Role of the Cotyledons in the Phototropic Response of Lavatera cretica Seedlings

Young seedlings of Lavatera cretica L. exhibit positive phototropism. The hypocotyl perceives unilateral illumination with blue light and curves towards the light source by unequal growth. In addition, the cotyledonary laminas perceive the vectorial component of unilateral illumination with blue light and reorient normal to the beam by creating a turgor differential in their pulvini. Excision of one cotyledon resulted in negative organotropic curvature of the hypocotyl, away from the remaining cotyledon. Illumination of the cotyledonary lamina did not participate in the phototropic curvature of the hypocotyl, so long as the lamina was free to reorient to face the beam. When the lamina was continuously exposed to vectorial photoexcitation, elongation of the hypocotyl on the side carrying the cotyledon could be enhanced, or inhibited, depending on the direction of the beam striking its lamina.     

Mechanism of rapid suppression of cell expansion in cucumber hypocotyls after blue-light irradiation

Rapid suppression of hypocotyl elongation by blue light in cucumber (Cucumis sativus L.) was studied to examine possible hydraulic and wall changes responsible for diminished growth. Cell-sap osmotic pressure, measured by vaporpressure osmometry, was not decreased by blue light; turgor pressure, measured by the pressureprobe technique, remained constant during the growth inhibition; and stem hydraulic conductance, measured by dynamic and static methods, was likewise unaffected by blue light. Wall yielding properties were assessed by the pressure-block technique for in-vivo stress relaxation. Blue light reduced the initial rate of relaxation by 77%, but had little effect on the final amount of relaxation. The results demonstrate that blue irradiation acts to decrease the wall yielding coefficient, but not the yield threshold. Stress-strain (Instron) analysis showed that irradiation of the seedlings had little effect on the mechanical extensibilities of the isolated wall. The results indicate that blue light can reduce cell-wall loosening without affecting bulk viscoelastic properties, and indicate a chemorheological mechanism of cell-wall expansion.Abbreviations and symbols BL blue light - wall yield coefficient - Y wall yield threshold - P turgor pressure - L hydraulic conductance - g radial water-potential gradient supporting cell expansion - osmotic pressure - P_i initial chamber pressure needed to stop growth - P_f final chamber pressure needed to stop growth     

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.     

Cell Wall Free Space of Cucumis Hypocotyls Contains NAD and a Blue Light-Regulated Peroxidase Activity

Solutions were obtained from the cell wall free space of red light-grown cucumber (Cucumis sativus L.) hypocotyl sections by a low-speed centrifugation technique. The centrifugate contained NAD and peroxidase but no detectable cytoplasmic contamination, as indicated by the absence of the activity of glucose-6-phosphate dehydrogenase from the cell wall solution. Peroxidase activity centrifuged from the cell wall of red light-grown cucumber hypocotyl section could be resolved into at least three cathodic isoforms and two anodic isoforms by isoelectric focusing. Treatment of red light-grown cucumber seedlings with a 10-minute pulse of high-intensity blue light increased the level of cell wall peroxidase by about 60% and caused a qualitative change in the anodic isoforms of this enzyme. The increase in peroxidase activity was detectable within 25 minutes after the start of the blue light pulse, was maximal at 35 minutes, and declined to control levels by 45 minutes of irradiation. The inhibitory effect of blue light on hypocotyl elongation was more rapid than the effect of blue light on total wall peroxidase activity, leading to the conclusion that growth and peroxidase activity are not causally related.     

Signal Integration by ABA in the Blue Light-Induced Acidification of Leaf Pavement Cells in Pea (Pisum sativum L. var. Argenteum)

Leaf pavement cell expansion in light depends on apoplastic acidification by a plasma membrane proton-pumping ATPase, modifying cell wall extensibility and providing the driving force for uptake of osmotically active solutes generating turgor. This paper shows that the plant hormone ABA inhibits light-induced leaf disk growth as well as the blue light-induced pavement cell growth in pea (Pisum sativum L.). In the phytochrome chromophore-deficient mutant pcd2, the effect of ABA on the blue light-induced apoplastic acidification response, which exhibits a high fluence phase via phytochrome and a low fluence phase via an unknown blue light receptor, is still present, indicating an interaction of ABA with the blue light receptor pathway. Furthermore, it is shown that ABA inhibits the blue light-induced apoplastic acidification reversibly. These results indicate that the effect of ABA on apoplastic acidification can provide a mechanism for short term, reversible adjustment of leaf growth rate to environmental change.Key Words: ABA, apoplastic acidification, blue light, epidermal pavement cell growth, leaf growth, pea (Pisum sativum L.), signal integration     

