14C-Studies on Apple Trees. IV. Photosynthate Consumption in Fruits in Relation to the Leaf-Fruit Ratio and to the Leaf-Fruit Position

The photosynthate consumption in apple fruits in relation to the leaf/fruit ratio was studied in sections of branches of the Graasten and Golden Delicious varieties by exposure to ¹⁴CO₂during July and August. A significant, negative correlation was found between the fixation of ¹⁴C in the fruits in terms of percent of the total amount of ¹⁴C absorbed and the leaf/fruit ratio of the branch sections. The leaf area required for the saturation of one fruit was found to be ca. 190 and 230 cm² (14 and 17 leaves) in the case of Golden Delicious, and ca. 400 and 670 cm² (25 and 42 leaves) for Graasten in July and August, respectively, calculated under conditions of large leaf areas per fruit. In such cases a fairly good, reverse proportionality exists between the ¹⁴C fixation in the fruit in terms of percent and the leaf area expressed in multiples of saturation area. At low leaf/fruit ratios, however, the actual saturation area is found to be lower than the theoretically computed one. The translocation of the ¹⁴C assimilated in the leaves of the extension shoots or of spurs without fruits to fruits on other spurs was on the whole promoted by a decreasing leaf/fruit ratio in the parts in question; similarly the greatest transport took place on the side where the leaf/fruit ratio was lowest. The fixation was often greatest in the fruit closest to the treated leaves, hut in a number of cases the value was higher for the second closest or even further removed fruits. In this connection the importance of the size of the fruit and the vascular connections is discussed.     

The regulation of sweet cherry fruit abscission by polar auxin transport

Inconsistency of cropping is an important problem for UK sweet cherry production. Premature fruit abscission in Prunus can reduce yields severely, however, the environmental cues and hormonal signals that trigger abscission have not been identified. Auxin (IAA) is known to delay abscission by reducing the sensitivity of cells in the abscission zone to ethylene, a promoter of abscission. Therefore, the capacity for polar auxin transport (PAT) through sweet cherry pedicels was examined in relation to fruit abscission. Cherry ‘spurs’ (short shoots) with similar leaf areas and different fruit numbers were phloem-girdled to restrict assimilate movement. Abscission from spurs with many fruit (eight or more) occurred within 14 days of girdling, whereas abscission from spurs with few (two) fruit was minimal. The pedicels’ capacity for PAT in spurs with different fruit numbers was determined 1, 3 and 9 days after girdling (DAG). Fruit were analysed for endogenous IAA concentration 3, 5, 7 and 9 DAG. PAT inhibitors 2,3,5-triiodobenzoic acid or 1-N-naphthylphtalamic acid were applied to pedicels of fruit not expected to abscise, i.e. on spurs with few fruit. The effect of these inhibitors on fruit abscission was determined 14 DAG. The proportion of the transported [³H]-IAA was lower from the outset in pedicels from spurs with many fruit. By 9 DAG, symptoms of fruit abscission were apparent and 40% less [³H] -IAA was transported through pedicels on spurs with many fruit. Fruit endogenous IAA concentrations were similar in the two groups of spurs. Application of PAT inhibitors shortly after girdling increased fruit abscission by 30%. The results suggest that although a decline in PAT is not the only cause of fruit abscission, the maintenance of PAT contributes to fruit retention.     

Adventitious root formation in relation to irradiance and auxin supply

High irradiance during treatment of mung bean cuttings favours root formation in response to supplied auxin, whether the latter is IAA or IBA. On the other hand it is inhibitory towards root formation in the absence of supplied auxin. Light promotes the uptake of¹⁴C-IAA into cuttings and its upward movement into the leaves. When¹⁴C-IAA is applied to leaves of cuttings high irradiance favours movement of radioactivity into the epicotyl and hypocotyl. This movement is also enhanced by concomitant supply of IBA to the base of the cuttings. The irradiance under which stock plants are raised also affects the extent of root formation on cuttings. When cuttings are held in darkness without a supply of exogenous auxin they root best if prepared from seedlings raised under high irradiance. However, transport of¹⁴C-IAA out of leaves of cuttings is favoured when cuttings are prepared from seedlings grown under low irradiance. These observations are discussed in relation to auxin transport, photodestruction and, possibly, metabolism.     

