Large plasma-membrane depolarization precedes rapid blue-light-induced growth inhibition in cucumber

Blue-light (BL)-induced suppression of elongation of etiolated Cucumis sativus L. hypocotyls began after a 30-s lag time, which was halved by increasing the fluence rate from 10 to 100 mol·m^-2·s^-1. Prior to the growth suppression, the plasma-membrane of the irradiated cells depolarized by as much as 100 mV, then returned within 2–3 min to near its initial value. The potential difference measured with surface electrodes changed with an identical time course but opposite polarity. The lag time for the change in surface potential showed an inverse dependence on fluence rate, similar to the lag for the growth inhibition. Green light and red light caused neither the electrical response nor the rapid inhibition of growth. The depolarization by BL did not propagate to nonirradiated regions and exhibited a refractory period of about 10 min following a BL pulse. Fluenceresponse relationships for the electrical and growth responses provide correlational evidence that the plasma-membrane depolarization reflects an event in the transduction chain of this light-growth response.Abbreviations and symbols BL blue light - V _m membrane potential - V ₈ surface potential     

Phototropic fluence-response relations for Avena coleoptiles on a clinostat

Phototropism of Avena sativa L. has been characterized using a clinostat to negate the gravitropic response. The kinetics for development of curvature was measured following induction by a single pulse of blue light (BL), five pulses of BL at 20-min intervals, and this same pulsed-light regime following a 2-h red light (RL) pre-irradiation. A final curvature of about 14° is expressed within 180 min following the single pulse; a final curvature of about 62° in about 240 min following five pulses without pre-irradiation; and a curvature of over 125° in 360 min following five pulses after the RL pre-irradiation. For seedlings not pre-irradiated, the final curvature to five pulses of BL at a total fluence of 9.4 pmol·cm^-2 increases with time of darkness between pulses up to 15 min; with seedlings pre-irradiated with RL, curvature increased more slowly with time of darkness between pulses to a maximum at 35 min. The final curvature induced by a constant fluence of 9.4 pmol·cm^-2 increases linearly with time between the first pulse and last pulse of a five-pulse sequence. The curvature induced by a single BL pulse with a 5-min RL co-irradiation increases with fluence to a maximum of about 60° at about 10 pmol·cm^-2, and then decreases to 0° at about 200 pmol·cm^-2. Curvature induced by five BL pulses following a 2-h RL pre-irradiation increased with fluence from a threshold of about 0.05 pmol·cm^-2 to a maximum of 90° at about 10 pmol·cm^-2, and then gradually decreased with fluence to 50° at 1 000 pmol·cm^-2. Based on these data, it is concluded that the initial photoproduct formed by a BL pulse has a limited lifetime, that there is a kinetic limitation downstream of the photoreceptor pigment for phototropism, and that the additivive effect of pulsed BL is distinct from the potentiating effect of RL on phototropism. Thus, any degree of curvature from 0° to over 90° may be induced by a fluence in the ascending arm of what is traditionally called the first positive phototropic response.Abbreviations BL blue light - RL red light     

Phycomyces: Phototropism and light-growth response to pulse stimuli

The relationship between phototropism and the light-growth response of Phycomyces blakesleeanus (Burgeff) sporangiophores was investigated. After dark adaptation, stage-IVb sporangiophores were exposed to short pulses of unilateral light at 450 nm wavelength. The sporangiophores show a complex reaction to pulses of 30 s duration: maximal positive bending at 3·10^-4 and 10^-1 J m^-2, but negative bending at 30 J m^-2. The fluence dependence for the light-growth response also is complex, but in a different way than for phototropism; the first maximal response occurs at 1.8·10^-3 J m^-2 with a lesser maximum at 30 J m^-2. A hypertropic mutant, L85 (madH), lacks the negative phototropism at 30 J m^-2 but gives results otherwise similar to the wild type. The reciprocity rule was tested for several combinations of fluence rates and pulse durations that ranged from 1 ms to 30 s. Near the threshold fluence (3·10^-5 J m^-2), both responses increase for pulse durations below 67 ms and both have an optimum at 2 ms. At a fluence of 2.4·10^-3 J m^-2, both responses decrease for pulse durations below 67 ms. The hypertropic mutant (madH), investigated for low fluence only, gave similar results. In both strains, the time courses for phototropism and light-growth response, after single short pulses of various durations, show no clear correlation. These results imply that phototropism cannot be caused by linear superposition of localized light-growth responses; rather, they point to redistribution of growth substances as the cause of phototropism.     

