Gravitropism in Higher Plant Shoots : IV. Further Studies on Participation of Ethylene

Ethylene at 1.0 and 10.0 cubic centimeters per cubic meter decreased the rate of gravitropic bending in stems of cocklebur (Xanthium strumarium L.) and tomato (Lycopersicon esculentum Mill), but 0.1 cubic centimeter per cubic meter ethylene had little effect. Treating cocklebur plants with 1.0 millimolar aminoethoxyvinylglycine (AVG) (ethylene synthesis inhibitor) delayed stem bending compared with controls, but adding 0.1 cubic centimeter per cubic meter ethylene in the surrounding atmosphere (or applying 0.1% ethephon solution) partially restored the rate of bending of AVG-treated plants. Ethylene increases in bending stems, and AVG inhibits this. Virtually all newly synthesized ethylene appeared in bottom halves of horizontal stems, where ethylene concentrations were as much as 100 times those in upright stems or in top halves of horizontal stems. This was especially true when horizontal stems were physically restrained from bending. Ethylene might promote cell elongation in bottom tissues of a horizontal stem or indicate other factors there (e.g. a large amount of `functioning' auxin). Or top and bottom tissues may become differentially sensitive to ethylene. Auxin applied to one side of a vertical stem caused extreme bending away from that side; gibberellic acid, kinetin, and abscisic acid were without effect. Acidic ethephon solutions applied to one side of young seedlings of cocklebur, tomato, sunflower (Helianthus annuus L.), and soybean (Glycine max [L.] Merr.) caused bending away from that side, but neutral ethephon solutions did not cause bending. Buffered or unbuffered acid (HCl) caused similar bending. Neutral ethephon solutions produced typical ethylene symptoms (i.e. epinasty, inhibition of stem elongation). HCl or acidic ethephon applied to the top of horizontal stems caused downward bending, but these substances applied to the bottom of such stems inhibited growth and upward bending—an unexpected result.     

Laser micromarking technique in studying the negative gravitropism in pea stem

A laser micromarking technique on plant epidermis was developed to study how a plant can reduce the stress in bending behavior by controlling the growth and morphogenesis. The negative gravitropism in a pea seedling (Pisum sativum L.) was discussed based on the time-dependent displacement of laser marking points which were formed by spatially-selective laser ablation of the cuticle layer that covers the outer surface of a plant. The elongation of the stem in the horizontal direction was remarkable in the first half of the gravitropism. The elongation percentages of the stem length between laser-marking points at around upper surface, middle, and bottom surface were evaluated to be 2.57, 4.87, and 7.70%, respectively. The characteristic feature of the stem bending in gravitropism is the elongation even at the upper surface region, that is, inside of the bending. This is a different feature from cantilever beams for structural materials like metals and polymers, where the compression of the upper surface and elongation of the bottom surface are caused by bending. Another laser micromarking technique was developed to improve the resolution of a dot-matrix pattern by fluorescent material transfer to a plant through a masking film with a micro-hole matrix pattern. Similar time-dependent displacement behavior was observed for a fluorescent dot-marked stem showing a feedback control loop in the mechanical optimization. These results suggested that plants solve the problem of the stress in stem bending through growth. The laser micromarking is an effective method for studying the mechanical optimization in plants.     

Gravitropism in Higher Plant Shoots : II. Dimensional and Pressure Changes during Stem Bending

Dimensional changes during gravitropic bending of cocklebur (Xanthium strumarium L.) dicot stems were measured using techniques of stereo photogrammetry. The differential growth is from an increased growth rate on the bottom of the stem and a stopping or contraction of the top.

Contraction of the top was especially evident upon release and immediate bending of horizontal stems that had been restrained between stiff wires for 36 hours. The energy for this could have been stored in both the top and bottom, since the bottom elongated, and the top contracted.

Forces developed during bending were measured by fastening a stem tip to the end of a bar with attached strain gauges and recording electrical output from the strain gauges. Restrained mature cocklebur stems continued to accumulate potential energy for bending for about 120 hours, after which the recorded force reached a maximum.