The free movement of14C-labelled organic compounds from intact apple seeds to growing fruitlets and shoots

Indole-3-acetic acid (IAA-1¹⁴C, IAA-2¹⁴C) and gamma-aminobutyric acid labelled with¹⁴C were applied in lanoline to the surface of intact seeds or inserted into seeds of growing apple fruitlets or to the cut surface of the pericarp. Their translocation in trees was checked by means of autoradiography or by a low background Geiger-Müller counter. Auxin applied to the top of intact seeds, or inserted into the seeds was translocated and distributed within the pericarp, spur tissues and shoots below. The translocation of gamma-aminobutyric acid from seeds suggests that the capacity of apple seeds for extensive interchange with the surrounding tissues of the fruit and the spur is more general than suspected. A much poorer or no translocation to the spur was found when radioactive compounds were applied to the cut surface of the pericarp, while the seeds were left untouched. The results suggest two mechanisms of metabolite translocation in the apple fruit: two-directional for seeds and one-directional for the pericarp.     

Evidence of 4-Cl-IAA and IAA Bound to Proteins in Pea Fruit and Seeds

The auxins 4-chloroindole-3-acetic acid (4-Cl-IAA) and indole-3-acetic acid (IAA) occur naturally in pea vegetative and fruit tissues (Pisum sativum L.). Previous work has shown that 4-Cl-IAA can substitute for the seeds in the stimulation of pea pericarp growth, whereas IAA is ineffective. Both auxins are found as free acids and as low-molecular-weight conjugates from organic solvent-soluble extracts from pea fruit. Here we present evidence for an additional conjugated auxin species that was not soluble in organic solvent and yielded 4-Cl-IAA and IAA after strong alkaline hydrolysis, suggestive of auxin attachment to pea seed and pericarp proteins. The solvent-insoluble conjugated 4-Cl-IAA in young pericarp was on average 15-fold greater than solvent-soluble 4-Cl-IAA. The solvent-insoluble conjugated IAA was approximately half the levels reported for the solvent-soluble IAA fraction. To identify putative 4-Cl-IAA-bound proteins, polyclonal antibodies were raised to 4-Cl-IAA linked to bovine serum albumin protein (BSA). Immunoblots probed with anti-4-Cl-IAA-BSA antiserum detected three to four unique bands (32–40 kDa) in primarily maternal tissues, and a different set of protein bands were detected in mainly embryonic tissues (ca. 65–74 kDa in mature seed). 4-Cl-IAA and IAA were also identified from protein fractions separated by polyacrylamide gel electrophoresis using GC-MS. These data show that the majority of 4-Cl-IAA, the growth-active auxin in young pea pericarp, and significant levels of IAA are linked to protein fractions. Auxin-proteins may function in regulation of free bioactive 4-Cl-IAA and IAA levels, and/or 4-Cl-IAA or IAA may be targeted to specific proteins post-translationally to modify protein function or stability.     

TRANSPORT OF IAA AND ACC IN FLORAL ORGANS OF IPOMOEA NIL (CONVOLVULACEAE)

The involvement of the stamens as transporters of plant growth regulators in flowers was examined by measuring the movement of ¹⁴C-indole-3-acetic acid (IAA) and ^l4C-l-aminocyclopropane-1-carboxylic acid (ACC) through floral organs of Ipomoea nil. During the transport of ¹⁴C-IAA through isolated filament segments, the polar accumulation of ¹⁴C-IAA in receiver blocks increased with time during filament development, which correlated with polar efflux rates at older stages of filament development. An inhibitor of polar IAA transport, 2,3,5-triiodobenzoic acid, disrupted the polarity of auxin transport by reducing the movement of ¹⁴C- IAA from filaments into receiver blocks. Transport of both ¹⁴C-IAA and ^l4C-ACC through filaments into other floral organs also was monitored in isolated flower buds in the laboratory and intact buds in the greenhouse. In isolated and intact buds 21 hr before anthesis, substantially higher levels of isotope were recovered in corolla tissue when ¹⁴C-ACC was transported through the filaments than when ¹⁴C-IAA was transported from the filaments. In isolated buds, substantial levels of both isotopes accumulated in the pistil (69 hr and 45 hr before anthesis), but minimal amounts were observed in receptacle and calyx tissues (69 hr to 21 hr before anthesis). In intact buds, high levels of both isotopes were recovered in receptacle, calyx, and pistil tissues (69 hr to 21 hr before anthesis). The results from this study support the hypothesis that Ipomoea stamens are transporters for ACC and IAA to regulate ethylene production in the corolla and other floral tissues.     

