Effects of Moderately Low Temperature (20°C) on Phytochrome Responses During Preirradiation: Anthocyanin Synthesis in Sorghum bicolor at High- and Low-Pfr/Ptot Ratios

Abstract— Effects on phytochrome-mediated anthocyanin synthesis of moderately low temperature (MLT) given during the preirradiation culture period were studied with seedlings of broom sorghum ( Sorghum bicolor Moench, cvs. Acme Broomcorn and Sekishokuzairai-Fukuyama). Seedlings were grown in the dark at 20°C (MLT) and 24°C (control). The MLT treatment strikingly enhanced the action induced by a red light (R) pulse above ca 200 μmol m⁻² and suppressed the action induced by an R pulse below ca 30 μmol m⁻² and by a far-red light (FR) pulse alone. We refer to these MLT-affected distinct responses as "high-Pfr/Ptot response" and "low-Pfr/Ptot response," as they have features different from the high-irradiance and very-low-fluence responses, respectively. The destruction rate of spectroscopically detectable phytochrome (phyA) and the time course of escape of anthocyanin synthesis from FR reversibility did not match, and hence the possibility of phyA being involved in high-Pfr/Ptot response was rejected, although it might be involved in low-Pfr/Ptot response. Possible mechanisms for the two distinct phytochrome responses are discussed.     

PHOTOREVERSIBLE Ca2+-DEPENDENT AGGREGATION OF PURIFIED PHYTOCHROME FROM ETIOLATED PEA AND RYE SEEDLINGS

The aggregation of phytochrome purified from etiolated pea ( Pisum satirum cv. Alaska) and rye ( Secale cereale cv. Cougar) tissues was investigated by centrifugation and turbidimetry. Purified pea phytochrome (A₆₆₉/A₂₈₀= 0.88), if irradiated with red light, became precipitable in the presence of CaCl₂. The precipitation upon red-light irradiation was optimal at a Ca^2- or Mg²⁺ concentration of 10–20 m M , was greater at increased phytochrome concentration or lower pH values, and was inhibited by 0.1 M KG. The precipitated phytochrome slowly became soluble after far-red light exposure.
Turbidity of pea phytochrome solutions after red-light irradiation also increased rapidly in the presence of either Ca²⁺ or Mg²⁺. Far-red light exposure after the red light cancelled the turbidity increase. Rye phytochrome showed less turbidity increase than pea phytochrome and occurred only in the presence of Ca²⁺. Partially degraded pea phytochrome produced by endogenous proteases in the extract did not show the turbidity increase. Undegraded pea phytochrome also associated with microsomal fractions under conditions similar to those described above, but the partially degraded phytochrome did not.     

SPECIFIC PHOTOREGULATION BY PHYTOCHROME OF EPINASTY and LIGHT-INDUCED ETHYLENE PRODUCTION IN MARCHANTIA POLYMORPHA*

Abstract— A study is made of the influence of phytochrome on light-induced ethylene production and epinasty in green Marchantia polymorpha thalli. Ethylene production of thalli irradiated with terminal far-red is not substantially affected by the presence or absence of CO₂, in contrast with the controls which show a clearcut CO₂ dependency. Photoreversibility and fiuence response data indicate the involvement of the low energy red'far-red reversible type of phytochrome action for both the light-induced ethylene production and the control of epinasty. The far-red effect is further characterized by a difference in escape from reversibility between light-induced ethylene production and the elimination of epinasty.     

PHOTOBIOLOGIE DU MÉSOCOTYLE D'AVOINE—II. STIMULATION DE L'ÉLONGATION PAR LA LUMIÈRE ROUGE-SOMBRE EN PRÉSENCE DE DIVERSES GIBBÉRELLINES

Abstract— Far-red light has the property of promoting the elongation of Avena mesocotyl sections in the presence of a gibberellin. Part of this effect is not reversible by red light (around 655 mμ ). The wavelengths which are most effective are those in the neignbourhood of 720 mμ , as revealed by the action spectrum. Amongst the 8 gibberellins tested, only A₁, A₃, and A₉, are capable of producing, after far-red irradiation, a greater elongation than the one they cause in the dark.     

