Nyctinastic Closure Research Articles

Calcium changes in Robinia pseudoacacia pulvinar motor cells during nyctinastic closure mediated by phytochromes.

Potassium pyroantimonate precipitation, transmission electron microscopy, and X-ray microanalysis were used to investigate the subcellular localization of loosely bound calcium in Robinia pseudoacacia pulvinar motor cells during phytochrome-mediated nyctinastic closure. Calcium localization was carried out in pulvini collected in white light 2h after the beginning of the photoperiod, immediately after a red light or a far-red light pulse applied 2h after the beginning of the photoperiod and after 15 or 25min of darkness respectively. Calcium antimonate precipitates were found in all the pulvinar tissues from the epidermis to the vascular bundle, independent of the light treatment. At subcellular level, precipitates were found mainly in the intercellular spaces, the inner surface of the plasma membrane, cytoplasm, colloidal vacuoles, and nuclei. Red light enhanced the nyctinastic closure of leaflets and caused an asymmetric distribution of cytosolic calcium precipitates between the extensor and flexor motor cells. Both the number and area of the cytosolic calcium precipitates drastically increased in the extensor cells compared to the flexor motor cells. Red light had a rapid and transient effect on the distribution of cytosolic calcium precipitates, which occurred during or at the end of the irradiation, before leaflet closure. By contrast, the distribution of cytosolic loosely bound calcium was similar between the extensor and flexor motor cells after irradiation with far-red light. Our results demonstrate that red light causes specific calcium mobilization in pulvinar motor cells and suggest the involvement of cytoplasmic Ca2+ as a second messenger for phytochrome during nyctinastic closure.

Nitric-oxide inhibits nyctinastic closure through cGMP in Albizia lophantha leaflets

Nitric oxide (NO) is a highly reactive radical that acts as a direct or indirect cellular signalling molecule in plant growth, development and environmental responses. Here we studied the contribution of NO to the control of leaflet movements during nyctinastic closure. For this purpose, we tested the effect of NO donors and an NO scavenger, all supplied in light, on Albizia lophantha leaflet closure after transferral to darkness. Exogenous NO, applied as four donors [sodium nitroprusside (SNP), diethylammonium (Z)-1-(N,N-diethylamino) diazen-1-ium-1,2-diolate (DEA-NONOate), S-nitroso-N-acetylpenicillamine (SNAP) and S-nitrosoglutathione (GS-NO)], inhibited nyctinastic leaflet closure while the application of an NO scavenger [2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO)] plus SNP cancelled the effect of the latter. The inclusion of Nω-nitro-l-arginine methyl ester (l-NAME) or sodium tungstate in the incubation media enhanced nyctinastic closure and also resulted in a decrease in the nitrate plus nitrite released by leaflets into the incubation solution. These results support the notion that NO is involved in regulating the nyctinastic closure of A. lophantha leaflets. Cellular perception of NO did not appear to be mediated by calcium. Pharmacological application of inhibitors of soluble guanylate cyclase (sGC) [1H-[1,2,4]-oxadiazole-[4,3-a]-quinoxalin-1-one (ODQ) and 6-anilino-5,8-quinolinequinone (Ly83583)], phosphodiesterase type 5 (PDE5) (Sildenafil) and the cyclic guanosine monophosphate (cGMP) analogue 8-bromoguanosine-3',5'-cyclomonophosphate sodium salt (8-Br-cGMP) indicated that cGMP was downstream of the NO signalling cascade during nyctinastic closure.

Microautoradiographic localisation of [3H]sucrose and [3H]mannitol in Robinia pseudoacacia pulvinar tissues during phytochrome-mediated nyctinastic closure

We have analysed the incorporation of [(3)H]sucrose and [(3)H]mannitol in pulvinar motor cells of Robinia pseudoacacia L. during phytochrome-mediated nyctinastic closure. Pairs of leaflets, excised 2 h after the beginning of the photoperiod, were fed with 50 mM [(3)H]sucrose or [(3)H]mannitol, irradiated with red (15 min) or far-red (5 min) light and placed in the dark for 2-3 h. Label uptake was measured in whole pulvini by liquid scintillation counting. The distribution of labelling in pulvinar sections was assessed by both light and electron microautoradiography. [(3)H]Sucrose uptake was twice that of [(3)H]mannitol incorporation in both red- and far-red-irradiated pulvini. In the autoradiographs, [(3)H]sucrose and [(3)H]mannitol labelling was localised in the area from the vascular bundle to the epidermis, mainly in vacuoles, cytoplasm, and cell walls. Extensor and flexor protoplasts displayed a different distribution of [(3)H]sucrose after red and far-red irradiation. Far-red light drastically reduced the [(3)H]sucrose incorporation in extensor protoplasts and caused a slight increase in internal flexor protoplasts. After red light treatment, no differences in [(3)H]sucrose labelling were found between extensor and flexor protoplasts. Our results indicate a phytochrome control of sucrose distribution in cortical motor cells and seem to rule out the possibility of sucrose acting as an osmoticum.