Inhibition of aggregation by light in the cellular slime mold Dictyostelium discoideum

Aggregation of Dictyostelium amoebae is inhibited by light. White light intensities 10² W · cm^-2 cause an inhibition which reaches a saturation at 2 · 10³ W · cm^-2. The action spectrum, based on photon fluence-response curves, shows a major peak around 405 nm and extends through most of the visible spectrum with a secondary maximum at about 530 nm. The action spectrum of the inhibition of aggregation resembles the action spectrum of accumulations of amoebae in light traps and the action spectrum of photodispersal from light traps; it does not resemble the action spectrum of phototaxis in pseudoplasmodia.     

Calcium influx at the plasmalemma of isolated guard cells of Commelina communis

The influx of ⁴⁵Ca into isolated guard cells of Commelina communis L. has been measured, using short uptake times, and washing in ice-cold La³⁺-containing solutions to remove extracellular tracer after the loading period. Over 0.5–4 min the uptake was linear with time, through the origin. Over 20–200M external Ca²⁺ the influx measured with 10–20 mM external KCl was in the range 0.3–2.3 pmol·cm^-2·s^-1 (on the basis of estimated guard-cell area); with only 1 mM KCl externally the ⁴⁵Ca influx was significantly reduced, in the range 0.3–1.1 pmol·cm^-2·s^-1 for external Ca²⁺ of 50–100 M. The results indicate that the Ca-channel is voltage-sensitive, opening with depolarisation. No consistent effect of the addition of abscisic acid could be found. In different experiments, on the addition of 0.1 mM abscisic acid the Ca²⁺ influx was sometimes stimulated by 28–79%, was sometimes unaffected, and was sometimes inhibited by 16–29%. The results rule out a long-lasting stimulation of ⁴⁵Ca influx by ABA, but they do not rule out a transient stimulation followed by inhibition, perphaps as a consequence of down-regulation of Ca²⁺ influx by increasing cytoplasmic Ca²⁺. The hypothesis that ABA may act via an action on Ca²⁺ influx, increasing cytoplasmic Ca²⁺, with consequent effects on voltage-dependent and Ca²⁺-dependent ion channels in both plasmalemma and tonoplast, is neither proved nor disproved by these results.Abbreviations ABA abscisic acid - Ca_o, K_o external Ca and K concentrations     

The effect of organic calcium channel modulators on the germination ofVaucheria longicaulis aplanospores

Summary InVaucheria longicaulis var.macounii aplanospore germination and filament growth are severely inhibited by the Ca²⁺-channel antagonists (–)202–791, diltiazem, nifedipine and verapamil, whereas the agonists (+)202–791, Bay K-8644 and CGP-28392 stimulate those processes. Both antagonist and agonist actions suggest that voltage-controlled Ca²⁺ influx plays a major role in the regulation of the initial events of germination and filament growth. Increases in⁴⁵Ca²⁺ influx are observed after pretreatment of the aplanospores with low temperature shocks of brief duration or FCCP. Both agents are known to depolarize the surface membrane.⁴⁵Ca²⁺ influx is reduced in material treated with FC, an agent known to hyperpolarize cell membranes. The results indicate that Ca²⁺ influx takes place through voltage-sensitive Ca-channels.Abbreviations Bay K-8644 methyl 1,4-dihydro-2,6-dimethy1-3-nitro-4-(2-trifluoromethylphenyl)-pyridine-5-carboxylate - CGP CGP-28392 - CTC chlorotetracycline - dil diltiazem - DMSO dimethyl sulphoxide - DTE dithioerythritol - EGTA ethyleneglycol-bis-(-aminoethylether) N,N¹-tetraacetic acid - FC fusicoccin - FCCP p-fluoromethoxy-(carbonylcyanide)phenylhydrazone - nif nifedipine - PCMB p-chloromercuribenzoate - ver verapamil     