"ENERGY"-DEPENDENT QUENCHING OF CHLOROPHYLL FLUORESCENCE and THERMAL ENERGY DISSIPATION IN INTACT LEAVES DURING INDUCTION OF PHOTOSYNTHESIS

Abstract— Intact leaves, previously adapted to darkness for a prolonged period of time, were suddenly illuminated with a strong, photosynthetically saturating, white light (ca 1500 μmol m⁻² s^_1), resulting in the rapid establishment of a large energy-dependent chlorophyll fluorescence quenching (q_E) as shown by in vivo fluorescence measurements with a pulse amplitude modulation technique. Two different photothermal methods, photoacoustic spectroscopy and photothermal deflection spectroscopy, were used to monitor thermal deactivation of excited pigments during the dark-light transitions. The in vivo photothermal signals measured with both techniques were shown to remain constant during induction of photosynthesis under high light conditions, suggesting that, in contrast to current hypotheses, energy-dependent quenching q_E is not associated with significant changes in thermal dissipation of absorbed light energy in the chloroplasts. When photosynthesis was induced with a low-intensity modulated light, a noticeable decrease in the heat emission yield was observed resulting from the progressive activation of the competing photochemical processes.     

PHYTOCHROME CONVERSION BY ULTRAVIOLET LIGHT

Abstract— Light absorbed primarily by the protein of phytochrome is active in transforming both the red and far-red absorbing forms. P _r and P _fr. The ratio of quantum yields for the conversions of P _r and P _fr by u.v. radiation (φ_r/φ_fr)_u.v.= 1.5 and does not differ significantly from the ratio obtained with red and far-red light absorbed directly by the chromophores (φ_r/φ_fr)v_vis. Thus, the efficiency of energy transfer from protein to chromophore is essentially the same for both forms of the chromoprotein. The ratio of the relative quantum yields for u.v. and visible light (φ^r)^u.v./(φ^r)^vis was 0.32 indicating that 30–35 per cent of the light energy absorbed by the protein was transferred to the chromophore.     

ON THE PRIMARY PHOTOPROCESS OF 124-kdalton PHYTOCHROME

The photoreaction between P_τ and the first detectable intermediate, lumi-R, of 124-kdalton oat phytochrome has been investigated at low temperatures. The temperature dependence of the quantum yields of the photoreactions, P_τ to lumi-R and lumi-R to P_τ, has been determined. From measurements over a temperature range from 119 to 155 K, an activation barrier of 3.6 ± 0.5 kJ mol ₁ is found for the photoreaction of P_τ with 661-nm actinic light. A higher value (5.7 ± 0.7 kJ mol ^-1) is found for the photoreaction of lumi-R to P^τ. with 698-nm actinic light. Increased quantum yields are found in deuterated buffer solutions at low temperatures. The activation energies for deuterated phytochrome (3.2 ± 0.7 kJ mol^–1 for P_τ with 661-nm irradiation and 6.2 ± 1.2 kJ mol^-1 for lumi-R at 698-nm irradiation) are identical within the limits of error with those of protonated phytochrome. The lack of a deuterium effect for the activation energies favors the Z,E-isomerization rather than proton transfer or tautomerization for the chromophore photochemistry during P_τ⇄lumi-R conversion.     

COMPARATIVE PHOTOCHEMICAL ANALYSIS OF HIGHLY PURIFIED 124 KILODALTON OAT and RYE PHYTOCHROMES in vitro

Abstract A direct comparison of the photochemical interconversions between red (P_r-) and far-red (P_fr-) absorbing forms of highly-purified 124 kDa oat and rye phytochromes under identical experimental conditions was performed. In two different buffer systems at 5°C, the quantum yields for the P_r to P_tr and P_fr to P_r phototransformations under constant red and far-red illumination, φ _r and φ_fr respectively, were determined to be 0.152-0.154 and 0.060-0.065 for oat preparations and 0.172-0.174 and 0.074-0.078 for rye preparations. These values as well as the wavelength dependence of the photoequilibrium produced under continuous illumination throughout the visible and near-ultraviolet spectrum were based on the absorption spectra of the two phytochrome preparations and revised molar absorption coefficients. The molar absorption coefficients were estimated by quantitative amino acid analysis and shown to be identical for the two monocot phytochromes (i.e. 132 mM ⁻¹ cm⁻¹ at the red absorption maximum for the P_r form). Because these measurements were performed under identical experimental conditions, including buffer, temperature, light fluence rate, and instrumentation, the differences observed must reflect structural features inherent to the two different monocotyledonous phytochromes.     