Control of circadian rhythm-regulated nyctinastic movement in water lily (Nymphea stellataWilld.) flowers

SummaryFlowers of Nymphea stellataWilld., like other water lilies, are subject to circadian clock-controlled petal closure in the afternoon. To investigate the effects of plant growth regulators, and various other chemicals, in arresting nyctinastic movement (petal closure), cut flowers of N. stellata were exposed to different concentrations of gibberellic acid (GA), benzyl amino purine (BAP), naphthalene acetic acid (NAA), abscisic acid (ABA), ethanol, AgNO3, NaCl or RinsoTM (laundry detergent powder). The most effective treatment to control the biological clock of petal movement was keeping flowers continuously in 25 mg l–1 GA.This treatment kept flowers open, to an acceptable cut flower standard, for over 28 h. Continuous exposure to 5 mg l–1 BAP also kept flowers open to the same acceptable standard for over 15 h. Considering low-cost applications, immersing flowers in 25 g l–1 NaCl for 1 h kept them open for almost 17 h, to an acceptable cut flower standard. Immersing in 2 g l–1 RinsoTM (Unilever, Sri Lanka) for 1 h also kept flowers open for 14 h in an acceptable condition. By inhibiting nyctinastic closure using any of the above chemicals, N. stellata could now be considered and exploited as a cut flower.

Phytochrome‐mediated closure of Albizzia lophantha leaflets as affected by sorbitol, mannitol and fusicoccin

The effect of sorbitol and mannitol and their interaction with the effect of fusiccocin has been investigated in phytochrome‐mediated nyctinastic closure of Albizzia lophantha leaflets. Treatments were commenced 2 h into the photoperiod. Pairs of leaflets were excised, floated on test solutions, and transferred to darkness immediately after red or far red irradiation. Leaflet angles were measured in darkness for 3 h. Fusicoccin (1 to 100 μM) inhibited nyctinastic closure and enhanced reopening in the presence of Pft (far‐red‐absorbing form of phytochrome). Fusicoccin effects on far red treated leaflets were not constant and whichever fusicoccin concentration was applied, the effects were small. These results confirm the role of the plasma membrane proton pump in extensor cells during opening. Sorbitol and mannitol (25–100 mM) increased nyctinastic closure for about 2 h and then enhanced reopening in the presence of Pfr. When sugar alcohols were applied together with fusicoccin, sorbitol and mannitol enhanced the fusicoccin inhibition of closure and a marked synergistic effect was observed. These results indicate that both fusiccocin and the reduction of external osmotic potential caused by sorbitol and mannitol supply may stimulate the activity of the extensor H+ pump promoting the reopening.

Effect of EGTA on Nyctinastic Closure Mediated by Phytochrome in <italic>Albizzia lophantha</italic>

The role of extracellular calcium on nyctinastic closure of Albizzia lophantha leaflets has been studied by testing the effect of EGTA and its reversibility by calcium

Role of Calcium in Phytochrome-Controlled Nyctinastic Movements of Albizzia lophantha Leaflets

The involvement of Ca(2+) on phytochrome-controlled nyctinastic closure in Albizzia lophantha has been studied by testing the effect of the calcium ionophore 6S-[6alpha(2S(*),3S(*)),8beta(R(*)),9beta,11alpha]-5- methyl-amino)-2-[[3,9,11-trimethyl-8-[-1-methyl-2-oxo-2-(1H-pyrrol-2-yl) ethyl]-1,7-dioxaspiro[5.5]-undec-2yl] methyl]-4-benzoxazolecarboxylic acid (A23187) and the intracellular calcium antagonist 8-(diethylamino)octyl 3,4,5-trimethoxybenzoate hydrochloride (TMB-8). An external supply of Ca(2+) or calcium ionophore A23187 to the Albizzia leaflets emulates the effect of red light irradiation and counteracts the inhibitory effect of far red light. The intracellular calcium antagonist TMB-8 supplied to Albizzia leaflets inhibits the effect of red light, but had no effect on far red irradiated plants. This suggests a dependence between phytochrome action and intracellular free Ca(2+). We suggest that calcium acts as a phytochrome messenger on control of ion fluxes that drive turgor changes in pulvinular motor cells.