Action spectra of the light-growth response of Phycomyces

The light-growth response of Phycomyces blakesleeanus (Burgeff) is a transient change in elongation rate of the sporangiophore caused by a change in light intensity. Previous investigators have found that the light-growth response has many features in common with phototropism; the major difference is that only the light-growth response is adaptive. In order to better understand the light-growth response and its relationship to phototropism, we have developed a novel experimental protocol for determining light-growth-response action spectra and have examined the effect of the reference wavelength and intensity on the shape of the action spectrum. The null-point action spectrum obtained with broadband-blue reference light has a small peak near 400 nm, a flat region from 430 nm to 470 nm, and an approximately linear decline in the logarithm of relative effectiveness above 490 nm. The shape of the action spectrum is different when 450-nm reference light is used, as has been shown previously for the phototropic-balance action spectrum. However, the action spectrum of the light-growth response differs from that for phototropic balance, even when the same reference light (450 nm) is used. Moreover, for the light-growth response, the relative effectiveness of 383-nm light decreases as the intensity of the 450-nm reference light increases; this trend is the opposite of that previously found for phototropic balance. The dependence of the lightgrowth-response action spectrum on the reference wavelength, its difference from the phototropic-balance action spectrum, and the reference-intensity dependence of the relative effectiveness at 383 nm may be attributable to dichroic effects of the oriented photoreceptor(s), and to transduction processes that are unique to the light-growth response.I dedicated to Masaki Furuya on the occasion of his 65th birthdayThis work was supported by a grant from the National Institutes of Health (GM29707) to E.D. Lipson. Anuradha Palit, Promod Pratap, and Benjamin Horwitz participated in the early phases of this work. We thank Leonid Fukshansky and Benjamin Horwitz for helpful discussions, David Durant for computer programming, and Steven Block for providing us with a C-language program of Reinsch's procedure for cubic spline interpolation. One of us (R.S.) gratefully acknowledges a junior faculty fellowship leave from the Department of Physics at Yale University.     

HCO 3 - and OH-transport across the plasmalemma ofChara

Internodal and whorl (branch) cells of the green alga,Chara corallina Klein ex Willd., em. R.D.W., were studied with the extracellular vibrating probe for measuring transmembrane ion currents, and with an extracellular pH microprobe for measuring the surface pH profile. Bands of positive inward current (OH^- efflux) 1–3 mm wide were separated by wider bands of outward current (HCO ₃ ^- influx) along the length of the cell. The measured peaks of inward current ranged from 20 to 60 A cm^-2 (98 m from the cell surface) which would correspond to a surface ionic flux of 270–800 pmol cm^-2 s^-1. The peaks of outward current (HCO ₃ ^- influx) ranged from 10 to 30 A cm^-2 which would correspond to a surface ionic flux of 140–400 pmol cm^-2 s^-1. The inward current bands matched the regions of surface alkalinity very well. The outward current (HCO ₃ ^- influx) was reduced at least 10-fold in low-HCO ₃ ^- medium, with a commensurate readjustment in the strength and pattern of inward current (OH^- efflux). (Although these experiments involved a manipulation of the external pH, it is felt that the main adjustment in current patterns was in response to the reduction in exogenous HCO ₃ ^- ). The presence of the vibrating probe perturbed the inward current region when vibrating with a 26-m amplitude, but this perturbation was eliminated when a 7-m amplitude was used. The perturbation was usually observed as a reduction in the number of inward current peaks with an increase (approximate doubling) in the amplitudes of the one or two remaining peaks. Both the inward and outward currents were light-dependent, falling off within seconds of light removal.     

Negative phototropism in the piloboloid mutants of Phycomyces blakesleeanus Bgff.

