Gibberellic Acid-Promoted Lignification and Phenylalanine Ammonia-lyase Activity in a Dwarf Pea (Pisum sativum)

The effects of gibberellic acid on lignification in seedlings of a dwarf and a tall cultivar of pea (Pisum sativum) grown under red or white light or in the darkness, were studied. Gibberellic acid (10⁻⁶-10⁻⁴ m) promoted stem elongation in both light and dark and increased the percentage of lignin in the stems of the light-grown dwarf pea. The gibberellin had no effect on the lignin content of the tall pea although high concentrations (10⁻⁴ m) promoted growth of the tall plants. Time course studies indicated that the enhanced lignification in the gibberellin-treated dwarf plants occurred only after a lag period of several days. It was concluded that gibberellic acid-enhanced ligmification had no direct relation to gibberellic acid-promoted growth. The activity of phenylalanine ammonia-lyase (E.C. 4.3.1.5) was higher in gibberellin-treated dwarf plants grown under white or red light than in untreated dwarf plants. Gibberellic acid had no detectable effect on the activity of this enzyme when the plants were grown in darkness, just as it had no effect on lignification under dark conditions. The data suggest that in gibberellin-deficient peas the activity of phenylalanine ammonia-lyase is one of the limiting factors in lignification.     

Evidence for the selective replication of dwarf pea DNA evoked by exogenous gibberellin application

Characteristics of the total DNA preparations isolated from apical parts of dwarf pea seedlings untreated and treated with gibberellic acid (GA3) were compared. Analytical centrifugation in a self-generated CsCl density gradient revealed the occurrence of a heavy satellite DNA band (p = 1.712 g X cm-3) in addition to the main DNA band (p = 1.696 g X cm-3) in the DNA preparation extracted from GA3-treated seedlings, that could not be detected in the DNA isolated from untreated plants. The existence of this GC-rich DNA fraction was additionally confirmed by means of derivative DNA melting profiles. Comparison of the reassociation kinetics obtained for control DNA with DNA from GA3-treated plants showed changes in the percentage distribution of three main DNA sequence classes, with different repetition frequency in the haploid pea genome. It is postulated that such a variation in the percentage of different C0t families might reflect the selective DNA replication evoked by hormonal treatment of dwarf pea plants.     

Synergism between Gibberellin and Auxin in the Growth of Intact Tomato

Gibberellin A_4&7 was more effective than gibberellic acid in increasing shoot elongation when applied to the apex of intact Lycopersicum esculentum seedlings of Tiny Tim, a dwarf cultivar, and Winsall, a tall cultivar. After 14 days, gibberellic acid and gibberellin A_4&7 stimulated growth of the dwarf more than the tall tomato. In tall tomato the application of indole-3-acetic acid alone (6.1 μg/plant) showed an inhibitory growth effect, but when applied with 17.5 μg per plant of gibberellic acid, it had a synergistic effect at 7 days but not at 14 days. When the auxin concentration was reduced to 0.61 μg per plant a synergistic effect was observed on tall plants at 7 and 14 days between indole-3-acetic acid and gibberellic acid. Application of gibberellin A_4&7 with auxin did not give a synergistic response in tall or dwarf tomato.     

Chloroplastic activity in response to gibberellic acid treatment of dwarf and normal cultivars of pea (Pisum sativum L.)

Levels of ferricyanide reduction, cyclic and non-cyclic photophosphorylation were measured in chloroplasts of two cultivars of pea and a comparison of their P/2e⁺ ratios were made. No differences were observed in cyclic photophosphorylation or ferricyanide reduction but non-cyclic photophosphorylation was lower in chloroplasts from the dwarf than the normal cultivar. Thus the P/2e⁺ ratio of the dwarf was lower than the normal. Dwarf seedlings treated with gibberellic acid (GA₃) had similar rates of cyclic photophosphorylation as the untreated dwarf but non-cyclic photophosphorylation was lower as was ferricyanide reduction. This resulted in P/2e⁺ ratios that were higher in chloroplasts from the GA₃ treated dwarf seedlings than the untreated, and were the same as the untreated normal. Addition of GA₃ directly to the chloroplasts did not alter the activity in any way. Hence gibberellins do not directly affect changes in chloroplastic activity but may conceivably be involved in a feed-back control system.     

Auxin-gibberellin interaction in apical dominance: Experiments with tall and dwarf varieties of pea and bean

Summary Seedlings of dwarf and tall varieties of pea and bean, growing in John Innes Compost No. 2, were studied in relation to the effects of decapitation, indole-3-acetic acid (IAA), and gibberellic acid (GA₃) on axillary bud growth. In all varieties, GA₃ antagonized the inhibitory influence of IAA on bud growth when both hormones were applied to the upper cut end of the stem. Thus, GA₃ caused a reduction in IAA-induced correlative bud inhibition in tall, as well as in dwarf, plants. These results agree with those obtained by several workers, but contrast with some recent reports of increased apical dominance in a tall pea variety when seedlings were treated with GA₃ in addition to IAA. An attempt was made to identify the cause of opposite results being obtained by different workers, and it is considered that possibly the most important factor is mineral nutrition.     