Indole-3-acetic acid concentration and ethylene evolution during early fruit development in peach

Ethylene evolution was measured from greenhouse-grown Jerseyglo peach fruits beginning 29 days after anthesis. Indole-3-acetic acid (IAA) levels were measured in the pericarp and seed tissues of individual fruits on a single shoot when variable ethylene evolution was noted. Despite hand-pollinating all flowers on the same day, variability within the shoot existed in fruit fresh weight, IAA levels, and ethylene evolution. Seed IAA concentration increased as fruit and seed fresh weight increased and ranged from 106 to 1572 ng. g^–1. As pericarp fresh weight increased, IAA levels in this tissue decreased. Ethylene evolution rates ranged from 0.21 to 1.07 nl. g.^–1 h^–1 and were not correlated with IAA concentration in seed, pericarp, or the whole fruit. High rates of ethylene evolution from the whole fruit occurred prior to increased IAA concentration in the seed.Fruits were excised from field-grown Redskin peach trees beginning 40 days after full bloom. Fruits from field sampled shoots appeared to be more physiologically advanced than the greenhouse-grown Jerseyglo fruits. Pericarp IAA concentration was low, ranging from 2.8 to 6.5 ng. g^–1. Seed concentrations accounted for 75% of the IAA found in the fruit and ranged from 239 to 1042 ng. g^–1. As with greenhouse-grown samples, whole fruit IAA concentration tended to decrease as fruits increased in fresh weight.     

Disruption of the Polar Auxin Transport System in Cotton Seedlings following Treatment with the Defoliant Thidiazuron

The effect of the defoliant thidiazuron (TDZ) on basipetal auxin transport in petiole segments isolated from cotton (Gossypium hirsutum L. cv LG102) seedlings was examined using the donor/receiver agar block technique. Treatment of intact seedlings with TDZ at concentrations of 1 micromolar or greater resulted in a dose-dependent inhibition of ¹⁴C-IAA transport in petiole segments isolated 1 or 2 days after treatment. Using 100 micromolar TDZ, the inhibition was detectable 19 hours after treatment and was complete by 27 hours. Both leaves and petiole segments exhibited a marked increase in ethylene production following treatment with TDZ at concentrations of 0.1 micromolar or greater. The involvement of ethylene in this TDZ response was evaluated by examining the effects of two inhibitors of ethylene action: silver thiosulfate, 2,5-norbornadiene. One day after treatment, both inhibitors effectively antagonized the TDZ-induced inhibition of auxin transport. Two days after TDZ treatment both inhibitors were ineffective. The decrease in IAA transport in TDZ treated tissues was associated with increased metabolism of IAA. The transport of ¹⁴C-2,4-dichlorophenoxyacetic acid was also inhibited by TDZ treatment. This inhibition was not accompanied by increased metabolism. Incorporation of TDZ into the receiver blocks had no effect on auxin transport. The ability of the phytotropin N-1-naphthylphthalamic acid to stimulate IAA uptake from a bathing medium was reduced in TDZ-treated tissues. This reduction is thought to reflect a decline in the auxin efflux system following TDZ treatment.     

The biosynthesis and translocation of 14C-IAA in Ricinus communis

The biosynthesis of ¹⁴C-IAA from ¹⁴C-tryptophan applied to abraded leaves of Ricinus communis and its subsequent export through the phloem were studied. Phloem sap was collected at intervals from incisions made in the stem below the IAA fed leaf. Any upward movement of label through the phloem or downward movement of phloem mobile compounds from leaves above the treated one were restricted by bark-ringing the plants.TLC and HPLC analyses of the collected sap indicate that some conversion of ¹⁴C-tryptophan to ¹⁴C-IAA had occurred. Subsequent GC-MS analysis of the HPLC purified samples of phloem sap revealed high levels of endogenous IAA transported from the fed leaf. The high ratio of unlabelled/labelled IAA in the phloem sap makes unequivocal confirmation by GC-MS of the predicted biosynthesis of ¹⁴C-IAA impossible. It is postulated that IAA is synthesised from tryptophan in mature leaves and exported to developing sink tissues with the flow of photoassimilates in the phloem.     