The sporangiophore (spph) of a piloboloid mutant, genotype pil, of Phycomyces ceases elongation and expands radially in the growth zone shortly after reaching the developmental stage IV b. The pil spph is always negatively phototropic to unilateral visible light when its diameter exceeds 210 m. Photoinduction of spph initiation, light-growth response, threshold of light energy fluence rate for the negative phototropism, avoidance and gravitropism in the pil mutant are all normal. In liquid paraffin, the pil spph shows negative phototropism as does the wild-type spph. Genetic analyses indicate that the negative phototropism of the pil mutant is governed by the phenotypic characteristics of pil but not by specific gene(s) responsible for negative phototropism. These facts imply that the reverse phototropism of the pil mutant results from a loss of the convergent lens effect of the cell because of the increase in cell diameter.Abbreviations spph(s) sporangiophore(s) - wt(s) wild type(s)     

Some regulatory aspects of [14C]methylamine influx intoPisum sativum L. cv. Feltham First seedlings

[¹⁴C]Methylamine influx intoPisum sativum L. cv. Feltham First seedlings showed Michaelis-Menten-type kinetics with apparentV _max=49.2 mol·g^-1 FW·h^-1 and apparentK _m=0.51 mM. The competitive interactions between ammonium and methylamine were most obvious when biphasic kinetics were assumed with saturation of the first phase at 0.05 mM. The inhibitor constant for ammonium (K _i)=0.027 mM. When [¹⁴C]methylamine was used in trace amounts with ammonium added as substrate, the influx of tracer showed Michaelis-Menten-type kinetics with apparentV _max=3.46 mol·g^-1 FW·h^-1 and apparentK _m=0.15 mM. The initial rate of net ammonium uptake corresponded with that found when [¹⁴C]methylamine was used to trace ammonium influx. The latter was also stimulated by high pH_o and inhibited by nitrate. Ammonium pretreatment±methionine sulphoximine or glutamine pretreatment of the seedlings inhibited subsequent [¹⁴C]methylamine influx, while methylamine or asparagine pretreatment stimulated [¹⁴C]methylamine influx. There was also a stimulatory effect of prior inoculation withRhizobium. The results are discussed in terms of current models for the regulation of ammonium uptake in plants.     

Regulation of aplanospore germination in Vaucheria

Germination of aplanospores in Vaucheria longicaulis Hoppaugh var. macounii Blum proceeds through three stages of development. Stage I begins with the initiation of germination and lasts approx. 2 h. During this stage germinating filaments grow at an accelerated rate (266 ± 12 m · h^–1). Stage II is characterized by a sharp decline in the growth rate of germinating filaments (96 ± 4 m · h^–1) and lasts 4 h. This is followed, during the next 4 h, by a recovery in the growth rate (168 ± 8 m · h^–1) of germinating filaments, stage III. Growth rates stabilize and remain unchanged during subsequent development (Oliveira and Fitch, 1988, J. Submicrosc. Cytol. Pathol. 20, 397–406). The Ca²⁺-influx modulators LaCl₃, nifedipine and methyl 1,4-dihydro-2,6-dimethyl-3-nitro-4 (2-trifluoromethylphenyl)-pyridine-5-carboxylate (Bay K-8644), the ionophore calcimycin (A23187), the intracellular Ca²⁺-release antagonist 8-N-N'-(diethylamino)-octyl-3,4, 5-trimethoxybenzoate (TMB-8), the Ca²⁺-uptake inhibitor ruthenium red and the phosphoinositide-cycle modulators LiCl and myo-inositol show that the events required for the initiation are distinct from those required for the completion of each stage of germination. These studies in conjunction with microinjection of germinating filaments with inositol 1,4,5-trisphosphate, the natural ligand for Ca²⁺ release from Ca-storing organelles (endoplasmic reticulum, vacuole), and treatment with chlorotetracycline (CTC), to visualize the distribution of membrane-bound Ca²⁺ reveal that both the initiation and completion of each stage of germination are controlled by Ca²⁺ signals which are restricted to well-defined time intervals and are modulated by the origin (source) of Ca²⁺.Abbreviations BAPTA 1,2-bis(2-aminophenoxy)ethane-N,N,N,N-tetraacetic acid - Bay K-8644 methyl 1,4-dihydro-2,6-dimethyl-3-nitro-4(2-trifluoromethylphenyl)-pyridine-5-carboxylate - CTC chlorotetracycline - InsP₃ inositol 1,4,5-trisphosphate - RR ruthenium red - TMB-8 8-N-N-(diethylamino)-octyl-3,4,5-trimethoxybenzoate The author wishes to express his gratitude to the technical group of the Immunocytochemistry Unit for their help with the microinjection studies. This work was supported by a grant from the Natural Sciences and Engineering Research Council of Canada (grant A-7844).     