ACTION OF GIBBERELLIC ACID ON CELL PROLIFERATION IN THE SUBAPICAL SHOOT MERISTEM OF WATERMELON SEEDLINGS

Rates of cell proliferation were about 2.5 fold greater in gibberellic acid-treated seedlings of dwarf watermelon than in untreated dwarf seedlings as determined by direct counts of pith cells in elongating hypocotyls of watermelon. The higher rates of cell proliferation observed in normal and GA-treated dwarf seedlings were due both to shorter cell cycle times and increased growth fractions of cells than in dwarf seedlings. The shorter duration of the cell cycle in GA-treated seedlings was primarily due to a reduced S period.     

The Promotion of Indole-3-acetic Acid Oxidation in Pea Buds by Gibberellic Acid and Treatment

Terminal buds of dark-grown pea (Pisum sativum) seedlings have an indole-3-acetic acid oxidase which does not require Mn(2+) and 2,4-dichlorophenol as cofactors. Oxidase activity is at least 50 times higher in buds of tall peas than in dwarf seedlings. Administration of gibberellic acid to dwarf peas stimulates both growth and indoleacetic acid oxidase activity to the same levels as in tall seedlings. By contrast, indoleacetic acid oxidation assayed in the presence of Mn(2+) and 2,4-dichlorophenol proceeds at similar rates regardless of gibberellin application. Treatment of tall peas with the growth retardant AMO-1618 reduces growth and oxidase activity. Such treated seedlings are indistinguishably dwarf. The enzyme does not appear to be polyphenol oxidase, nor do the results suggest that reduced activity in dwarf buds is due to higher levels of a dialyzable inhibitor. The peroxidative nature of the oxidase is probable.     

Activity of mitochondria isolated and purified from dwarf and tall,light-grown pea seedlings

Mitochondria from dwarf and tall light-grown peas have been separated from contaminating chloroplasts and other organelles on discontinuous sucrose gradients. A comparison between activities of the mitochondria from the two cultivars of light-grown pea seedlings was made. It was observed that both respiratory control (R.C.) indices and phosphate to oxygen (P : O) ratios were superior in mitochondria from the tall cultivar than the dwarf cultivar. The results are discussed in relation to energy status and possible gibberellic acid content in the two cultivars.     

Effects of Gibberellin on the Arrangement and the Cold Stability of Cortical Microtubules in Epidermal Cells of Pea Internodes

Longitudinal microtubules are predominant in epidermal cellsof the 3rd internodes of dwarf pea (Pisum sativum L. cv. LittleMarvel) seedlings. In more than 50% of the cells, cortical microtubulesare running parallel to the cell axis. GA₃ promotes elongation of the internodes and gives rise toa predominance of transverse microtubules. In more than 60%of the GA₃-treatd cells, cortical microtubules are running transverseto the cell axis. Longitudinal microtubules in the GA₃-untreated cells are resistantto low-temperature treatment, but transverse microtubules inthe GA₃-treated cells are sensitive to it. Longitudinal microtubulesare present in GA₃-treated epidermal cells with low frequency.They are resistant to low-temperature treatment. Longitudinal, oblique and transverse microtubules are presentwith almost the same frequency in epidermal cells of the 3rdinternodes of tall pea (cv. Early Alaska) seedlings. GA₃ promoteselongation of the internodes also in tall pea seedlings, butit does not alter the direction of cortical microtubules sodistinctly as it does in dwarf pea seedlings. As in dwarf pea seedlings, longitudinal microtubules are resistantto low-temperature treatment, and transverse microtubules aresensitive to it in tall pea seedlings. (Received September 19, 1986; Accepted December 26, 1986)     

The influence of waterlogging at different temperatures on penetration depth and porosity of roots and on stomatal diffusive resistance of pea and maize seedlings

The 8 days old seedlings of pea (cv. Ilowiecki) and maize (cv. Alma F1) were subjected to differentiated aeration conditions (control — with pore water tension about 15 kPa and flooded treatment) for 12 days at three soil temperatures (7, 15 and 25 °C). The shoots were grown at 25 °C while the soil temperature was differentiated by keeping the cylinders with the soil in thermostated water bath of the appropriate temperature. Lowering the root temperature with respect to the shoot temperature caused under control (oxic) conditions a decrease of the root penetration depth, their mass and porosity as well as a decrease of shoot height, their mass and chlorophyll content; the changes being more pronounced in maize as compared to the pea plants. Flooding the soil diminished the effect of temperature on the investigated parameters; the temperature effect remaining significant only in the case of shoot biomass and root porosity of pea plants. Root porosity of pea plants ranged from 2 to 4 % and that of maize plants — from 4 to 6 % of the root volume. Flooding the soil caused an increase in the root porosity of the pea plants in the entire temperature range and in maize roots at lower temperatures by about 1 % of the root volume. Flooding the soil caused a decrease of root mass and penetration depth as well as a decrease of plant height, biomass and leaf chlorophyll content.     