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     

Abscisic Acid accumulates at positive turgor potential in excised soybean seedling growing zones

Abscisic acid (ABA) accumulated in soybean (Glycine max [L.] Merr. cv Williams) hypocotyl elongating regions when seedlings were transferred to low water potential vermiculite (Ψ = −0.3 megapascals) even though positive turgor is retained in this tissue. Accumulation of ABA in growing zones could occur from de novo biosynthesis within this tissue or transport from adjacent nongrowing zones. Both growing and nongrowing hypocotyl and root tissues accumulated significant levels of ABA when excised and dehydrated to reduce turgor. Surprisingly, excised growing zones (which experienced no water loss) also accumulated ABA when incubated in darkness for 4 hours at 100% relative humidity and 29°C. Induction of ABA accumulation in the excised elongating region of the hypocotyl was not caused by disruption of root pressure or wounding. While excision of hypocotyl elongating regions induced ABA accumulation, no change in either extensin or p33 mRNA levels was observed. Accumulation of extensin or p33 mRNA required more severe wounding. This suggests that ABA is not involved in the response of these genes in wounded tissue and that wound signals are not causing ABA accumulation in excised tissue. Accumulation of ABA in excised elongating regions was correlated with growth inhibition and a decline in turgor to the yield threshold (Ψ;_p = 0.37 megapascals; R Matyssek, S Maruyama, JS Boyer [1988] Plant Physiol 86: 1163-1167). Inhibiting hypocotyl growth by transferring seedlings to lower temperatures or light did not cause ABA accumulation. We conclude that induction of ABA accumulation in growing zones is more sensitive to changes in turgor than the induction which occurs in mature tissues.     

Endogenous Abscisic Acid Promotes Hypocotyl Growth and Affects Endoreduplication during Dark-Induced Growth in Tomato (Solanum lycopersicum L.)

Dark-induced growth (skotomorphogenesis) is primarily characterized by rapid elongation of the hypocotyl. We have studied the role of abscisic acid (ABA) during the development of young tomato (Solanum lycopersicum L.) seedlings. We observed that ABA deficiency caused a reduction in hypocotyl growth at the level of cell elongation and that the growth in ABA-deficient plants could be improved by treatment with exogenous ABA, through which the plants show a concentration dependent response. In addition, ABA accumulated in dark-grown tomato seedlings that grew rapidly, whereas seedlings grown under blue light exhibited low growth rates and accumulated less ABA. We demonstrated that ABA promotes DNA endoreduplication by enhancing the expression of the genes encoding inhibitors of cyclin-dependent kinases SlKRP1 and SlKRP3 and by reducing cytokinin levels. These data were supported by the expression analysis of the genes which encode enzymes involved in ABA and CK metabolism. Our results show that ABA is essential for the process of hypocotyl elongation and that appropriate control of the endogenous level of ABA is required in order to drive the growth of etiolated seedlings.     

Similarity between Cytokinin and Blue Light Inhibition of Cucumber Hypocotyl Elongation

The cytokinin benzyladenine inhibited endogenous hypocotyl elongation in intact etiolated seedlings of cucumber (Cucumis sativus L.). In hypocotyl segments, the inhibitory effect of benzyladenine on growth was clearly detectable in the presence of indoleacetic acid. Fusicoccin-induced elongation was unaffected by the presence of cytokinin. The effect of cytokinin on elongation of the segments was determined by measuring changes in fresh weight, a linear function of extension growth. The effect of benzyladenine on hypocotyl growth was at least as large in segments prepared from red-light-grown seedlings as in those from seedlings grown in total darkness. A comparison was made between the inhibitory effects of cytokinin and blue light. The use of the calcium chelator ethyleneglycol-bis(β-aminoethyl ether)-N, N′-tetraacetic acid indicated that calcium ions are required for manifestation of benzyladenine-induced inhibition.     