Characterization of the Na+-dependent taurine influx in flounder erythrocytes

About 92% of the taurine influx in flounder erythrocytes at physiological conditions in vitro (330 mosmol·l^-1, 145 mmol·l^-1 Na⁺, 0.30 mmol·l^-1 taurine) is Na⁺-dependent. This influx is highly specific for taurine. The -amino compounds hypotaurine and -alanine were the only compounds which mimicked the inhibitory effect of taurine on influx of [¹⁴C]taurine, the former more than the latter. Counterexchange of taurine was also mediated by the taurine transporters. Reduction of osmolality per se did not affect the activity of these transporters. Non-linear regression analysis of the influx values revealed the presence of two different influx systems: a system with high affinity and low capacity and another with low affinity and high capacity. However, we cannot exclude the possibility that this influx of taurine was mediated by only one transporter which operated in different modes depending on the extracellular Na⁺ concentration. On the assumption that the Na⁺-dependent influx was mediated by two separate systems, the maximal velocity of the low capacity system was 2.55 nmol·g dry weight^-1·min^-1 at 145 mmol·ll^-1 extracellular Na⁺. This capacity was about 50% lower than that of the high capacity system. The Michaelis constants were 0.013 and 1.34 mmol·l^-1, respectively. Reduction of the extracellular Na⁺ concentration reduced maximal velocity and the affinity to taurine of both transport systems. At 10 mmol·l^-1 Na⁺ or lower concentrations the high capacity system did not seem to operate. The activation method suggested that each taurine molecule transported by the high capacity system was accompanied by two Na⁺. The stoichiometry of the low capacity system was 1 taurine: 1 Na⁺. The Hill-coefficient for both transport systems was 1.00.Abbreviations cpm counts per minute - dw dry weight - GABA -amino-n-butyric acid - K _m Michaelis constant - pK _b basic dissociation constant - SD standard deviation - -ABA Dl--amino-n-butyric acid - V _max maximal velocity - ww wet weight     

Ionic basis of currents in somatic embryos of Daucus carota

A vibrating probe was used to measure extracellular electrical currents around developing somatic embryos in two lines (RCC27, RCC48) of cultured cells of Daucus carota L. at the heart and torpedo stages. At pH 5.5, an inward current of 1.2±0.1 A·cm^-2 (n=23) was detected at the cotyledon, and an outward current of 1.0±0.1 A·cm^-2 (n=22) was found at the radicle in torpedostage embryos from the RCC27 line. At a pH of 5.75 the currents increased by 0.2–0.3 A·cm^-2 (n=60–62). In a few cases an additional small inward current was detected at the tip of the radicle in toepedo-stage embryos from RCC27 line. Such an inward current at the radicle seemed to appear earlier, some time after the heart stage, in embryos from the RCC48 line.Both extracellular pH measurements (using microelectrodes filled with ion-sensitive resin) and ion-substitution studies were carried out in order to ascertain the ionic composition of the currents in torpedo-stage embryos from the RCC27 line. Regions adjacent to the cotyledon and radicle, at the points of current entry and exit, were found to be more acidic by 0.02±0.01 (n=14) and 0.07±0.01 (n=12) pH units, respectively, than the bulk medium. Removal of K⁺ from the medium reversibly reduced the currents to about 25% of their original value at both cotyledon and radicle. Deletion of Cl^- decreased the currents slightly. Removal of Ca²⁺ resulted in a rapid doubling of currents. Addition of either N,N-dicyclohexylcarbodiimide or tetraethyl ammonium chloride substantially reduced overall currents, and their removal resulted in partial recovery of the currents. It is suggested that the inward current at the cotyledon is comprised largely of K⁺ influx and the outward current at the radicle is mainly the result of active H⁺ efflux.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - MS Murashige and Skoog     