Correlative Studies on Plant Growth and Metabolism II. Effect of Light and of Gibberellic Acid on the Changes in Protein and Soluble Nitrogen in Lettuce Seedlings

Protein and soluble nitrogen distribution in different parts of lettuce seedling was studied in light and darkness and in presence and absence of gibberellic acid. In dark, applied gibberellic acid failed to show any marked effect on the nitrogen changes in lettuce. Light inhibits translocation of nitrogen reserves from the cotyledons. Gibberellic acid reverses the light inhibition of longitudinal growth but has no effect on the inhibition of translocation from the cotyledons. Light grown, gibberellic acid treated seedlings exhibit a pattern of protein and soluble-N which is characteristic of the dark grown seedlings. Thus gibberellic acid not only causes morphological reversal of light inhibition but also shifts the nitrogen metabolism of light grown plants, close to that of plants grown in darkness.     

Gibberellin-auxin interaction in pea stem elongation

Joint application of gibberellic acid and indole-3-acetic acid to excised stem sections, terminal cuttings, and decapitated plants of a green dwarf pea results in a markedly synergistic growth response to these hormones. Synergism in green tall pea stem sections is comparatively small, although growth is kinetically indistinguishable from similarly treated dwarf sections.

Gibberellin-induced growth does not appear to be mediated through its effect on auxin synthesis, since gibberellin pretreatment of dwarf cuttings fails to elicit an enhanced tryptophan-induced growth response of sections, whereas auxin-induced growth is strongly enhanced. Also, tryptophan-gibberellin synergism is not significant in sections and cuttings of green dwarf peas, while auxin-gibberellin synergism is.

Administration of gibberellic acid prior to indole-3-acetic acid results in greatly increased growth. In reversed order, the application fails to produce any synergistic interaction. This indicates that gibberellin action must precede auxin action in growth regulation.

     

Comparison ofin vivo activity of L-tryptophan synthas in plants with a low and a high content of L-tryptophan

The activity of L-tryptophan synthase (TS, E.C.4.2.1.20) was comparedin vivo in seedlings of plants high in L-tryptophan (L-trp) (pea and kohlrabi) and low in this amino acid (maize). In maize the TS was studied both in the normal and in the opaque-2 genotype that forms an endosperm richer in essential amino acids. The activity of TS was determined on the basis of the increase in radioactivity of the chromatographically purified L-trp-¹⁴C, synthetized after vacuum infiltration of L-serine-¹⁴C-(U) and ineubation for 24 h. As regards the TS activity in seedlings, maize is comparable to pea and kohlrabi; in contrast to this TS is less active in pea seedlings, which can be attributed to the presence of TS inhibitor (CHEN and BOLL 1969). In ripening maize kernels and leaves adjacent to the ear the TS activity is about 20 times lower than in seedlings. The differences in the activity of TS in the genotypes of maize could not be detected, even at the period of seed ripening. Therefore the differences in L-trp content in the investigated plants cannot be explained by a differing activity of TS. TS is probably not the determining regulator of L-trp level in plants, its activity is relatively high even in plants low in L-trp.     

Physiological Effects of Gibberellic Acid. IX. Recovery of Gibberellic Acid Following Incubation with Endosperm

To determine the fate of gibberellic acid (GA₃), solutions were incubated for 24 hours with or without barley endosperm and were subsequently applied to dwarf maize seedlings. Hormone activity appeared to increase as a result of incubation with endosperm. This apparent increase in GA₃ concentration was probably due to a synergistic interaction between GA₃ and endosperm constituents, particularly carbohydrate, released during the incubation period. It is concluded that relatively little hormone is inactivated during the initiation of endosperm mobilization.     

Plant Growth Substances Produced by Micro-organisms of Soil and Rhizosphere

S ummary : Micro-organisms isolated from rhizospheres and rhizoplanes of wheat plants, and from root-free soil, produced growth regulating substances with the properties of gibberellins and indolyl-3-acetic acid (IAA). Substances inhibiting extensions of pea plant internodes and lettuce hypocotyls were also produced, especially by bacteria from the root region of seedlings 6 days old. Bacteria producing growth promoting substances were most abundant on roots of older plants. Seedlings grown aseptically with added gibberellic acid (GA₃) and IAA, or grown with a soil inoculum, developed similarly and differed in their morphology from those grown aseptically without additives.     