Effect of Far-Red and Green Irradiation on the Nyctinastic Closure of Albizzia Leaflets

The nyctinastic closing of Albizzia julibrissin pinnules is delayed by exposure to far-red radiation at 710 and 730 nanometers, with the former more effective than the latter. Far-red radiation at 750 and 770 nanometers has no effect on the process. Red light at 660 nanometers, which by itself has no effect, delayed closure when given before or simultaneously with far-red radiation at 750 or 770 nanometers. Low doses of green light, on the other hand, prevented all far-red radiations from delaying closure when given together with one of them. Effectiveness peaks at 550 nanometers. Green light by itself has no effect on the closing process.From these and previous results, it is concluded that phytochrome is one of two photoreceptors in the process, that the other photoreceptor is an unknown pigment, and that the unknown photoreceptor requires some prior effect of the far-red-absorbing form of phytochrome before its action. Predictions are made of some of the properties of the unidentified pigment.

Role of Boron in the Far-Red Delay of Nyctinastic Closure of Albizzia Pinnules

Boron has been found to be necessary for the delaying action of 710 nanometer irradiation on the nyctinastic closing of Albizzia julibrissin pinnules. It is effective only over a narrow micromolar range. In contrast, the delaying action of 470 nanometer irradiation on closure is inhibited by boron. The effect of boron on leaflet closing occurs rapidly. Boron is suggested to be involved in a phytochrome action that precedes and is required for a delaying effect on pinnule closing subsequently induced by an unknown far-red absorbing pigment.

Effect of the lithium chloride on the leaf movements of Cassia fasciculata

LiCl inhibits the nyctinastic closure of folioles of excised leaves and enhances their opening, if there are 3 h before the light is switched off or on; the minimal concentration for significant effects on closure is 3·10(-4) M, and on opening 3·10(-3). The use of chlorides of other cations and other Li salts showed specificity of the lithium for the closure movement, the effect being reversed by KCl and NaCl. For the opening movement the Li effect is less specific. These results are compared to those obtained in other phenomena.

Potassium flux and leaf movement in Samanea saman. II. Phytochrome controlled movement.

Phytochrome, a membrane-localized biliprotein whose conformation is shifted reversibly by brief red or far-red light treatments, interacts with the rhythmic oscillator to regulate leaflet movement and potassium flux in pulvinal motor cells of Samanea. Darkened pinnae exposed briefly to red light (high P(fr) level) have less potassium in motor cells in the extensor region, more potassium in motor cells in the flexor region, and smaller angles than those exposed to far-red light (low P(fr) level). Increase in temperature from 24 degrees to 37 degrees increases the differential effect of the light treatments during opening (the energetic phase) but not during closure, implying that phytochrome controls an energetic process. It seems likely that phytochrome interacts with rhythmically controlled potassium pumps in flexor and extensor cells. During nyctinastic closure of white-illuminated pinnae, exposure to far-red light before darkening results in larger angles than does exposure to red. As in rhythmic opening, the angles of all pinnae and the differential effect of the light treatments increases with increasing temperature.

Rhythmic Potassium Flux in Albizzia

Rhythmic leaflet movement in Albizzia is controlled by rhythmic K(+) flux in pulvinal motor cells. The angle assumed by darkened leaflets during the open phase of the rhythm can be altered by various compounds and changes in temperature; such treatments are ineffective during the closed phase. In all cases, effects on leaflet angle are correlated with and probably a consequence of K(+) flux in pulvinal motor cells. Incubation at low temperature (6C) or on sodium azide (1.0 mm) reduces K(+) in the ventral motor region and increases K(+) in the dorsal motor region, thereby decreasing leaflet angle. Incubation on cycloheximide (0.1 mm) or sodium acetate (0.05 m) inhibits protein synthesis; if the incubation period immediately precedes the opening phase, these compounds prevent both K(+) flux into the ventral motor cells and leaflet opening. Magnesium nitrate (0.05 m), supplied after leaflets have started to open, promotes K(+) secretion from the dorsal motor cells and increases the angle of opening.The data support the following hypothesis. Active K(+) transport into the ventral motor cells and out of the dorsal motor cells leads to opening; K(+) leakage in the opposite directions causes closure; and the interaction of these processes results in a rhythmic oscillation. Proteins in the ventral cell membranes that are required for active transport or membrane integrity turn over rhythmically and are resynthesized before opening. The availability of divalent cations determines the phase relationships between the K(+) rhythms in the dorsal and ventral motor cells.White light phases the rhythm. The "light on" signal turns on a K(+) secreting pump in the dorsal motor cells, while the "light off" signal initiates a period of protein synthesis which in turn leads to active K(+) transport into the ventral motor cells. Aminophylline (0.1-6.0 mm) inhibits white light-promoted opening and nyctinastic closure.