Ion transport across leech integument

The dorsal skin of the leech Hirudo medicinalis was used for electrophysiological measurements performed in Ussing chambers. The leech skin is a tight epithelium (transepithelial resistance = 10.5±0.5 k· cm^-2) with an initial short-circuit current of 29.0±2.9 A·cm^-2. Removal of Na⁺ from the apical bath medium reduced short-circuit current about 55%. Ouabain (50mol·l^-1) added to the basolateral solution, depressed the short-circuit current completely. The Na⁺ current saturated at a concentration of 90 mmol Na⁺·l^-1 in the apical solution (K _M=11.2±1.8 mmol·l^-1). Amiloride (100 mol·l^-1) on the apical side inhibited ca. 40% of the Na⁺ current and indicated the presence of Na⁺ channels. The dependence of Na⁺ current on the amiloride concentration followed Michaclis-Menten kinetics (K _i=2.9±0.4 mol·l^-1). The amiloride analogue benzamil had a higher affinity to the Na⁺ channel (K _i=0.7±0.2 mol·l^-1). Thus, Na⁺ channels in leech integument are less sensitive to amiloride than channels known from vertebrate epithelia. With 20 mmol Na⁺·l^-1 in the mucosal solution the tissue showed an optimum amiloride-inhibitable current, and the amiloride-sensitive current under this condition was 86.8±2.3% of total short-circuit current. Higher Na⁺ concentrations lead to a decrease in amiloride-blockade short-circuit current. Sitmulation of the tissue with cyclic adenosine monophosphate (100 mol·l^-1) and isobutylmethylxanthine (1 mmol·l^-1) nearly doubled short-circuit current and increased amiloride-sensitive Na⁺ currents by 50%. By current fluctuation analysis we estimated single Na⁺ channel current (2.7±0.9 pA) and Na⁺ channel density (3.6±0.6 channels·m^-2) under control conditions. After cyclic adenosine monophosphate stimulation Na⁺ channel density increased to 5.4±1.1 channels·m^-2, whereas single Na⁺ channel current showed no significant change (1.9±0.2 pA). These data present a detailed investigation of an invertebrate epithelial Na⁺ channel, and show the similarities and differences to vertebrate Na⁺ channels. Whereas the channel properties are different from the classical vertebrate Na⁺ channel, the regulation by cyclic adenosine monophosphate seems similar. Stimulation of Na⁺ uptake by cyclic adenosine monophosphate is mediated by an increasing number of Na⁺ channels.Abbreviations slope of the background noise component - ADH antidiuretic hormone - cAMP cyclic adenosine monophosphate - f frequency - f _c coner frequency of the Lorentzian noise component - Hepes N-hydroxyethylpiperazine-N-ethanesulphonic acid - BMX isobutyl-methylxanthine - i _Na single Na⁺ channel current - I _Na max, maximal inhibitable Na⁺ current - I _SC short circuit current - K _i half maximal blocker concentration - K _M Michaelis constandard error of the mean - S _(f) power density of the Lorentzian noise component - S ₀ plateau value of the Lorentzian noise component - TMA tetramethylammonium - Trizma TRIS-hydroxymethyl-amino-methane - V _max maximal reaction velocity - V _T transepithelial potential - K half maximal blocker concentration     

Long-term kinetics of the light-dependent regulation of ribulose-1,5-bisphosphate carboxylase/oxygenase activity in plants with and without 2-carboxyarabinitol 1-phosphate

The light-dependent modulation of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) activity was studied in two species: Phaseolus vulgaris L., which has high levels of the inhibitor of Rubisco activity, carboxyarabinitol 1-phosphate (CA1P), in the dark, and Chenopodium album L., which has little CA1P. In both species, the ratio of initial to fully-activated Rubisco activity declined by 40–50% within 60 min of a reduction in light from high a photosynthetic photon flux density (PPFD; >700 mol · m^–2 · s^–1) to a low PPFD (65 ± 15 mol · m^–2 · s^–1) or to darkness, indicating that decarbamylation of Rubisco is substantially involved in the initial regulatory response of Rubisco to a reduction in PPFD, even in species with potentially extensive CA1P inhibition. Total Rubisco activity was unaffected by PPFD in C. album, and prolonged exposure (2–6 h) to low light or darkness was accompanied by a slow decline in the activity ratio of this species. This indicates that the carbamylation state of Rubisco from C. album gradually declines for hours after the large initial drop in the first 60 min following light reduction. In P. vulgaris, the total activity of Rubisco declined by 10–30% within 1 h after a reduction in PPFD to below 100 mol · m^–2 · s^–1, indicating CA1P-binding contributes significantly to the reduction of Rubisco capacity during this period, but to a lesser extent than decarbamylation. With continued exposure of P. vulgaris leaves to very low PPFDs (< 30 mol · m^–2 · s^–1), the total activity of Rubisco declined steadily so that after 6–6.5 h of exposure to very low light or darkness, it was only 10–20% of the high-light value. These results indicate that while decarbamylation is more prominent in the initial regulatory response of Rubisco to a reduction in PPFD in P. vulgaris, binding of CA1P increases over time and after a few hours dominates the regulation of Rubisco activity in darkness and at very low PPFDs.Abbreviations CA1P 2-carboxyarabinitol 1-phosphate - CABP 2-carboxyarabinitol 1,5-bisphosphate - k_cat substrate-saturated turnover rate of fully carbamylated enzyme - PPFD photosynthetically active photon flux density (400–700 nm) - Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase - RuBP ribulose-1,5-bisphosphate     

