Water potentials induced by growth in soybean hypocotyls

Gradients in water potential form the driving force for the movement of water for cell enlargement. In stems, they are oriented radially around the vascular system but should also be present along the stem. To test this possibility, growth, water potential, osmotic potential, and turgor were determined at intervals along the length of dark-grown soybean (Glycine max L. Merr., cv. Wayne) hypocotyls. Transpiration was negligible in the dark, humid conditions, so that all water uptake was for growth. Elongation occurred in the terminal 1.5 centimeters of the hypocotyl. Water potential was −3.5 bars in the elongating region but −0.5 bar in the mature region, both in intact plants and detached tissue. There was a gradual transition between these values that was related to the growth profile along the hypocotyl. Tissue osmotic potentials generally paralleled tissue water potentials, so that turgor was the same throughout the length of the hypocotyl. If the elongating zone was excised, growth ceased immediately. If the elongating zone was excised along with mature tissue, however, growth continued, which confirmed the presence of a water-potential gradient that caused longitudinal water movement from the mature zone to the elongating zone. When the plants were grown in vermiculite having low water potentials, tissue water potentials and osmotic potentials both decreased, so that water potential gradients and turgor remained undiminished. It is concluded that growth-induced water potentials reflect the local activity for cell enlargement and are supported by appropriate osmotic potentials.     

Auxin-induced changes in the population of translatable messenger RNA in elongating sections of soybean hypocotyl

In vitro translation products of polyadenylated RNA from untreated and auxin-treated elongating sections of soybean (Glycine max var. Wayne) hypocotyl were analyzed by two-dimensional polyacrylamide gel electrophoresis. The levels of translatable messenger RNA for at least ten in vitro translation products are increased by auxin treatment. The induction by auxin occurs rapidly (within 15 minutes), and the amounts of the induced in vitro translation products increase with time of auxin treatment. Indoleacetic acid has the same effect on the population of translatable messenger RNA as 2,4-dichlorophenoxyacetic acid. The auxin-induced in vitro translation products disappear rapidly when Actinomycin D is present during the last two hours of a three-hour auxin treatment.     

Auxin- and ethylene-induced changes in the population of translatable messenger RNA in Basal sections and intact soybean hypocotyl

In vitro translation products of polyadenylated RNA from untreated and auxin-treated basal sections of soybean (Glycine max var. Wayne) hypocotyl were analyzed by two-dimensional polyacrylamide gel electrophoresis. Within one hour of 2,4-dichlorophenoxyacetic acid treatment, the translatable messenger RNAs for at least twelve in vitro translation products are modulated upward. In vitro translation products of polyadenylated RNA from untreated, auxin-treated and Ethephon-treated intact soybean hypocotyl were also analyzed. Within two hours of treatment with either 2,4-dichlorophenoxyacetic acid or Ethephon, the translatable messenger RNAs for a group of high molecular weight in vitro translation products are modulated upward. There is a particular set of translatable messenger RNA, encoding in vitro translation products in the 24,000 to 32,000 molecular weight range, that is specifically modulated upward by auxin treatment in intact soybean hypocotyl and in hypocotyl sections.     

Effects of abscisic Acid on growth, RNA metabolism, and respiration in germinating bean axes

The effect of abscisic acid on growth, respiration, the ATP pool, and rate and amount of RNA synthesis in aseptically cultured axes of Phaseolus vulgaris during the first 24 hours of germination has been measured in experiments where the duration of abscisic acid application and its concentration have been varied. At concentrations from 10(-7) to 10(-4)m, abscisic acid inhibits synthesis of RNA with maximal inhibition (80%) at 10(-5)m. RNA synthesis is inhibited by abscisic acid at all times examined (12, 18, and 24 hours), but the extent of inhibition is maximal at 18 hours. In 18-hour axes RNA synthesis is inhibited 42%, ATP pool size is reduced 3%, and O(2) consumption is decreased by 6% after 75 minutes of abscisic acid treatment. Inhibition of RNA synthesis is complete by 2 hours of treatment with abscisic acid, and recovery to near control levels occurs by the 3rd hour after removal from abscisic acid.     

Reduction of turgor induces rapid changes in leaf translatable RNA

The turgor of pea (Pisum sativum) leaves was reduced by exposing excised pea shoots to a stream of 23°C air for 20 min. Poly(A)⁺ RNA was isolated from control and wilted shoots, translated in vitro and radiolabeled translation products separated by electrophoresis on two-dimensional (isoelectric focusing-sodium dodecyl sulfate) polyacrylamide gels. This analysis showed that the levels of several poly(A)⁺ RNAs increased in wilted plants. Most of the poly(A)⁺ RNAs induced in wilted plants did not accumulate in response to heat shock or exogenously applied ABA even though endogenous ABA levels were found to increase in shoots 30 min after wilting and by 4 h had increased 50-fold (1 versus 0.02 microgram per gram fresh weight). A λgt10 cDNA library was constructed using poly(A)⁺ RNA from wilted shoots which had been incubated for 4 hours. Differential screening of the library identified four clones corresponding to poly(A)⁺ RNAs which are induced in wilted shoots.     

