The Influence of Leaf Water Status and ABA on Leaf Growth and Stomata of Phaseolus Seedlings with Hypoxic Roots

The objective of this study was to assess the relative rolesof leaf water status and root-sourced signals in mediating beanleaf responses to root hypoxia. To do so, the roots of beanplants under varied VPD (0.95 kPa to 0.25 KPa) were made hypoxic.Under all conditions, leaf growth rates and stomatal conductanceswere reduced. There was a transitory decline in leaf water potentialat high VPD which accounted for the initial reduction in leafgrowth rates and stomatal conductance. At low VPD, no waterdeficits were detected. Leaf growth inhibition and reduced stomatalconductance under low VPD treatments were unrelated to leafwater status and must be induced by some other factor. In vitrogrowth of leaf discs was reduced by xylem sap collected fromhypoxic roots. Exogenously applied ABA, at high concentrationsin KCl and sucrose, or at low concentrations diluted in xylemsap from aerated plants, inhibited in vitro growth of leaf discs.Applications of ABA in the transpiration stream reduced stomatalconductance.     

Photocontrol of Dark Circadian Rhythms in Stomata of Phaseolus vulgaris L

Stomatal diffusion resistance in primary leaves of Phaseolus vulgaris L. which had been grown in light:dark cycles followed a marked circadian rhythm when the plants were transferred to continuous darkness. Reentrainment of the rhythm required more than one inductive change in photoperiod. The phasing of the rhythm of dark stomatal opening was contolled primarily by the light-on (dawn) signal, whereas the rhythm of dark closure was related to the light-off (dusk) signal. The evidence points to a dual control of the circadian clock in which a product of photosynthesis plays a major role. No evidence for phytochrome involvement in the phasing of the rhythm was found. An influence of phytochrome on the amplitude of the stomatal rhythm was observed in which removal of phytochrome-far-red absorbing form caused rapid damping.     

Identification of Cytokinins from Xylem Exudate of Phaseolus vulgaris L

The principal biologically active cytokinins in xylem exudate of young Phaseolus vulgaris L. plants were identified by bioassay, high-performance liquid chromatography, enzymic degradation and combined gas chromatography-mass spectrometry (selected ion monitoring) a zeatin riboside, zeatin nucleotide, dihydrozeatin riboside, dihydrozeatin nucleotide, O-glucosyl zeatin, O-glycosyl dihydrozeatin, O-glucosyl dihydrozeatin riboside, and O-glucosyl dihydrozeatin nucleotide. Trace amounts of O-glucosyl zeatin riboside and O-glucosyl zeatin nucleotide were also detected.     

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     

Cell-Wall Proteins Induced by Water Deficit in Bean (Phaseolus vulgaris L.) Seedlings

In the last few years, much attention has been given to the role of proteins that accumulate during water deficit. In this work, we analyzed the electrophoretic patterns of basic protein extracts, enriched for a number of cell-wall proteins, from bean (Phaseolus vulgaris L.) seedlings and 21-d-old plants subjected to water deficit. Three major basic proteins accumulated in bean seedlings exposed to low water potentials, with apparent molecular masses of 36, 33, and 22 kD, which we refer to as p36, p33, and p22, respectively. Leaves and roots of 21-d-old plants grown under low-water-availability conditions accumulated only p36 and p33 proteins. In 21-d-old plants subjected to a fast rate of water loss, both p33 and p36 accumulated to approximately the same levels, whereas if the plants were subjected to a gradual loss of water, p33 accumulated to higher levels. Both p36 and p33 were glycosylated and were found in the cell-wall fraction. In contrast, p22 was not glycosylated and was found in the soluble fraction. The accumulation of these proteins was also induced by abscisic acid (0.1-1.0 mM) treatment but not by wounding or by jasmonate treatment.     

Comparative Water Relations of Phaseolus vulgaris L. and Phaseolus acutifolius Gray

Leaf area expansion, dry weight, and water relations of Phaseolus vulgaris L. and P. acutifolius Gray were compared during a drying cycle in the greenhouse to understand the characteristics which contribute to the superior drought tolerance of P. acutifolius. Stomates of P. acutifolius closed at a much higher water potential than those of P. vulgaris, delaying dehydration of leaf tissue. P. acutifolius had a more deeply penetrating root system, which also contributes to its drought tolerance. Root-shoot ratios did not differ between the two species either under well watered or water stressed conditions. Leaf osmotic potential was also similar in the two species, with no apparent osmotic adjustment during water stress. These results indicate that P. acutifolius postpones dehydration and suggest that sensitive stomates and a deeply penetrating root system are characteristics which, if incorporated into cultivated beans, might increase their drought tolerance.     