Pressures within castor bean (Ricinus communis L.) stems were also measured with 3.5-millimeter diameter pressure transducers. As expected, the pressure on the bottom of the restrained plants increased with time; pressures decreased in vertical controls, tops of restrained stems, and bottoms of free-bending stems. Pressures increased in tops of free-bending stems. When restrained plants were released, pressure on the bottom decreased and pressure on the top increased. Results suggest a possible role for cell contraction in the top of stems bending upward in response to gravity.

     

Growth patterns and endogenous indole-3-acetic acid concentrations in current-year coppice shoots and seedlings of two Betula species

Apical dominance, internode elongation, radial growth and xylem cell size in coppice and apical shoots of Betula pubescens B. Pendula were determined and related to endogenous indole-3-acetic acid (IAA) levels, measured by gas chromatography-selected ion monitoring in the apical bud and at three positions along the stem. The effects of defoliation and debudding on morphological and anatomical characters and endogenous IAA levels were also investigated. The coppice shoots displayed superior stem elongation and increased branching during the initial phase of growth, after which their growth pattern was similar to that of the seedlings; however, their radial growth was greater throughout the experiment. Both plant types produced smaller-sized xylem cells at the top of the shoot than at the bottom with coppice shoots tending to form larger tracheids and smaller vessels than the seedlings. There was no consistent difference in IAA concentration between the coppice shoots and the seedlings. Defoliation and debudding reduced the IAA level in the stem within 36 h and it was still low after 25 days. Although the extent of the IAA decrease was similar in both coppice shoots and seedlings, the treatments affected the morphological and anatomical characters differently in the two plant types. The results suggest that the observed differences between seedlings and coppice shoots were not mediated through a drastic change in IAA level.     

Cell elongation, turgor and osmotic pressure in developing sunflower hypocotyls

The relationship between cell elongation, change in turgor andcell osmotic pressure was investigated in the sub-apical regionof hypocotyls of developing sunflower seedlings (Helianthusannuus L.) that were grown in continuous white light. Cell turgorwas measured with the pressure probe. The same hypocotyl sectionswere used for determination of osmotic pressure of the tissuesap. Acceleration of cell elongation during the early phaseof growth was accompanied by a 25% decrease in both turgor andosmotic pressure. During the linear phase of growth both pressuresremained largely constant. The difference between turgor andosmotic pressure (water potential) was –0.10 to –0.13MPa. Excision of one cotyledon had no effect on growth, turgorand osmotic pressure. However, after removal of both cotyledonscell elongation ceased and a substantial decrease in both pressureswas measured. In addition, we determined the longitudinal tissuepressure in seedlings from which one or both cotyledons hadbeen removed. Tissue pressure and turgor were very similar quantitiesunder all experimental conditions. Our results demonstrate thatturgor and cell osmotic pressure show a parallel change duringdevelopment of the stem. Cessation of cell elongation afterremoval of the cotyledons is attributable to a decrease in turgor(tissue) pressure, which provides the driving force for growthin the hypocotyl of the intact plant. Key words: Cell elongation, Helianthus annuus, osmotic pressure, tissue pressure, turgor     

Fungal elicitor-induced retardation and its restoration of root growth in tobacco seedlings

In this study, we investigated responses of growing and intact tobacco (N. tabacum cv Xanthi) seedlings to a fungal elicitor, a xylanase from Trichoderma viride (TvX). In addition to the induction of defense gene expression, TvX treatment caused the retardation of growth of seedlings. In the TvX-treated seedlings, growth of primary roots was markedly reduced through repression of cell division and longitudinal cell elongation in a meristematic zone and an elongation zone, respectively. However, cell differentiation to form vascular bundles and root hairs continued. In the TvX-treated root cap, disappearance of starch granules in columella cells and aggregation of border cells were observed. Furthermore, the TvX-induced growth retardation was restored after removal of the elicitor, resulting in a plastic alteration of root architecture. Therefore, the fungal elicitor might act as an environmental cue that regulates root growth and development as well as the ordinary defense responses in plant seedlings. These findings suggest a novel aspect of plant growth regulation via a plant–microbe interaction in the rhizosphere.     