Benzyladenine reversal of abscisic Acid inhibition of growth and RNA synthesis in germinating bean axes

The effect of benzyladenine on growth, ATP pool size and specific radioactivity, and the rate and amount of RNA synthesis in aseptically cultured axes of Phaseolus vulgaris during the first 24 hours of germination were measured in experiments where the duration of benzyladenine application and its concentration were varied. Maximum promotion of growth (25%) occurs at 10(-5)m benzyladenine. Maximum promotion of RNA synthesis (44%) occurs at 10(-5)m benzyladenine. Benzyladenine has little effect on the size or specific radioactivity of the ATP pool. Benzyladenine can completely counteract abscisic acid inhibition of growth and RNA synthesis, and these reversals are measurable in 2 hours. GA(1) and GA(3) do not promote growth or counteract abscisic acid inhibition of growth in germinating bean axes in these experiments.     

Fractionation of RNA polymerase subspecies from soybean hypocotyls by isoelectric focusing in Sephadex

RNA polymerase enzymes isolated from chromatin of 6-day-old soybean hypocotyls are resolved into two major and one minor species of activity by DEAE-Sephadex chromatography. A soluble form of the enzyme, isolated from the postchromatin supernatant fraction, shows a broad peak of activity when fractionated by this method. The elution characteristics and α-amanitin sensitivity data indicate the two major chromatin-bound activities to be Class I and III enzymes, while the minor chromatin-bound activity and the soluble enzyme are representative of the Class II enzymes. In contrast to these profiles, fractionation of these enzyme preparations by the new method of isoelectric focusing in Sephadex G-15 yields five distinct chromatin-bound and four soluble subspecies. The relationships of these observed activities to their parent DEAE classes are investigated, showing two subspecies within the Class I and III RNA polymerase enzymes, respectively, and four subspecies within the Class II enzymes.     

Polyadenylated RNA sequences which are reduced in concentration following auxin treatment of soybean hypocotyls

Previous work has shown that any effect of exogenous auxin on gene expression in soybean hypocotyl tissue must be restricted to a relatively small fraction of the polyadenylated RNA. However, kinetic hybridization analysis with cDNA probes revealed that a minor abundant class of sequences is markedly reduced in concentrations in the auxin-treated polyadenylated RNA. Recombinant plasmids containing copies of polyadenylated RNA species were constructed using the G-C tailing procedure and clones of auxin-regulated sequences were detected by differential in situ hybridization with cDNA of polyadenylated RNA from auxin-treated or untreated hypocotyls. Although the 12 clones which were selected all contained different size inserts, and were therefore independent, 11 of these apparently hybridized to just two different RNA species. The rate constant of the auxin-sensitive abundant component of the untreated polyadenylated RNA/DNA hybridization was similar to that of the reaction between the two major groups of clones and untreated polyadenylated RNA. This indicates that these cloned sequences are homologous with that cDNA fraction. The twelfth clone is thought to be representative of a group of less abundnt auxin-regulated polyadenylated mRNA species which had been detected in an earlier analysis of the in vitro translation products of soybean hypocotyl RNA. Both the timing and the extent of the influence of auxin on the relative concentration of these cloned sequences are quite consistent with a close relationship between growth regulation by auxin and its effects on gene expression.     

Chromatin RNA polymerase activity from soybean hypocotyls treated with gibberellic acid and AMO-1618

Chromatin isolated from control, AMO-1618 [2′-isopropyl-4′-(trimethylammonium chloride)-5′-methylphenyl piperidine carboxylate] and gibberellic acid (GA) treated soybean hypocotyl tissue incorporates labeled nucleoside triphosphates into acid-insoluble RNA. Gibberellic acid, sprayed on intact soybean hypocotyls, is shown to have enhanced the level of chromatin RNA polymerase activity while chromatin isolated from hypocotyls pretreated with AMO-1618 exhibits a lower polymerase activity relative to the control. Chromatin extracted from the treated or untreated seedlings are all sensitive to the inhibition (in varying degrees) by the presence of actinomycin D, pyrophosphate, or ribonuclease. Thus enhanced (or decreased) RNA-synthesizing capacity of chromatin in response to chemical treatments may be due to enhanced (or decreased) synthesis of RNA polymerase.     