Porphyrins and Related Compounds as Photoactivatable Insecticides I. Phototoxic Activity of Hematoporphyrin Toward Ceratitis capitata and Bactrocera oleae

The photodynamic sensitizer hematoporphyrin (HP) IX was efficiently accumulated by Ceratitis capitata (Mediterranean fruit fly) and Bactrocera oleae (olive fly) when the insects were fed with a sugar/protein bait containing micromolar amounts of porphyrin. Hematoporphyrin appeared to be mainly accumulated in the midgut, Malpighian tubes, adipose tissue and cuticle and was gradually cleared from the organism in a 24–48 h time interval. Exposure of the HP-fed flies to light mimicking the solar spectrum caused a decrease in the survival whose extent was modulated by the HP concentration in the bait, the irradiation fluence rate and the total light dose. For 8 μmol/mL HP in the bait 100% mortality during 1 h exposure to light was obtained using a fluence rate of 1220 μE s⁻¹ m⁻² for C. capitata and 2080 μE s⁻¹ m⁻² for B. oleae . The latter fly was somewhat less photosensitive than C. capitata possibly owing to the smaller amount of ingested HP and/or darker pigmentation. Studies are in progress in order to extend these investigations from the laboratory to the field scale.     

PHYTOCHROME-MEDIATED PHOTOTROPISM IN PROTONEMATA OF THE MOSS Ceratodon purpureus BRID

Abstract— Protonemata of the moss Ceratodon purpureus cultured in white light were transferred to darkness for 3 days and then used for phototropic experiments. Irradiation of the apical region of vertically position protonemata with small beams (0.2 mm) of red light induced a growth response towards the irradiated side (positive phototropism). The phototropic response showed irradiance dependence. The effect of red light was completely reversed by far-red light following red light irradiations, demonstrating that phytochrome was the photoreceptor pigment. Far-red light or UV-blue light had no influence on either bulging or phototropism. Experiments with linearly polarized red or far-red light showed a different dichroic distribution of phytochrome in its different forms, the red-absorbing form, P_r and the far-red-absorbing form, P_fr. Red light with a vibration plane parallel to the long axis of the filaments was most effective. The effectiveness of far-red light was expressed best when its vibration plane was 90° to the electrical vector of the inductive red light.     

PHOTOPHYSIOLOGY OF TURION GERMINATION IN Spirodela polyrhiza (L.) SCHLEIDEN–V. DEMONSTRATION OF A CALCIUM-REQUIRING PHASE DURING PHYTOCHROME-MEDIATED GERMINATION

Abstract— Etiolated turions of Spirodela polyrhiza are positively photoblastic and show a phytochrome-mediated low fiuence germination response. The far-red light (FR) reversibility decreased with the delay of FR irradiation (lag phase 1.06 ± 0.03 days after red light irradiation; half-maximal response 1.9 days). The action of the far-red-absorbing form of phytochrome (Pfr) was only realized by a germination response if exogenously applied Ca²+ was present. Calcium step-down (from 1 mM to 0.9 μ M Ca²+) and Ca²+ step-up (from 0.9 μ M to 1 m M Ca²+) experiments were carried out to determine the Ca²+-sensitive phase. There was no time gap between the two phases determined by the step-down and step-up experiments but a clear coincidence of both curves. Pulse treatments (24 h) with Ca²+ (1 m M ) showed the upper part of this common curve to represent the most Ca²+-sensitive phase. The Ca²+-sensitive phase was within the Pfr-requiring phase. After reversion of Pfr by FR pulses there was only a negligible response to the high Ca²+-concentration, independent of the delay between the red light (R) and FR pulses. These results are compatible with the assumption of Ca²+ acting as a second messenger of Pfr. However, the Ca²+-insensitivity in the first 12 h after the R pulse points against this hypothesis.     

ADENOSINE 5-TRIPHOSPHATE CONTENT AND ENERGY CHARGE DURING PHOTOMORPHOGENESIS OF THE MUSTARD SEEDLING SIN APIS ALBA L.