Sulphate influx in wheat and barley roots becomes more sensitive to specific protein-binding reagents when plants are sulphate-deficient

When young wheat (Triticum aestivum L.) or barley (Hordeum vulgare L.) plants were deprived of an external sulphate supply (-S plants), the capacity of their roots to absorb sulphate, but not phosphate or potassium, increased rapidly (derepression) so that after 3–5 d it was more than tenfold that of sulphate-sufficient plants (+S plants). This increased capacity was lost rapidly (repression) over a 24-h period when the sulphate supply was restored. There was little effect on the uptake of L-methionine during de-repression of the sulphate-transport system, but S input from methionine during a 24-h pretreatment repressed sulphate influx in both+S and-S plants.Sulphate influx of both+S and-S plants was inhibited by pretreating roots for 1 h with 4,4-diisothiocyanatostilbene-2,2-disulphonic acid (DIDS) at concentrations > 0.1 mol · m^-3. This inhibition was substantially reversed by washing for 1 h in DIDS-free medium before measuring influx. Longer-term pretreatment of roots with 0.1 mol·m^-3 DIDS delayed de-repression of the sulphatetransport system in-S plants but had no influence on+S plants in 3 d.The sulphydryl-binding reagent, n-ethylmaleimide, was a very potent inhibitor of sulphate influx in-S roots, but was much less inhibitory in +S roots. Its effects were essentially irreversible and were proportionately the same at all sulphate concentrations within the range of operation of the high-affinity sulphate-transport system. Inhibition of influx was 85–96% by 300 s pretreatment by 0.3 mol·m^-3 n-ethylmaleimide. No protection of the transport system could be observed by including up to 50 mol·m^-3 sulphate in the n-ethylmaleimide pre-treatment solution. A similar differential sensitivity of-S and+S plants was seen with p-chloromercuriphenyl sulphonic acid.The arginyl-binding reagent, phenylglyoxal, supplied to roots at 0.25 or 1 mol·m^-3 strongly inhibited influx in-S wheat plants (by up to 95%) but reduced influx by only one-half in+S plants. The inhibition of sulphate influx in-S plants was much greater than that of phosphate influx and could not be prevented by relatively high (100 mol·m^-3 sulphate concentrations accompanying phenylglyoxal treatment. Effects of phenylglyoxal pretreatment were unchanged for at least 30 min after its removal from the solution but thereafter the capacity for sulphate influx was restored. The amount of new carrier appearing in-S roots was far greater than in+S roots over a 24-h period.The results indicate that, in the de-repressed state, the sulphate transporter is more sensitive to reagents binding sulphydryl and arginyl residues. This suggests a number of strategies for identifying the proteins involved in sulphate transport.Abbreviations DIDS 4,4-diisothiocyanatostilbene-2,2-disulphonic acid - NEM n-ethylmaleimide - PCMBS p-chloromercuriphenyl sulphonic acid     

Effects of six trace metals on calcium fluxes in brown trout (Salmo trutta L.) in soft water