Auxin-induced changes in the population of translatable messenger RNA in elongating maize coleoptile sections

In-vitro translation products of polyadenylated RNA from untreated and indole-3-acetic acid (IAA)-treated elongating sections of maize (Zea mays L.) coleoptiles were analyzed by twodimensional polyacrylamide gel electrophoresis. Treatment with IAA results in an increased amount of at least four in-vitro translation products. The amounts of two of these translation products are increased within 10 min of IAA treatment.Abbreviation IAA indole-3-acetic acid     

Effect of shading individual soybean reproductive structures on their abscisic Acid content, metabolism, and partitioning

Pod set in soybean is related to carbon partitioning and may be, at least partially, regulated by abscisic acid (ABA) concentrations. The studies reported here examine the relationship between carbon and ABA partitioning, reproductive abscission and ABA metabolism. The partitioning of radiolabeled ABA and photoassimilates from leaves to flowers and endogenous ABA concentrations were determined in shaded and unshaded reproductive structures. Aluminum foil was gently placed over individual soybean reproductive structures for 48 hours at 0, 4, 12, 17, and 22 days after anthesis (DAA). Shading of flowers at 12, 17, and 22 DAA resulted in significantly reduced concentration of ABA. However, shading had no effect on the catabolism of exogenously supplied [³H] ABA. The shading treatment on the first four of the five dates reduced partitioning of photoassimilates and ABA from the subtending leaf to the flower. Shading of reproductive structures also caused a significant reduction in the amount of assimilate exported from the subtending leaf, at 17 DAA. We conclude that shade-induced premature reproductive abscission in soybean is not stimulated by high levels of ABA within reproductive structures, but that ABA may inhibit abscission of reproductive structures by playing a role in preferential assimilate partitioning.     

Cold-acclimation induced changes in freezing tolerance and translatable RNA content in Citrus grandis and Poncirus trifoliata

Cold-acclimation-induced changes in freezing tolerance and translatable RNA content were compared in seedlings of a relatively cold sensitive citrus species, Citrus grandis L. Osb. cv. Thong Dee (pummelo), and the cold-hardy citrus relative, Poncirus trifoliata L. Raf. cv. Pomeroy (trifoliate orange). Cold acclimation of pummelo (10 days at 15°C followed by 4 weeks at 10°/5°C, day/night) resulted in a decrease in LT₅₀ from −6 to −8°C, while in trifoliate orange (acclimated for 7 weeks at 5°C), the LT₅₀ decreased from −9 to −18°C. Qualitative changes in the in vitro translation profile, revealed by two-dimensional gel electrophoresis, were observed following cold acclimation in both species. An mRNA for a large polypeptide (ca 160 kDa) was detected following cold acclimation of trifoliate orange. A similar change was not observed in pummelo following cold acclimation.     

Fruit age and changes in abscisic Acid content, ethylene production, and abscission rate of cotton fruits

The relationships of fruit age, abscisic acid (ABA) concentration, ethylene evolution, and abscission rates were studied in an effort to determine why cotton (Gossypium hirsutum L., cv. Deltapine 16) fruits rarely abscise more than 15 days after anthesis. Because abscission of cotton fruits is increased by conditions that limit photosynthesis, greenhouse-grown plants with fruits of various ages were placed in dim light for 3 days to induce high rates of fruit abscission. Abscission rates, ABA concentrations, and ethylene evolution rates were determined for fruits of various ages. Almost all of the young fruits abscised, but abscission rate declined with age until almost no abscission was observed in fruits that were 15 or more days past anthesis.     

Effects of pod removal on the transport and accumulation of abscisic Acid and indole-3-acetic Acid in soybean leaves

Concentrations of abscisic acid (ABA) and indole-3-acetic acid (IAA) in the second most recently expanded trifoliolate leaf were determined during reproductive development of soybean (Glycine max [L.] Merr cv `Chippewa 64'). The concentration of ABA in leaves was constant during most of the seed filling period until the seeds began to dry. The concentration of IAA in the leaves decreased throughout development. Removal of pods 36 hours prior to sampling resulted in increased concentrations of ABA in leaves during the period of rapid pod filling but had little effect on the concentration of IAA in leaves. ABA appears to accumulate in leaves after fruit removal only when fruits represent the major sink for photosynthate.

ABA and IAA moving acropetally and basipetally in petioles of soybean were estimated using a phloem exudation technique. ABA was found to move mostly in the basipetal direction in petioles (away from laminae). IAA, primarily in the form of ester conjugate(s), was found to be moving acropetally (toward laminae) in petioles. The highest amount of IAA ester(s) was found in petiole exudate during the mid and late stages of seed filling. Removal of fruits 36 hours prior to exudation reduced the amount of IAA ester recovered in exudate, suggesting that fruits were a source of the IAA conjugate in petiole exudate.

     

Induction of heat shock protein messenger RNA in maize mesocotyls by water stress, abscisic Acid, and wounding

Exposure of the excised growing region of the mesocotyl of young corn seedlings to heat shock stimulated the production of specific heat shock proteins and the intensification of synthesis of two proteins with a molecular weight of approximately 70,000. Water stress and abscisic acid also stimulated synthesis of these 70,000-dalton proteins, and other unique proteins distinct from those induced by heat shock. Growing tissues of intact corn mesocotyls exposed to heat shock, water stress, or abscisic acid accumulated mRNA species homologous to a cloned genomic probe of the 5′ end of the 70,000-dalton Drosophila heat shock protein gene. Since cut segments of the mesocotyl under unstressed conditions produced a similar mRNA, we suggest that the hsp 70 gene is activated in corn by a variety of diverse stresses. Production of the mRNA is rapid, but transient, being induced within 3 hours of the imposition of the stress, but declining after reaching a maximum at 9 hours.