Levels of indole-3-acetic acid in vigorous and genetic dwarf apple trees

Seasonal levels of indole-3-acetic acid (IAA) present in buds, meristematic tissues, and leaves of 1-year-old shoots of two selections (vigorous and dwarf) of a F₂ generation apple population with original “Goldspur” × “Redspur” parentage were determined using gas chromatography—selected ion monitoring—mass spectrometry (GC-SIM-MS) with a¹³C-IAA internal standard. A comparison of IAA levels through one growing season was made in the shoots of two trees differing in growth. The levels of IAA in the leaves of the vigorous tree were higher in the early and late growing seasons, as compared to those in the dwarf tree. Levels of IAA in buds of the dwarf tree shoots were found to be higher than in those of the vigorous tree shoots. On leave from Indian Institute of Horticultural Research, Bangalore-560080, India.     

Induced parthenocarpy—a way of changing the levels of endogenous hormones in tomato fruits (Lycopersicon esculentum Mill.) 1. Extractable hormones

Ethylene, indol-3-acetic acid (IAA), gibberellin-like substances (GAs) and abscisic acid (ABA) were analysed in extracts from normal, seed-containing and parthenocarpic tomato fruits throughout fruit development. Parthenocarpic fruit growth was induced with an auxin (4-CPA), morphactin (CME) or gibberellic acid (GA₃) and compared with that of pollinated control fruits. Fruit growth was only affected by the treatment with GA₃, decreasing size and fresh weight by 60%. The peak sequence of hormones during fruit development was ethylene-GAs-IAA-ABA. Seeded fruits contained the highest levels of IAA and ABA but the lowest levels of GAs. Also, in seeded fruits, a high proportion of IAA and ABA was found in the seeds whereas this was not the case for GAs.Hormone levels of tomato fruits may be successfully, easily and reproducibly altered by inducing parthenocarpic fruit growth and thus eliminating development of seeds which are a major source of hormone synthesis. In spite of markedly changed hormone levels, there was no obvious relationship between fruit growth and extractable hormones per se. However, the results indicate that a high ratio of GAs: auxins is unfavourable for growth of tomato fruits.     

Hormone and seed-specific regulation of pea fruit growth

Growth of young pea (Pisum sativum) fruit (pericarp) requires developing seeds or, in the absence of seeds, treatment with gibberellin (GA) or auxin (4-chloroindole-3-acetic acid). This study examined the role of seeds and hormones in the regulation of cell division and elongation in early pea fruit development. Profiling histone H2A and gamma-tonoplast intrinsic protein (TIP) gene expression during early fruit development identified the relative contributions of cell division and elongation to fruit growth, whereas histological studies identified specific zones of cell division and elongation in exocarp, mesocarp, and endocarp tissues. Molecular and histological studies showed that maximal cell division was from -2 to 2 d after anthesis (DAA) and elongation from 2 to 5 DAA in pea pericarp. Maximal increase in pericarp gamma-TIP message level preceded the maximal rate of fruit growth and, in general, gamma-TIP mRNA level was useful as a qualitative marker for expanding tissue, but not as a quantitative marker for cell expansion. Seed removal resulted in rapid decreases in pericarp growth and in gamma-TIP and histone H2A message levels. In general, GA and 4-chloroindole-3-acetic acid maintained these processes in deseeded pericarp similarly to pericarps with seeds, and both hormones were required to obtain mesocarp cell sizes equivalent to intact fruit. However, GA treatment to deseeded pericarps resulted in elevated levels of gamma-TIP mRNA (6 and 7 DAA) when pericarp growth and cell enlargement were minimal. Our data support the theory that cell division and elongation are developmentally regulated during early pea fruit growth and are maintained by the hormonal interaction of GA and auxin.     