Turgor and Longitudinal Tissue 12Helianthus annuus

The quantitative relationship between turgor and the pressureexerted by the inner tissues (cortex, vascular tissue, and pith)on the peripheral cell walls (longitudinal tissue pressure)was investigated in hypocotyls of sunflower seedlings (Helianthusannuus L.) In etiolated hypocotyls cell turgor pressures, asmeasured with the pressure probe, were in the range 0·38to 0·55 MPa with an average of 0·48 MPa. In irradiatedhypocotyls turgor pressures varied from 0·40 to 0·57MPa with a, mean at 0·49 MPa. The pressure exerted bythe inner tissues on the outer walls was estimated by incubatingpeeled sections in a series of osmotic test solutions (polyethyleneglycol 8000). The length change was measured with a transducer.In both etiolated and irradiated hypocotyls an external osmoticpressure of 0·5 MPa was required to inhibit elongationof the inner tissues, i.e. the average cell turgor and the longitudinaltissue pressure are very similar quantities. The results indicatethat the turgor of the inner tissues is displaced to and borneby the thick, growth-limiting peripheral cell walls of the hypocotyl. Key words: Helianthus annuus, hypocotyl growth, tissue pressure, turgor pressure, wall stress     

Turgor Pressure and Phototropism in Sinapis alba L. Seedlings

Rich, T. C. G. and Tomos, A. D. 1988. Turgor pressure and phototropismin Sinapis alba L. seedlings.—J. exp. Bot 39: 291-299. Phototropic responses were studied in light-grown mustard hypocotyls.Phototropism was induced by adding 0.27 µmol m^–2s^–1 unilateral blue light to a background of low pressuresodium (SOX) lamp light. Curvatures of some 6° from thevertical were reached by 60 min, the curvature rate between20 min and 60 min being 0.14° min^–1. From the axialgrowth rate and tissue geometry the local growth rates of illuminatedand shaded sides of the hypocotyl were calculated to be 1.5and 4.5 µmin^–1 respectively. Turgor pressures ofexpanding cells in control plants and in the shaded and illuminatedsides of the blue light illuminated hypocotyls were measuredto be 0.40-0.55 MPa with a pressure probe. No changes in turgorpressure were observed on initiation of curvature. The decayof pressure in the cells of non-transpiring plants followingexcision indicated that the yield stress threshold of the tissuemay be as low as 0.1 MPa. These results indicate that the phototropicgrowth response in this tissue is not mediated by changes inturgor pressure. Key words: Sinapis alba L., phototropism, turgor pressure     

The effect of lateral illumination on growth oscillations of pine seedling hypocotyls (Pinus silvestris L.)

Phototropic sensitivity of forest wood seedlings to lateral illumination was proposed as an early assay for distinquishing various ecotypes of woody plants of the same species. Statistical analysis showed that results were significantly influenced by heterogenity of experimental material caused by an interference of phototropic movements and natural oscillations of hypocotyl. Both movements of pine seedlings (Pinus silvestris L.) were registered by phase photography and their mechanism was analyzed. The apical part of growing hypocotyl illuminated from above oscillates in a space spiral with frequency 3.3 h at mean growth rate 0.66 mm h^?1. The mean size of spiral amplitudes is 2.9 mm. The oscillation rhythm is disturbed after the lateral illumination and a phase shift was observed as a result. A new oscillation rhythm with frequency 3.9 h and mean growth rate 0.69 mm h^?1 was stabilized after a period of time equal to one nutation turn. Oscillation amplitudes were increased to 4.3 mm. In addition to the radiation intensity the appearance of the phototropic response to light pulse is first of all effected by the actual position of the apical part of hypocotyl in the course of endogenous circumnutations. A uniform plant material for the early assay may be obtained by selection of seedlings which are at the beginning of lateral illumination in the same phase of nutation rhythm. Under such conditions the deviations of longitudinal axis of oscillating spirals characterize the actual phototropic curvature.     

Effect of light on ethylene production and hypocotyl growth of soybean seedlings

The apical 1-cm hypocotyl of dark-grown `Clark' soybean (Glycine max [L.] Merr.) seedlings produced ethylene at rates of 7 to 11 nanoliters per hour per gram when attached to the cotyledons. Such physiologically active rates occurred prior to the deceleration of hypocotyl elongation caused by the temperature of 25 C.

Daily exposure of the etiolated seedlings to red light promoted hypocotyl elongation and prevented its lateral swelling. Red light treatment also caused a 45% decrease in ethylene production. Far red irradiation following the red treatment reversed the red effects, suggesting that the ethylene intervenes as a regulator in the phytochrome control of `Clark' soybean hypocotyl growth at 25 C.