Physiological Stress and Crystallites in Leaf Plastids of Phaseolus vulgaris L.

Crystalline inclusions were found in leaf plastids of Phaseolusvulgaris L. cultivar Limburg when excised plant parts were used.Removal of the root system induced crystalloid production afteran incubation period of optimal length. In agreement with thefindings of other authors physiological stress seems to be theunderlying condition of crystal formation in plastids.     

Abscisic Acid Decreases Leaf Na+ Exclusion in Salt-Treated Phaseolus vulgaris L.

Previous results showed that in short-term NaCl-treated beans increased leaf abscisic acid (ABA) concentration was triggered by Na⁺ but not by Cl^-. In this work, the specificity of ABA signaling for Na⁺ homeostasis was studied by comparing the plant’s responses to solutions that modified accumulation of ABA and/or Na⁺ uptake and distribution, such as supplemental Ca²⁺, increased nutrient strength, different isosmotic composition, application of exogenous ABA, fluridone (an ABA inhibitor) and aminooxiacetic acid (AOA, an ethylene inhibitor). After fluridone pretreatment, salt-treated beans had lower Na⁺ uptake and higher leaf Na⁺ exclusion capacity than non-pretreated plants. Moreover, Na⁺ uptake was increased and leaf Na⁺ exclusion was decreased by AOA and ABA. NaCl and KCl similarly increased leaf ABA and decreased transpiration rates, whereas supplemental Ca²⁺ and increased strength nutrient solution decreased leaf ABA and leaf Na⁺. These results show (1) a non-ion-specific increase in ABA that probably signaled the osmotic component of salt, and (2) increased ABA levels that resulted in higher leaf Na⁺ concentrations due to lower Na⁺ exclusion or increased root-shoot Na⁺ translocation.     

Water permeability and revolving movement in Phaseolus vulgaris L. twining shoots

Osmotic water permeability (Pos) was measured in protoplasts isolated from different tissues of Phaseolus vulgaris twining shoot. Parenchyma protoplasts exhibited more frequently high Pos values than epidermis protoplasts did. Water channels could facilitate water movement between parenchyma cells whereas cell-to-cell water transport mostly occurs through plasmodesmata in epidermis.     

In Vivo Nitrite Reduction in Leaf Tissue of Phaseolus vulgaris L

Experiments were performed to establish a procedure for in vivo measurement of nitrite utilization by leaf tissue of bean (Phaseolus vulgaris L. cv. Top Crop).     

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     

Stomatal Behaviour and Water Relations of Chilled Phaseolus vulgaris L. and Pisum sativum L.

Leaf diffusion resistance interpreted as stomatal resistance,leaf water potential (_w), solute potential (_s) and leaf turgorpotential (_p) of the chilling sensitive species Phaseolus vulgariswere determined during chilling at 4 °C in the light. Bothchill-hardened and non-hardened plants were used. For comparison,the chilling resistant species Pisum sativum was also used. The results for chilled P. sativum were similar to those obtainedfor chill-hardened P. vulgaris plants receiving a chilling treatment.In both cases a reduction in stomatal aperture and the maintenanceof a positive leaf turgor were the responses to chilling. Leavesof chilled but non-hardened P. vulgaris plants were found tomaintain open stomata throughout the chilling treatment despitea severe wilt developing after 7 h at 4 °C. This was incontrast to the chill-resistant P. sativum. which showed a rapidclosing and subsequent re-opening of the stomata to a new reducedaperture. During the first 12 h of chilling _wof P. vulgaris leaves changedfrom –0.47 MPa to –1.24 MPa. On more prolonged chilling_w tended to return to pre-chilling values. In addition. _p decreasedfrom 0.42 MPa to zero after only 9 h of chilling, and remainedat this value for the remainder of the chilling period, _s, changedrapidly from –0.89 MPa to –1.35 MPa in the first7.5 h, and after 9 h. _w and _s, were equal, i.e. zero _p. In contrast,the chilling resistant plant P. sativum maintained a positive_p throughout the chilling period, and there was little differencebetween values of _w, and _s in control and chilled leaves. Key words: Chilling, Stomata, ater relations, Phaseolus vulgaris, Pisum sativum     