Systemic Invasion of Immature Sweet Cherry Fruit by Pseudomonas syringae pv. morsprunorum Through Blossoms1

Cherry blossoms inoculated with a rifampicin-resistant strain of Pseudomonas syringae pv. morsprunorum died or gave rise to fruits containing necrotic spots at or near the blossom ends. Scanning electron microscopy of developing fruits indicated that the pathogen had invaded the entire pericarp, including the endocarp. Bacteria also spread to the fruit stalk and, to a lesser extent, to the spurs. Mesocarp cells below the lesion collapsed. Infected fruit, stalks, and spurs contained, respectively, ca. 10⁹, 10⁷, and 10² colony forming units of P. syringae pv. morsprunorum as determined by a dilution plate method on an agar medium supplemented with 50 μg/ml rifampicin. This is the first report of systemic spread of P. syringae from blossoms to developing fruit of a deciduous crop.     

Diurnal water balance of the cowpea fruit

The vascular network of the cowpea (Vigna unguiculata [L.] Walp.) fruit exhibits the anatomical potential for reversible xylem flow between seeds, pod, and parent plant. Feeding of cut shoots with the apoplast marker acid fuchsin showed that fruits imported regularly via xylem at night, less frequently in early morning, and only rarely in the afternoon. The dye never entered seeds or inner dorsal pod strands connecting directly to seeds. Root feeding (early morning) of intact plants with ³²PO₄ or ³H₂O rapidly (20 min) labeled pod walls but not seeds, consistent with uptake through xylem. Weak subsequent (4 hours) labeling of seeds suggested slow secondary exchange of label with the phloem stream to the fruit. Vein flap feeding of subtending leaves with [¹⁴C]sucrose, ³H₂O, and ³²PO₄ labeled pod and seed intensely, indicating mass flow in phloem to the fruit. Over 90% of the ¹⁴C and ³H of fruit cryopuncture phloem sap was as sucrose and water, respectively. Specific ³H activities of transpired water collected from fruits and peduncles were assayed over 4 days after feeding ³H₂O to roots, via leaf flaps, or directly to fruits. The data indicated that fruits transpired relatively less xylem-derived (apoplastic) water than did peduncles, that fruit and peduncle relied more heavily on phloem-derived (symplastic) water for transpiration in the day than at night, and that water diffusing back from the fruit was utilized in peduncle transpiration, especially during the day. The data collectively support the hypothesis of a diurnally reversing xylem flow between developing fruit and plant.     

Morphology and phenology of seeds and whole fruit eaten by Milne-Edwards’ sifaka,Propithecus diadema edwardsi,in Ranomafana National Park,Madagascar

Although it is an anatomical folivore, the diet of the Milne-Edwards’ sifaka (Propithecus diadema edwardsiA. Grandidier, 1871) in Ranomafana National Park contained 35% seeds, 30% whole fruit, and 28% leaves. Plant species used as seed sources differed from those used as whole fruit sources in terms of temporal variation in consumption, taxonomic affiliation, morphology, and phenology. Although seeds were destroyed in both exploitation styles used by the sifakas—seed and whole fruit-eating—the gross morphology of species used as seed sources conformed to the complex of traits typical for fruits experiencing seed predation, while species used as whole fruit sources conformed to traits typical for fruits that do not experience predispersal predation. Many of the 19 plant species from which the seed was extracted and eaten contained a single seed with moderate testa thickness, and fruits containing this type of seed were medium-sized with dry or fibrous flesh, moderate skin thickness, and a dull color. In contrast, brightly colored, juicy fruits with minimally protected seeds were characteristic of the 38 plant species from which both pericarp and seed were eaten. Compared to transectwide measures of fruit availability or patterns restricted to whole fruit sources, fewer species of seed sources produced fruit per month and fruiting activity was more seasonal.     

The Role of Endogenous Auxins and Ethylene in the Formation of Adventitious Roots and Hypocotyl Hypertrophy in Flooded Sunflower Plants (Helianthus annuus)