Abstract— The contents of ATP, ADP and AMP (enzymatic assay) and the rate of respiration (output of CO₂ per h, continuously measured with an infrared CO₂ analyzer) were determined in long-term experiments with intact dark-grown and far-red-light-grown mustard ( Sinapis alba L.) seedlings. While there is a strong effect of far-red light treatment on the respiratory rate (inhibition as well as promotion, depending on the onset of light), an effect on the contents of adenosine nucleotides, and therewith on energy charge, is hardly detectable. At most, there is a tendency that long-term far-red light slightly lowers ATP levels and energy charge. The results suggest that the adenylate system (and therewith the energy charge) in the mustard seedling is controlled by endogenous homoeostasis even under intense phytochrome-mediated photomorphogenesis. It is unlikely that the levels of adenosine nucleotides or the energy charge are links in any causal chain originating from P_fr and leading to phenomena of photomorphogenesis.     

INDEPENDENT EFFECTS OF PHYTOCHROME AND NITRATE ON NITRATE REDUCTASE AND NITRITE REDUCTASE ACTIVITIES IN MAIZE

Abstract— Involvement of phytochrome in the regulation of nitrate reductase (NR) and nitrite reductase (NIR) activities in excised, etiolated leaves of Zea mays (L.) variety 'Ganga-5' is demonstrated using low energy and high irradiance responses of phytochrome action. Photoreversibility by far-red light of red light stimulated increases in NR and NIR activities was lost by 2 h. Red light given to the leaves, when induction by NO^-₃, was saturated, further increased both enzyme activities. Even if red light was given 4–8 h before NO^-₃, it still increased both NR and NIR activities.     

THE PHOTOBIOLOGY OF Paphiopedilum STOMATA: OPENING UNDER BLUE BUT NOT RED LIGHT

Abstract— The responses of stomata from Paphiopedilum harrisianum , Orchidaceae, to light and CO₂ were studied in epidermal peels. Stomatal opening under red light was indistinguishable from that in darkness, whereas blue light promoted opening above dark levels. The ineffectiveness of red light in causing stomatal opening was confirmed in the presence of 100 μ M KCN; average apertures in both darkness and red light were 53% of those measured in the absence of the inhibitor, whereas under blue irradiation, the KCN inhibition was only 30%, with average apertures two-fold of those measured under red light or darkness. Fluence rate response curves under blue light were typical of a single photoreceptor; removal of CO₂ increased aperture values without a significant light-CO₂ interaction. The lack of a stomatal red light response contrasts with results obtained in species with chlorophyllous stomata in which red light consistently causes stomatal opening, and suggests that the previously reported red light responses in stomata from intact Paphiopedilum leaves resulted from indirect effects, such as depletion of intercellular CO₂ by mesophyll photosynthesis. In isolation, Paphiopedilum stomata appear to rely on a blue light photosystem for their responses to light and fail to open under red light because of their lack of guard cell chloroplasts.     

ANALYSIS OF MULTIPLE PHOTORECEPTOR PIGMENTS FOR PHOTOTROPISM IN A MUTANT OF Arabidopsis thaliana

Abstract
The shape of the fluence-response relationship for the phototropic response of the JK224 strain of Arabidopsis thaliana depends on the fluence rate and wavelength of the actinic light. At low fluence rate (0.1 μmol m^-2s^-1), the response to 450-nm light is characterized by a single maximum at about 9 μmol m^-2. At higher fluence rate (0.4 μmol m^-2s^-1), the response shows two maxima, at 4.5 and 9 μmol m^-2. The response to 510-nm light shows a single maximum at 4.5 μmol m^-2. Unilateral preirradiation with high fluence rate (25 μmol m^-2s^-1) 510-nm light eliminates the maximum at 4.5 μmol m^-2 in the fluence response curve to a subsequent unilateral 450-nm irradiation, while the second maximum at 9 μmol m^-2 is unaffected. Based on these results, it is concluded that a single photoreceptor pigment has been altered in the JK224 strain of Arabidopsis thaliana.     

SEASONAL VARIATION OF SOLAR UV-RADIATION AT A HIGH MOUNTAIN STATION

Abstract— At the high mountain station Jungfraujoch (3576 m), the maximum daily totals for erythemal dose (G_ER), UV-A radiation (G_UVA) and global radiation (G) are 29 Sunburn Units d⁻¹, l.7 MJ m⁻²d⁻¹ and 37 MJ m⁻² d⁻¹. The maximum instantaneous values at solar noon in midsummer are 4.2 Sunburn Unit h⁻¹, 53 W m⁻² and 1110 W m⁻². A significantly nonlinear relation between G_ER and G results from the influence of the irradiated ozone mass on the UV-B erythemal dose. In contrast, G_UVA and G are linearly proportional, which can be seen from the diurnal and seasonal courses of the ratios G_ER/G and G_UVA/G AND from their dependence on the optical air mass. UV-A radiation flux is less attenuated by cloudiness than is global radiation. This effect is masked for the erythemal dose by variations in the ozone concentration. Due to seasonal ozone layer thickness and effective pathlength variations, the ratio G_ER/G shows a significant asymmetry. At the autumn equinox it is about 16% higher than at the spring equinox.     

