embryo.     

Late Transplanting and the Yield of Phaseolus vulgaris L.

The prediction that very high seed yields of dry beans (Phaseolusvulgaris L.) would be produced by the delayed transplantingof large plants has been tested in a factorial experiment withfour dates of transplanting and eight plant populations. Therewere significant differences in yield between transplantingdates and between population densities, and there was a significantdate-density interaction. At low plant densities (up to about30 plants m^–2) the three transplanted treatments yieldedless than the hand-sown controls, and late transplanting yieldedless than early. At the highest density the situation was reversed;all three transplanted treatments out-yielded the controls andlate transplanting tended to out-yield plants transplanted early.The biggest yield was 340 g seed m^–2 from a transplantedcrop grown at 35 plants m^–2. The data on yield fitted a modified rectangular hyperbola ofthe form where y is yield per unit area, p is the number of plants perunit area, t is the number of days between sowing and transplanting,and B_o, n, m, and p are arbitrary parameters. This equationaccounted for 91 per cent of the variation in yield with t andp. It is suggested that late transplanting had adverse effects,due to transplanting ‘shock’ and which were mostmarked at low plant densities; and beneficial effects, ascribableto an effect on plant ‘plasticity’, which were mostmarked at high plant densities. Possible physiological mechanismsof these effects are discussed. Phaseolus vulgaris, yield, density, transplanting