The role of ethylene and auxins in flood-induced adventitious root formation and hypocotyl hypertrophy in sunflower (Helianthus annuus L. cv. Russian) plants was studied. Flooding without aeration (F) resulted in a steady increase in ethylene in hypocotyls, and flooding with aeration (FA) caused a transient increase. Low light intensity increased ethylene levels but decreased adventitious root formation. Treatment of shoots with benzyladenine (BA) increased ethylene content in non-flooded (NF) but not in F or FA shoots. Twenty-four hours of flooding brought about a rise of endogenous indole-acetic acid (IAA) in hypocotyls. ¹⁴C-IAA applied to the shoot accumulated more in F and FA hypocotyls than in NF hypocotyls, and BA reduced this accumulation. There was less IAA metabolism in F and FA than in NF hypocotyls. Tri-iodo benzoic acid (TIBA) applied to the hypocotyls of F plants inhibited root production. Benzyladenine (BA) applied to the leaves had similar effect but was not effective when supplied to the shoot apex. BA did not inhibit flood-induced hypocotyl hypertrophy. Ethrel did not affect adventitious root formation in NF plants but did increase hypocotyl thickening. It is concluded that flood-induced adventitious root formation is stimulated primarily by an accumulation of auxins in the hypocotyls. Increases in ethylene might cause this auxin build up. Hypocotyl hypertrophy is presently thought to be the result of an interaction of auxin and ethylene with ethylene being the major factor.     

Developmental and Hormonal Regulation of Gibberellin Biosynthesis and Catabolism in Pea Fruit

In pea (Pisum sativum), normal fruit growth requires the presence of the seeds. The coordination of growth between the seed and ovary tissues involves phytohormones; however, the specific mechanisms remain speculative. This study further explores the roles of the gibberellin (GA) biosynthesis and catabolism genes during pollination and fruit development and in seed and auxin regulation of pericarp growth. Pollination and fertilization events not only increase pericarp PsGA3ox1 message levels (codes for GA 3-oxidase that converts GA₂₀ to bioactive GA₁) but also reduce pericarp PsGA2ox1 mRNA levels (codes for GA 2-oxidase that mainly catabolizes GA₂₀ to GA₂₉), suggesting a concerted regulation to increase levels of bioactive GA₁ following these events. 4-Chloroindole-3-acetic acid (4-Cl-IAA) was found to mimic the seeds in the stimulation of PsGA3ox1 and the repression of PsGA2ox1 mRNA levels as well as the stimulation of PsGA2ox2 mRNA levels (codes for GA 2-oxidase that mainly catabolizes GA₁ to GA₈) in pericarp at 2 to 3 d after anthesis, while the other endogenous pea auxin, IAA, did not. This GA gene expression profile suggests that both seeds and 4-Cl-IAA can stimulate the production, as well as modulate the half-life, of bioactive GA₁, leading to initial fruit set and subsequent growth and development of the ovary. Consistent with these gene expression profiles, deseeded pericarps converted [¹⁴C]GA₁₂ to [¹⁴C]GA₁ only if treated with 4-Cl-IAA. These data further support the hypothesis that 4-Cl-IAA produced in the seeds is transported to the pericarp, where it differentially regulates the expression of pericarp GA biosynthesis and catabolism genes to modulate the level of bioactive GA₁ required for initial fruit set and growth.     

Indole-3-acetic acid metabolism and growth in young kiwifruit berry

The patterns of auxin concentration and metabolism were investigated in distinct kiwifruit portions and compared with the rate of fruit growth during early developmental stages. Indole-3-acetic acid (IAA) level was higher in inner fruit tissues, particularly in younger fruit, while the hormone was barely detectable in outer tissues. Modulation of free IAA concentration did not appear to depend tightly on conjugation of the hormone. Despite the lack of a strong correlation between the levels of IAA and enzymes involved in its catabolism, in some portions of the fruit a low hormone level corresponded to a higher IAA degradation activity. An inverse correlation was also observed between hormone levels and the appearance/increase in some bands with high mobility in peroxidase gel activity assay. Phenols, compounds with a potential auxin-protecting activity, appeared to be involved mostly in photoprotection of the fruit than in the regulation of IAA levels. Beyond catabolism and conjugation, other metabolic pathways, particularly those occurring in the developing seeds, may have decisively influenced auxin levels in fruit tissues, as well as the amount of the hormone exported from the fruit. The latter, estimated by analyzing the concentration of IAA in the sap exuded from the pedicel, showed a time course which was similar to that displayed by inner fruit tissues. Furthermore, similarities were found between the pattern of IAA concentration in inner fruit tissues and fruit growth rate. The possible role of IAA in promoting growth during early fruit development is discussed.