Phytochrome-controlled Nyctinasty in Albizzia julibrissin

Indole-3-acetic acid, alpha-naphthylacetic acid, and 2,4-dichlorophenoxyacetic acid (0.001 to 1.0 mm) inhibit the nyctinastic closure of excised Albizzia leaflet pairs; antiauxins and auxin analogs are ineffective, and the auxin effects seem not to be mediated by ethylene. Indoleacetic acid (0.001 to 0.1 mm) also promotes rhythmic opening in the dark, but is ineffective during that phase of rhythmic closure ("leaky phase") which is insensitive to azide. At these concentrations, all of the indoleacetic acid effects are reversible upon transfer of the tissue to water and are linked to alteration of potassium flux in pulvinule motor cells.A supraoptimal concentration of indoleacetic acid (1 mm) inhibits rhythmic opening as well as nyctinastic closure, although it has little or no effect on potassium flux in motor cells. These inhibitions cannot be completely reversed by transferring the leaflets to water.Although indoleacetic acid (0.01 to 1.0 mm) inhibits leaflet opening and potassium flux in dorsal and ventral motor cells when leaflets are transferred from darkness to light, it has no effect during other portions of the light period, implying that changes in endogenous auxin do not control leaflet angle in the light. Neither does auxin seem to be involved in the phytochrome-regulated process, since it does not alter phytochrome control of leaflet movement or potassium flux. However, endogenous auxin probably plays an important role in controlling potassium flux into ventral motor cells during the opening phase of rhythmic leaflet movement in the dark.

Effects of Ethylenediaminetetraacetic Acid, Auxin and Gibberellic Acid on Phytochrome Controlled Nyctinasty in Albizzia julibrissin

AbstractNyctinastic closure of Albizzia julibrissin pinnules is inhibited by 5 × 10−2M ethylenediaminetetraacetic acid. At least two hours of incubation are required for maximum inhibition and destruction of the phytochrome effect.Concentrations of 10−3 to 10−5M naphthaleneacetic acid reduced the nyctinastic closure of pinnules but not the phytochrome response. Similar results were obtained with indoleacetic acid and gibberellic acid. No appreciable differences in pinnule movements could be attributed to pH. Chelation or the inhibition of ion transport resulting in, or caused by, changes in membrane permeability are suggested as possible mechanisms involved in these effects.

Phytochrome-controlled Nyctinasty in Albizzia julibrissin

Prolonged irradiation during appropriate parts of the diurnal cycle promotes the opening of Albizzia julibrissin leaflets. Leaflets also open without illumination, but such opening starts later and is slower and less complete. Opening in the dark is accompanied by lower potassium efflux from dorsal pulvinule motor cells but equal or greater potassium movement into ventral motor cells than occurs during opening in the light. Far red-absorbing phytochrome inhibits opening in the dark, indicating that its action is similar during endogenously controlled opening and nyctinastic closure; i.e., a high far redabsorbing phytochrome level is associated with low potassium content in ventral motor cells, high potassium content in dorsal motor cells, and a small angle between leaflets.When open leaflets are darkened, there is an immediate and large potassium flux into dorsal motor cells. This is initially independent of red and far red preirradiation, but prior red light appears to promote continued potassium movement into dorsal cells during the latter part of a 90-minute dark period. The situation in ventral motor cells is different; here the effect of prior red or far red irradiation on potassium efflux is evident after 10 minutes of darkness. Phytochrome controls the direction of potassium movement in ventral motor cells during the early part of the dark period (to 25 minutes); potassium moves out of ventral motor cells if leaflets are preirradiated with red light and into these same cells if leaflets are preirradiated with far red light. Kinetic data are consistent with the suggestion that potassium leaving ventral cells enters dorsal cells. However, there must be an additional source of potassium entering dorsal cells since this potassium movement precedes potassium efflux from ventral cells.Pulvinules excised from the lamina or rachilla open and close in response to light and darkness and also move during extended periods of constant intensity light or uninterrupted darkness. This shows that the photoreceptor controlling opening and the oscillator controlling endogenously rhythmic leaflet movement are localized in the pulvinule. In addition, all the potassium that enters expanding cells during leaflet movement and the energy for potassium transport must be available from within the pulvinule.If leaflets are darkened late in the photoperiod, they close more rapidly and show lower inhibition by far red preirradiation or anaerobic conditions. A rhythmic increase in potassium efflux from ventral motor cells appears to be the basis for rhythmic promotion of nyctinastic closure. We suggest this is due to a rhythmic increase in the leakiness of ventral motor cells.

Phytochrome control of rapid nyctinastic movements and membrane permeability in Albizzia julibrissin

The rapid nyctinastic movements of Albizzia julibrissin pinnules are under the control of phytochrome. When given prior to a dark period, red light facilitates and far-red light inhibits pinnule closure in the dark. These light effects are mutually photoreversible. The opening reaction of the pinnules following a dark period is mediated mainly by blue light. The nyctinastic closure response is accompanied by an increased rate of electrolyte efflux from the cut pinna base. This observation, coupled with the fact that the rapid nyctinastic movement is not affected by actinomycin D, supports the view that phytochrome control of the sleep movement is not mediated through effects on RNA metabolism, but rather through changes in membrane permeability.