Gravitropism in Higher Plant Shoots : III. Cell Dimensions during Gravitropic Bending; Perception of Gravity

Cross and longitudinal sections were prepared for light microscopy from vertical control plants (Xanthium strumarium L. Chicago strain), free-bending horizontal stems, plants restrained 48 hours in a horizontal position, and plants restrained 48 hours and then released, bending immediately about 130°. Top cells of free-bending stems shrink or elongate little; bottom cells continue to elongate. In restrained stems, bottom cells elongate some and increase in diameter; top cells elongate about as much but decrease in diameter. Upon release, bottom cells elongate more and decrease in diameter, while top cells shorten and increase in diameter, accounting for the bend. During restraint, bottom cells take up water while tissue pressures increase; top cells fail to take up water although tissue pressures are decreasing.

Settling of amyloplasts was observed in cells of the starch sheath.

Removal of different amounts of stem (Xanthium; Lycopersicon esculentum Miller, cv Bonny Best; Ricinus communis L. cv Yolo Wonder) showed that perception of gravity occurs in the bending (elongation) zone, although bending of fourth and fifth internodes from the top was less than in uncut controls. Uniform application of 1% indoleacetic acid in lanolin to cut stem surfaces partially restored bending. Reversing the gradient in tension/compression in horizontal stems (top under compression, bottom under tension) did not affect gravitropic bending.

     

Photophysiology of the Elongated Internode (ein) Mutant of Brassica rapa: ein Mutant Lacks a Detectable Phytochrome B-Like Polypeptide

Several phytochrome-controlled processes have been examined in etiolated and light-grown seedlings of a normal genotype and the elongated internode (ein/ein) mutant of rapid-cycling Brassica rapa. Although etiolated ein seedlings displayed normal sensitivity to prolonged far-red light with respect to inhibition of hypocotyl elongation, expansion of cotyledons, and synthesis of anthocyanin, they displayed reduced sensitivity to prolonged red light for all three of these deetiolation responses. In contrast to normal seedlings, light-grown ein seedlings did not show a growth promotion in response to end-of-day far-red irradiation. Additionally, whereas the first internode of light-grown normal seedlings showed a marked increase in elongation in response to reduced ratio of red to far-red light, ein seedlings showed only a small elongation response. When blots of protein extracts from etiolated and light-treated ein and normal seedlings were probed with monoclonal antibody to phytochrome A, an immunostaining band at about 120 kD was observed for both extracts. The immunostaining intensity of this band was substantially reduced for extracts of light-treated normal and ein seedlings. A mixture of three monoclonal antibodies directed against phytochrome B from Arabidopsis thaliana immunostained a band at about 120 kD for extracts of etiolated and light-treated normal seedlings. This band was undetectable in extracts of ein seedlings. We propose that ein is a photoreceptor mutant that is deficient in a light-stable phytochrome B-like species.     

Photophysiology and phytochrome content of long-hypocotyl mutant and wild-type cucumber seedlings

Photomorphogenetic responses have been studied in a cucumber (Cucumis sativus L.) mutant (lh), which has long hypocotyls in white light (WL). While etiolated seedlings of this mutant have a similar phytochrome content and control of hypocotyl elongation as wild type, deetiolation is retarded and WL-grown seedlings show reduced phytochrome control. Spectrophotometric measurements exhibit that WL-grown tissues of the lh mutant (flower petals and Norflurazon-bleached leaves) contain 35 to 50% of the phytochrome level in the wild type. We propose that this is a consequence of a lack of light-stable phytochrome, in agreement with our hypothesis proposed on the basis of physiological experiments. The lh mutant lacks an end-of-day far-red light response of hypocotyl elongation. This enables the end-of-day far-red light response, clearly shown by the wild type, to be ascribed to the phytochrome, deficient in the lh mutant. Growth experiments in continuous blue light (BL) and continuous BL + red light (RL) show that when RL is added to BL, hypocotyl growth remains inhibited in the wild type, whereas the lh mutant exhibits significant growth promotion compared to BL alone. It is proposed that the hypocotyls fail to grow long in low fluence rate BL because photosynthesis is insufficient to sustain growth.     

New plant growth promoters,repraesentins A,B and C,from Lactarius repraesentaneus

Three new plant growth regulatory sesquiterpenoids were isolated from the Lactarius repraesentaneus fungus. Their structures were elucidated to be a protoilludene sesquiterpene, namely repraesentin A (1), and two related sesquiterpenes, namely repraesentins B (2) and C (3). Compounds 1-3 showed promotion activities toward the radicle elongation of lettuce seedlings by 136%, 118% and 184% at 67 ppm, respectively.     