Growth Activity of the Gibberellins of Dwarf French Bean, Potato, and Lettuce

Zones with similar Rf to gibberellic acid (GA) on chromatogramsof ethylacetate extracts of dwarf French bean and potato leavesand stems and lettuce heads all promoted growth of disks ofetiolated dwarf French bean leaves in darkness. The presence of a gibberellin was confirmed by tests with stemsof dwarf pea, but that the substance was not GA was shown bythe lack of effect in the tests with lettuce hypocotyls. Thetests with dwarf maize, effects on d-3 and d-5, but not d-1,identified it as gibberelliin A₅ (bean factor II). Materialthat promoted the growth of lettuce hypocotyls occurred in zonesclose to the starting-line of chromatograms of extracts of beanand lettuce leaves. Calibration of the growth tests with GA gave an estimate ofthe gibberellin content of the tissues extracted. In the dwarf-peatest the apparent gibberellin contents were similar to the valuesin the leaf-disk test, but very low values occurred in the maizetests.     

The Relation between Cell-wall and Protein Synthesis in Dwarf Pea Plants treated with Gibberellic Acid

The amount of protein and soluble nitrogen present in expandinginternodes of intact dwarf pea seedlings, was investigated atdifferent times after treatment of plants with gibberellic acid(GA). There was a marked increase in rate of protein synthesisfollowing GA treatment, but the rate of synthesis did not keeppace with internode expansion, so that the amount of proteinper unit length fell. The rate of cell-wall synthesis was alsoincreased, and, in contrast to protein, the amount of cell wallper unit length remained approximately constant during internodeexpansion. It is suggested that the increased rate of cell-wallsynthesis which follows GA treatment is mediated by a changein protein metabolism. The amount of soluble nitrogen presentin expanding internodes was also increased. There was littleeffect of GA upon the protein content of internodes which werealmost fully expanded at the time of treatment.     

Diversity of abundant mRNA sequences and patterns of protein synthesis in etiolated and greened pea seedlings

The diversity of abundant mRNA sequences in various parts of 4-d etiolated pea seedlings (Pisum sativum L. var. Rondo CB) was compared by a cell-free translation of the mRNAs in the presence of [³⁵S]methionine and by an analysis of the products by two-dimensional electrofocussing/ electrophoresis (2D separation). The various parts of the seedlings were also examined for the pattern of protein synthesis in vivo. Proteins were labeled by injection of [³⁵S]methionine into the cotyledons, followed by 2D separation of the products. Over 95% of the abundant mRNA sequences and newly synthesized abundant polypeptides were shared by all parts of etiolated seedlings, including the cotyledons. However, a few distinct differences were observed when comparing mRNAs of roots and shoots; the most prominent among these were a group of six abundant mRNA sequences found exclusively in shoots. Only about 30% of the polypeptides synthesized on isolated RNA could be traced in equivalent positions on the gels as the polypeptides synthesized in vivo. Analysis of total RNA from light-grown pea seedlings showed the appearance of some twenty-five translation products not found with total RNA from etiolated seedlings, while about nine other translation products disappeared. At least ten of the light-induced RNA sequences were also present after growth in low-intensity red light (>600 nm) and are therefore thought to be controlled by the phytochrome system. Comparison of 11-d light-grown pea plants with 4-d light-grown seedlings did not reveal additional translatable RNA sequences, indicating that the major morphogenetic changes that occur after 4 d are not accompanied by significant changes in the pattern of abundant RNA sequences.     

STS-95 space experiment for plant growth and development, and auxin polar transport.

The principal objective of the space experiment, BRIC-AUX on STS-95, was the integrated analysis of the growth and development of etiolated pea and maize seedlings in space, and the effect of microgravity conditions in space on auxin polar transport in the segments. Microgravity conditions in space strongly affected the growth and development of etiolated pea and maize seedlings. Etiolated pea and maize seedlings were leaned and curved during space flight, respectively. Finally the growth inhibition of these seedlings was also observed. Roots of some pea seedlings grew toward the aerial space of Plant Growth Chamber. Extensibilities of cell walls of the third internode of etiolated pea epicotyls and the top region of etiolated maize coleoptiles which were germinated and grown under microgravity conditions in space were significantly low. Activities of auxin polar transport in the second internode segments of etiolated pea seedlings and coleoptile segments of etiolated maize seedlings were significantly inhibited and extremely promoted, respectively, under microgravity conditions in space. These results strongly suggest that auxin polar transport as well as the growth and development of plants is controlled under gravity on the earth.