     

The effect of (2-chloroethyl)trimethylammonium chloride (CCC) and gibberellic acid on the growth and development of wheat cultivated under red or blue light

The effect of CCC and GA₃ on the growth and development of spring wheat (Triticum aestivum L.) cultivated under predominantly red (500–700 nm) or blue (400–500 nm) light was investigated. Red light enhanced the development of wheat during the exponential phase of growth. This effect presumably implicated the promotion of gibberellin synthesis under red light. The strong inhibitory action of CCC under red light (the inhibition was lower under blue light) might be interpreted in a similar way. The growth became more intensive under blue light after caring and was accompanied by increased susceptibility to giberellic acid treatment.     

Gibberellic Acid stimulation of cucumber hypocotyl elongation : effects on growth, turgor, osmotic pressure, and cell wall properties

Recently developed techniques have been used to reinvestigate the mechanism by which gibberellic acid (GA₃) stimulates elongation of light-grown cucumber (Cucumis sativus L.) seedlings. Osmotic pressure and turgor pressure were slightly reduced in GA₃-treated seedlings, which elongated 3.5 times faster than control seedlings. This indicated that GA₃ enhancement of growth was not controlled by changes in the osmotic properties of the tissues. Stress/strain (Instron) analysis revealed that plastic extension of the cell walls of GA₃-treated seedlings increased by up to 35% above the control values. Stress-relaxation measurements on frozen-thawed tissue showed that T₀ the minimum relaxation time, was reduced following application of GA₃. In vivo wall relaxation (measured by the pressure block technique) showed that the wall yield coefficient was increased, and the yield threshold was slightly reduced. Thus GA₃ affected both the mechanical (viscoelastic) and biochemical (chemorheological) properties of the cell walls of light-grown cucumber. The previous hypothesis, that GA₃ stimulates cucumber hypocotyl growth by increasing osmotic pressure and cell turgor, is contradicted by our results.     

Gradients of turgor, osmotic pressure, and water potential in the cortex of the hypocotyl of growing ricinus seedlings : effects of the supply of water from the xylem and of solutes from the Phloem

To evaluate the possible role of solute transport during extension growth, water and solute relations of cortex cells of the growing hypocotyl of 5-day-old castor bean seedlings (Ricinus communis L.) were determined using the cell pressure probe. Because the osmotic pressure of individual cells (πⁱ) was also determined, the water potential (ψ) could be evaluated as well at the cell level. In the rapidly growing part of the hypocotyl of well-watered plants, turgor increased from 0.37 megapascal in the outer to 1.04 megapascal in the inner cortex. Thus, there were steep gradients of turgor of up to 0.7 megapascal (7 bar) over a distance of only 470 micrometer. In the more basal and rather mature region, gradients were less pronounced. Because cell turgor ≈ πⁱ and ψ ≈ 0 across the cortex, there were also no gradients of ψ across the tissue. Gradients of cell turgor and πⁱ increased when the endosperm was removed from the cotyledons, allowing for a better water supply. They were reduced by increasing the osmotic pressure of the root medium or by cutting off the cotyledons or the entire hook. If the root was excised to interrupt the main source for water, effects became more pronounced. Gradients completely disappeared and turgor fell to 0.3 megapascal in all layers within 1.5 hours. When excised hypocotyls were infiltrated with 0.5 millimolar CaCl₂ solution under pressure via the cut surface, gradients in turgor could be restored or even increased. When turgor was measured in individual cortical cells while pressurizing the xylem, rapid responses were recorded and changes of turgor exceeded that of applied pressure. Gradients could also be reestablished in excised hypocotyls by abrading the cuticle, allowing for a water supply from the wet environment. The steep gradients of turgor and osmotic pressure suggest a considerable supply of osmotic solutes from the phloem to the growing tissue. On the basis of a new theoretical approach, the data are discussed in terms of a coupling between water and solute flows and of a compartmentation of water and solutes, both of which affect water status and extension growth.     

Streptomycin induced changes in growth and metabolism of etiolated seedlings

Effects of various concentrations of streptomycin sulphate either alone or in combination with different cations and hormones on mungbean (Phaseolus aureus L.) seedling growth were studied. The relative inhibition of root growth was stronger than that of hypocotyl growth. Root growth inhibition was completely overcome by calcium, while other cations were ineffective. Inhibition of hypocotyl elongation could not be prevented by cations. IAA and GA₃ were capable of relieving streptomycin inhibition but kinetin was ineffective. In the coleoptiles of streptomycin-treated rice (Oryza sativa L.) seedlings, there were accumulation of nucleic acids and decline in protein content resulting in increased RNA/protein and DNA/protein ratios. High nucleic acid content induced by streptomycin could be correlated with reduced activity of the nucleases.     