ABA Levels and Effects in Chilled and Hardened Phaseolus vulgaris

Leaf abscisic acid (ABA) levels of chilled P. vulgaris weremeasured after 18 h chilling at 5°C, at a saturation deficitof 1.24 g m^–3 (SD), and after chilling in a water-saturatedatmosphere. Changes were also followed during a chill hardeningperiod of 4 d at 12°C, 2.1 g m^–3 SD. It was foundthat hardening resulted in an almost 5. fold increase in ABAlevels after 3 d at 12°C, and this decreased to approximatelycontrol levels on the fourth day. Subsequent chilling of hardenedplants produced no change in ABA levels from that of controlplants (22° C). In contrast, non-hardened plants chilledat 1.24 g m^–3 SD had ABA levels almost 3 times the levelof control plants. However, chilling in a water-saturated atmosphereresulted in a decrease in ABA levels. In addition, the response of leaf diffusion resistance (LDR)to exogenous ABA fed via the transpiration stream was measuredat 5 ° C and 22° C in hardened and non-hardened plants.Use of tritium-labelled ABA was made to calculate the stomatalsensitivity to ABA. It was found that exogenous ABA caused anincreased in LDR at 22°C in both hardened and non-hardenedplants. However, the sensitivity of the hardened plants to ABAwas greater in terms of rate of closure and amount of ABA requiredto close the stomata. At 5°C, however, ABA caused stomatalopening and the maintainance of open stomata in non-hardenedplants. In hardened plants, ABA caused stomatal closure at 5°C.These results are discussed in relation to the locking-openresponse of chilled P. vulgaris stomata. Key words: Chilling, Stomata, ABA, Phaseolus vulgaris     

The Rhythmic Leaf Movements after Regeneration of Partially Excised Pulvinus in Phaseolus vulgaris L.

Unlike the petiole or stem, the laminar pulvinus of the primaryleaves of Phaseolus vulgaris L. regenerated after a partialexcision. The histological and physiological aspects of theseregeneration processes have been studied. On the third day afterthe excision of the flexor (or extensor) region, the pulvinuswas regenerated. When the major part of the extensor was cutaway, the period and phase of the circadian leaf movements wereunchanged whereas the amplitude was greatly reduced. When theflexor region was excised, period, phase and amplitude weremaintained. Some changes could be seen in the ultradian movementsafter excision of flexor as well extensor regions. (Received August 31, 1988; Accepted March 30, 1989)     

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.     

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.     

Hypobaric Control of Ethylene-Induced Leaf Senescence in Intact Plants of Phaseolus vulgaris L

A controlled atmospheric-environment system (CAES) designed to sustain normal or hypobaric ambient growing conditions was developed, described, and evaluated for its effectiveness as a research tool capable of controlling ethylene-induced leaf senescence in intact plants of Phaseolus vulgaris L.

Senescence was prematurely-induced in primary leaves by treatment with 30 parts per million ethephon. Ethephon-derived endogenous ethylene reached peak levels within 6 hours at 26°C. Total endogenous ethylene levels then temporarily stabilized at approximately 1.75 microliters per liter from 6 to 24 hours. Thereafter, a progressive rise in ethylene resulted from leaf tissue metabolism and release. Throughout the study, the endogenous ethylene content of ethephon-treated leaves was greater than that of nontreated leaves.

Subjecting ethephon-treated leaves to atmospheres of 200 millibars, with O₂ and CO₂ compositions set to approximate normal atmospheric partial pressures, prevented chlorophyll loss. Alternately, subjecting ethephon-treated plants to 200 millibars of air only partially prevented chlorophyll loss. Hypobaric conditions (200 millibars), with O₂ and CO₂ at normal atmospheric availability, could be delayed until 48 hours after ethephon treatment and still prevent most leaf senescence. In conclusion, hypobaric conditions established and maintained within the CAES prevented ethylene-induced senescence (chlorosis) in intact plants, provided O₂ and CO₂ partial pressures were maintained at levels approximating normal ambient availability.

An unexpected increase in endogenous ethylene was detected within nontreated control leaves 48 hours subsequent to relocation from winter greenhouse conditions (latitude, 42°00″ N) to the CAES operating at normal ambient pressure. The longer photoperiod and/or higher temperature utilized within the CAES are hypothesized to influence ethylene metabolism directly and growth-promotive processes (e.g. response thresholds) indirectly.

     

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.