Summary Calcium fluxes were measured simultaneously in brown trout fry maintained in an artificial soft water medium of [Ca] 20 mol·l^-1 and pH 5.6, and exposed to each of six trace metals (Al, Cu, Fe, Ni, Pb, and Zn). The trace metal concentrations represented typical and maximum levels found in acid waters experiencing declining fishery status. In the absence of trace metals, evidence is presented which suggests that ca. 91% of Ca taken up from the external medium was by extraintestinal active transport. Calcium efflux was stimulated by both concentrations of Al, Cu, Fe, and Pb. Efflux was also stimulated by [Ni] 170 nmol·l^-1 and [Zn] 3000 nmol·l^-1. In some cases, response to increased efflux was stimulation of influx. Lack of stimulation of influx resulted in negative net Ca fluxes. Net Ca losses were recorded at both concentrations of Al, Pb, and Ni, lower concentrations only of Fe, and higher concentrations only of Cu and Zn.Abbreviations J _in influx - J _net net flux - J _out efflux Henceforward in this paper, chemical elements are referred to by their chemical symbols rather than by full names     

Phototactic responses in Haematococcus lacustris and its modification by light intensity and the carotenoid biosynthesis inhibitor Norflurazon

At fluence rates below 45 W· m^-2 cells of the flagellate stage of Haematococcus lacustris react only positively phototactically with a rather high degree of orientation (indicated by r values up to 0.66 with the Rayleigh test). The directedness of orientation decreases with decreasing irradiance. The degree of directedness of the phototactic response depends on the intensity of preirradiation: Low light intensity applied after strong light application results in a dark reaction (low r values), low light given after darkness stimulates a rather high degree of directedness of positive phototaxis. Weak blue light (=483 nm; 0.4 W · m^-2) stimulates positive phototactic response, whereas comparable red light (=658 nm; 0.5 W · m^-2) does not.Cells which were grown in a medium containing 10^-4 M Norflurazon (effective in inhibition of carotenoid biosynthesis) although maintaining motility completely lose the ability to react positively phototactically. The possible role of carotenoids in the phototactic orientation is discussed.     

Redistribution of growth during phototropism and nutation in the pea epicotyl

First positive phototropism of the third internode of intact, 5-d-old pea (Pisum sativum L.) seedlings, grown under continuous, dim red light, showed maximal response following a photon fluence of 3 mol·m^-2 blue light. Greater or lesser fluences (with irradiation time 100 s or less) caused less bending, no response being detectable above 300 or below 0.03 mol·m^-2. Bilateral irradiation with blue light caused no detectable inhibition of growth rate over that range of fluences. The linear nutation of the pea third internode was shown to be driven by a balanced oscillation of growth rate such that the overall growth rate was little changed during the oscillation. Phototropic stimulation changed neither the amplitude nor the period of nutation. Nutation and phototropism probably regulate growth independently. Phototropism in response to the optimal blue light fluence was caused by concomitant depressed growth on the irradiated side and stimulated growth on the shaded side of the bending internode. These results are consistent with the Cholodny-Went hypothesis which states that unilateral blue light induces a lateral redistribution of a growth regulator.Abbreviations R red light - BL blue light Carnegie Institution, Department of Plant Biology paper No. 921     

Uptake of phosphate by symbiotic and free-living dinoflagellates

Uptake of phosphate in the light by Amphidinium carterae, Amphidinium klebsii, cultured and symbiotic Gymnodinium microadriaticum conformed to Michaelis-Menten type saturation kinetics with all organisms showing similar K _m values, namely 0.005 to 0.016 M phosphorus. V _max values were 0.009–0.32 nmol phosphorus · 10⁵ cells^-1 · 10 min^-1. Phosphate uptake by all the dinoflagellates was greater in the dark than in the light. The metabolic inhibitor 3-(3,4-dichlorophenyl) 1,1-dimethylurea stimulated phosphate uptake in the light by A. carterae and A. klebsii, but inhibited uptake by cultured and symbiotic G. microadriaticum. Carbonylcyanide 3-chlorophenylhydrazone (CCCP) inhibited phosphate uptake by A. carterae and A. klebsii under both light and dark conditions. Uptake of phosphate by cultured and symbiotic G. microadriaticum in the light, but not in the dark, was inhibited by CCCP. Low concentrations of arsenate (5 g As · l^-1) stimulated phosphate by A. carterae and A. klebsii, but inhibited uptake by cultured and symbiotic G. microadriaticum. High concentrations of arsenate (100 g As · l^-1) did not affect uptake of phosphate by A. carterae and A. klebsii.