Lunisolar tidal force and the growth of plant roots, and some other of its effects on plant movements

Background

Correlative evidence has often suggested that the lunisolar tidal force, to which the Sun contributes 30 % and the Moon 60 % of the combined gravitational acceleration, regulates a number of features of plant growth upon Earth. The time scales of the effects studied have ranged from the lunar day, with a period of approx. 24·8 h, to longer, monthly or seasonal variations.

Scope

We review evidence for a lunar involvement with plant growth. In particular, we describe experimental observations which indicate a putative lunar-based relationship with the rate of elongation of roots of Arabidopsis thaliana maintained in constant light. The evidence suggests that there may be continuous modulation of root elongation growth by the lunisolar tidal force. In order to provide further supportive evidence for a more general hypothesis of a lunisolar regulation of growth, we highlight similarly suggestive evidence from the time courses of (a) bean leaf movements obtained from kymographic observations; (b) dilatation cycles of tree stems obtained from dendrograms; and (c) the diurnal changes of wood–water relationships in a living tree obtained by reflectometry.

Conclusions

At present, the evidence for a lunar or a lunisolar influence on root growth or, indeed, on any other plant system, is correlative, and therefore circumstantial. Although it is not possible to alter the lunisolar gravitational force experienced by living organisms on Earth, it is possible to predict how this putative lunisolar influence will vary at times in the near future. This may offer ways of testing predictions about possible Moon–plant relationships. As for a hypothesis about how the three-body system of Earth–Sun–Moon could interact with biological systems to produce a specific growth response, this remains a challenge for the future. Plant growth responses are mainly brought about by differential movement of water across protoplasmic membranes in conjunction with water movement in the super-symplasm. It may be in this realm of water movements, or even in the physical forms which water adopts within cells, that the lunisolar tidal force has an impact upon living growth systems.     

Repraesentins D, E and F, new plant growth promoters from Lactarius repraesentaneus

Three new plant growth regulatory sesquiterpenes were isolated from the Lactarius repraesentaneus fungus. Their structures were elucidated to be lactarane sesquiterpenes, namely repraesentins D (1) and E (2), and a protoilludane-related sesquiterpene, namely repraesentin F (3). Repraesentin E (2) showed the strongest promotion activity, 164% at 3.6 microM, of the three compounds toward the radicle elongation of lettuce seedlings.     

Changes in cucumber hypocotyl cell wall dynamics caused by Azospirillum brasilense inoculation

We previously reported that Azospirillum brasilense induced a more elastic cell wall and a higher apoplastic water fraction in both wheat coleoptile and flag leaf. These biophysical characteristics could permit increased growth. Knowledge of the biochemical effects the bacteria could elicit in plant cell walls and how these responses change plant physiology is still scarce. The objective of this work was to analyze whether A. brasilense Sp245 inoculation affected elongation and extensibility of growing cucumber (Cucumis sativus) hypocotyls and ionically bound cell wall peroxidase activities. Hypocotyl tip and basal segments were excised from A. brasilense Sp245-inoculated cucumber seedlings growing in darkness under hydroponic conditions. Elongation, cell wall extensibility, cell wall peroxidase activities against ferulic acid and guaiacol and NADH oxidase activities were analyzed. Azospirillum-inoculated cucumber seedlings grew bigger than non-inoculated ones. Dynamic cell wall differences were detected between inoculated and non-inoculated hypocotyls. They included greater acid-induced cell wall extension and in vivo elongation when incubated in distilled water. Although there was no difference between treatments in either region of the hypocotyl NADH oxidase and ferulic acid peroxidase activities were lower in both regions in inoculated seedlings. These lesser activities could be delaying the stiffening of cell wall in inoculated seedlings. These results showed that the cell wall is a target for A. brasilense growth promotion.     

Reproduction of Heterodera zeae and Its Suppression of Corn Plant Growth as Affected by Temperature

Reproduction of the corn cyst nematode (Heterodera zeae) and its effect on growth of corn (Zea mays) was studied in plant growth chambers at 24, 27, 30, 33, and 36 C. Reproduction of H. zeae increased directly with increase in temperature from 24 to 36 C. Fifteen-cm-d pots of corn seedlings inoculated with a mixture of 5,000 eggs + J2 and maintained for 8 weeks in growth chambers contained an average of 7,042 cysts + females at 36 C, but only 350 cysts + females at 24 C. Fresh weights of plants without nematodes were highest at 27 C and lowest at 36 C. Nematodes suppressed plant fresh weight by an average of 30% at 27 C and by 27% at 33 C but did not suppress plant weight at 36 C. Heterodera zeae has the highest reported temperature optimum for reproduction of any cyst nematode.     