Boron-regulated hypocotyl elongation is affected in Arabidopsis mutants with defects in light signalling pathways

We studied the effect of elevated boron (B) concentrations on the growth and development of Arabidopsis thaliana in vitro with respect to different light conditions. Two basic responses were observed. At high concentrations (above 5 mM) a clear toxicity effect of B on plant growth was apparent. Seedlings were short, stunted and pale. However at concentrations between 1 and 3 mM H₃BO₃, hypocotyl elongation was stimulated in all Arabidopsis ecotypes tested relative to plants grown at 0.1 mM H₃BO₃. The stimulation of hypocotyl elongation by elevated B was proportionally greater with increasing irradiance. We also showed that blue light (BL) and red light (RL) did not alter the sensitivity of Arabidopsis hypocotyls to boron, but, dependent on genotype, BL and RL increased or reduced capacity of boron-induced hypocotyl elongation. Analysis of photomorphogenic mutants indicated the existence of an interaction between boron and light signalling pathways during plant growth and development. This interaction was supported by the observation that the expression of the BOR1 gene in Arabidopsis hypocotyls was stimulated by BL and RL. Our results suggest that in etiolated or light-grown seedlings the stimulation of hypocotyl growth by boron can be mediated by cryptochromes and phytochromes.     

Inhibition of stem elongation in cucumis seedlings by blue light requires calcium

The effects of blue light and calcium on elongation of hypocotyl segments of Cucumber (Cucumis sativa L. cv Burpee's Pickler) were studied. Cucumber seedlings grown in dim red light showed a rapid decline in the rate of hypocotyl elongation when irradiated with high intensity (100 micromoles per square meter per second) blue light. In intact, 4-day-old seedlings the inhibition began within 2 minutes after the onset of blue-light irradiation and reached a maximum of approximately 55% within 4 minutes. Hypocotyl segments cut from 4-day-old seedlings also showed an inhibition of elongation in response to blue light when segments were floated on aqueous buffer and exposed to blue light for 3 hours. In the presence of 2 micromolar indole-3-acetic acid, blue light caused a 50% inhibition of elongation. Buffering free calcium in the incubation medium with 0.1 millimolar ethylene glycol bis(-aminoethyl ether)- N,N,N′,N′-tetraacetic acid eliminated the blue-light inhibition of segment elongation. Several experiments confirmed a specific requirement for calcium for the blue-light-induced inhibition of segment elongation. Treating segments with 0.2 micromolar fusicoccin abolished the inhibition of elongation by blue light as did buffering the medium at pH 4. Adding 1 millimolar ascorbate to incubation medium also eliminated the inhibition of segment elongation caused by blue light. Several compounds implicated in cell-wall redox reactions alter the magnitude of the blue-light-induced inhibition. The activity of peroxidase isolated from the cell-wall free space of cucumber hypocotyls was inhibited by ascorbate and low pH. The results are consistent with the hypothesis that blue light inhibits elongation by inducing an increase in cell-wall peroxidase activity and implicate calcium ions in the response to blue light.     

Endogenous Phytohormones and Regulation of Morphogenesis of Arabidopsis thalianaby Blue Light

We studied the effects of blue light (BL) on the levels of endogenous phytohormones (IAA, ABA, gibberellins, and cytokinins) and morphogenesis of the 7-day-old Arabidopsis thaliana(L.) Heynh seedlings of wild type (Ler) and its hy4mutant with a disturbed synthesis of cryptochrome 1 (CRY1), which is a receptor for BL. In darkness, the mutant contained considerably less free IAA and zeatin, but much more ABA as compared to the wild-type seedlings. BL retarded the hypocotyl growth in the wild-type seedlings but stimulated it in the mutant. Elongation of mutant hypocotyls was accompanied by accumulation of free IAA and a decrease in the content of free ABA; the level of cytokinins did not change. We believe that the response of the hy4hypocotyls to BL is mediated by a BL receptor distinct from cryptochrome 1. The conclusion is that light and hormonal signals interact in the control of the hypocotyl growth in A. thalianaseedlings.