Growth, in vivo extensibility, and tissue tension in developing pea internodes

The relationship between growth, in vivo extensibility, and tissue tension in the first 3 internodes of 5, 6, and 7 day-old pea plants (Pisum sativum L. cv Alaska), grown under continuous red light was investigated. The upper 15 millimeters of each internode was marked with ink and its elongation growth measured over the next subsequent 8 hours. In vivo extensibility was measured by stretching living tissue at constant force (creep test) in a custom-built extensiometer. Tissue tension was determined by (a) measuring the rate of expansion of the isolated cortical cylinder after adding water and the amount of contraction of the epidermis after peeling, and (b) by use of the `split section test.' A good correlation between rate of elongation growth, in vivo extensibility, and tissue tension was established. The epidermis peeled from the growing third internode of 7 day-old plants and measured immediately showed a plastic extensibility (E<sub>pl</sub> twice that of peels from nongrowing excised sections. This high E<sub>pl</sub>-value was lost on incubation of the sections in distilled water, and was subsequently restored by incubating the sections in auxin (indole-3-acetic acid). We conclude that the in situ growth of the internodes is a function of tissue-tension, which provides the driving force of organ growth, and the extensibility (E<sub>pl</sub> of the outer epidermal wall, which is in the growing plant in a `loosened' state. We furthermore suggest that in the intact plant auxin is causally involved in the wall loosening process in the epidermis.     

Phytochrome-mediated phototropism in de-etiolated seedlings : occurrence and ecological significance

Phototropic responses to broadband far red (FR) radiation were investigated in fully de-etiolated seedlings of a long-hypocotyl mutant (lh) of cucumber (Cucumis sativus L.), which is deficient in phytochrome-B, and its near isogenic wild type (WT). Continuous unilateral FR light provided against a background of white light induced negative curvatures (i.e. bending away from the FR light source) in hypocotyls of WT seedlings. This response was fluence-rate dependent and was absent in the lh mutant, even at very high fluence rates of FR. The phototropic effect of FR light on WT seedlings was triggered in the hypocotyls and occurred over a range of fluence rates in which FR was very effective in promoting hypocotyl elongation. FR light had no effect on elongation of lh-mutant hypocotyls. Seedlings grown in the field showed negative phototropic responses to the proximity of neighboring plants that absorbed blue (B) and red light and back-reflected FR radiation. The bending response was significantly larger in WT than in lh seedlings. Responses of WT and lh seedlings to lateral B light were very similar; however, elimination of the lateral B light gradients created by the proximity of plant neighbors abolished the negative curvature only in the case of lh seedlings. More than 40% of the total hypocotyl curvature induced in WT seedlings by the presence of neighboring plants was present after equilibrating the fluence rates of B light received by opposite sides of the hypocotyl. These results suggest that: (a) phytochrome functions as a phototropic sensor in de-etiolated plants, and (b) in patchy canopy environments, young seedlings actively project new leaves into light gaps via stem bending responses elicited by the B-absorbing photoreceptor(s) and phytochrome.     

Root growth dynamics of Aleppo pine (Pinus halepensis Mill.) seedlings in relation to shoot elongation, plant size and tissue nitrogen concentration

Large and high nitrogen (N) concentration seedlings frequently have higher survival and growth in Mediterranean forest plantations than seedlings with the opposite traits, which has been linked to the production of deeper and larger root systems in the former type of seedlings. This study assessed the influence of seedling size and N concentration on root growth dynamics and its relation to shoot elongation in Aleppo pine (Pinus halepensis Mill.) seedlings. We cultivated seedlings that differed in size and tissue N concentration that were subsequently transplanted into transparent methacrylate tubes in the field. The number of roots, root depth, and the root and shoot elongation rate (length increase per unit time) were periodically measured for 10 weeks. At the end of the study, we also measured the twig water potential (ψ) and the mass of plant organs. New root mass at the end of the study increased with seedling size, which was linked to the production of a greater number of new roots of lower specific length rather than to higher elongation rate of individual roots. Neither plant size nor N concentration affected root depth. New root mass per leaf mass unit, shoot elongation rate, and pre-dawn ψ were reduced with reduction in seedling size, while mid-day ψ and the root relative growth rate were not affected by seedling size. N concentration had an additive effect on plant size on root growth but its overall effect was less important than seedling size. Shoot and roots had an antagonistic elongation pattern through time in small seedlings, indicating that the growth of both organs depressed each other and that they competed for the same resources. Antagonism between shoot and root elongation decreased with plant size, disappearing in large and medium seedlings, and it was independent of seedling N concentration. We conclude that root and shoot growth but not rooting depth increased with plant size and tissue N concentration in Aleppo pine seedlings. Since production of new roots is critical for the establishment of planted seedlings, higher absolute root growth in large seedlings may increase their transplanting performance relative to small seedlings. The lack of antagonism between root and shoot growth in large seedlings suggests that these plants can provide resources to sustain simultaneous growth of both organs.     

Interaction of ethylene and a cytokinin in promoting hypocotyl elongation in a dwarf strain of watermelon

Experiments were conducted to study the interaction of ethylene and the cytokinin N⁶-benzyladenine (BA) in promoting hypocotyl elongation in a dwarf strain of watermelon (Citrullus lanatus [Thunb] Matsu. and Nakai). Optimum promotion of hypocotyl elongation is elicited by an apical treatment with 0.2 microgram BA. At dosages above 0.3 microgram per apex, BA-enhancement of elongation is reduced concomitant with stimulation of ethylene production and lateral expansion of hypocotyls. Application of the ethylene biogenesis inhibitor, aminoethoxyvinylglycine, at dosages from 0.3 to 10 micrograms per apex inhibited BA-induced ethylene production. In seedlings treated with 0.2 microgram BA, 10 micrograms aminoethoxyvinylglycine per apex reduced ethylene production to about one-third of control levels and reduced BA stimulation of hypocotyl elongation by 74%. Exposure of watermelon seedlings to 60 ± 10 nanoliters per liter of ethylene in a flowing system nearly eliminated aminoethoxyvinylglycine inhibition of BA-promoted growth. The results suggest that physiological levels of internal ethylene are required for cytokinin promotion of hypocotyl elongation in watermelon.     

Biomechanical study of the effect of a controlled bending on tomato stem elongation: global mechanical analysis

An experiment was designed to apply a controlled bending to a tomato stem and simultaneously to measure its effect on stem elongation. Stem elongation was measured over 2 d until steady and equal rates were obtained for the control and the treated plants. Thereafter, the basal part of the stem was submitted to a transient controlled bending at constant displacement rate using a motorized dynamometer. After load removal, stem elongation was again measured for 2 d. The tested plants were mature (height visible internodes) and only the basal part of the stem, which had already finished elongation, was loaded (hypocotyl and the first three internodes). A few minutes after the application of bending, elongation stopped completely for 60 min. Thereafter it took 120-1000 min to recover a rate of elongation similar to the control. The growth response was exclusively due to the bending of the basal part of the stem. It was shown that the side mechanical perturbations on the roots and on the stem tissues interacting directly with the clamp were not significantly involved on the elongation response. These results give evidence for mechanical perception and plant signalling from the basal stem to the upper elongating zone. However, none of the variables characterizing the global mechanical state of the bent part of the stem (i.e. the maximal force, bending moment, inclination, mean curvature of the stem, stored mechanical energy) could quantitatively explain the variability of the growth response. A more local mechanical analysis is therefore needed to elucidate how the mechanical stimulus is perceived.     

Repraesentins D,E and F,New Plant Growth Promoters from Lactarius repraesentaneus

Three new plant growth regulatory sesquiterpenes were isolated from the Lactarius repraesentaneus fungus. Their structures were elucidated to be lactarane sesquiterpenes, namely repraesentins D (1) and E (2), and a protoilludane-related sesquiterpene, namely repraesentin F (3). Repraesentin E (2) showed the strongest promotion activity, 164% at 3.6 μM, of the three compounds toward the radicle elongation of lettuce seedlings.