PHOTOBIOLOGIE DU MÉSOCOTYLE D''AVOINE-III. DOUBLE EFFET DE LA LUMIÈRE BLEUE

Abstract— Using excise sections of oat first-internodes, a dual effect of blue light can be demonstrated on elongation when the sections are first irradiated in distilled H₂O, then incubated with gibberellic acid (GA). At low light energies (230 ergs/cm² per sec, for 2 min), a pretreatment with blue light enhances the GA effect above the elongation it can produce in the dark. At high energies (650 ergs/cm²/sec for 45 min), the same wavelengths cause an inhibition of the GA-induced elongation. An action spectrum for the two effects show a maximum near 435 mμ in both cases. Neither light effect is visible when indoyl-3-acetic acid is used instead of GA. Several physiological effects distinguish the two blue effects. The promotive effect is most marked in the young regions of the mesocotyl, whereas the maximum inhibitive effect is located in slightly older tissues. Time-course experiments showed that the promotive effect is partly due to an extension of the duration of elongation. The inhibitory effect is only temporary and vanishes about 30 hr after the beginning of the experiment. The promotive effect of blue light resembles the effect of far-red light, but the former can be observed with gibberellins A₂, A₄, A₅, A₆ and A₇ which are practically inactive after an irradiation with far-red light. The inhibitory effect of blue light is different from the red-light effect as shown by the time-course experiments.     

Cyclic-AMP levels in the lichen Evernia prunastri are modulated by light quantity and quality

Changes in the accumulation of cAMP levels were measured by the isotope dilution assay using protein kinase A in the lichen Evernia prunastri at varying light conditions. cAMP levels decreased following exposure to low irradiance (20 μmol quanta m⁻² s⁻¹, and below the compensation point for photosynthesis) of red light (600–710-nm wave length) and increased by 50% after far-red light illumination (15 μmol quanta m⁻² s⁻¹, 710–800-nm wavelength). Far-red partially reverted the effect of red light when the former was supplied after the latter. cAMP increased to its maximum level under high irradiance supplied by a non-photomorphogenic yellow light source (400 μmol quanta m⁻² s⁻¹, reaching photosynthetic saturation). The addition of small quantities of red and far-red light, however, had profound restricting effects on cAMP accumulation. The addition of inhibitors of electron transport chains did not promote any significant change in cAMP levels in any of the treatments, indicating that cAMP accumulation could not depend on ATP synthesis. We propose that the response of cAMP accumulation at low irradiance comprises the activation of a morphogenic pathway through a red/far-red photoreceptor. In addition, at high irradiance the response would occur most likely through photosystems II and I acting as sensors of light quantity, that can be strongly modified by the red/far-red photomorphogenic system. Thus, cAMP would be involved in sensing the overall light environment.     

A POSSIBLE CONTROL BY A PHYTOCHROME-LIKE PHOTORECEPTOR OF CHLOROPHYLL SYNTHESIS IN THE GREEN ALGA Ulva rigida

Chlorophyll synthesis is stimulated by red light pulses in the green alga Ulva rigida C. Aghard. Chlorophyll synthesis in darkness is greater after longer red light pulses (30 min) than after shorter red light pulses (5 min). Chlorophyll synthesis was higher after red light pulses of 14 Wm_-2 fluence rate than after those of 7 Wm_-2. The effect of red light showed some far-red reversibility. The reversion by far-red light was higher after red light pulses of 4 min than after those of 30 min. These results indicate the existence of a rapid induction of chlorophyll synthesis during the red light pulses and a fast escape from photoreversibility. The percentage of reversion is also affected by the fluence rate of the light pulses. The reversion was reduced by about 15% when the photon fluence rate was increased from 7 to 14 Wm_-2. Reversion was also observed when red and far-red light pulses were applied successively. Thus, phytochrome or a phytochrome-like photoreceptor could be involved in the induction of chlorophyll synthesis in Ulva rigida.     

PHOTOFRAGMENTATION OF α-OXOCARBOXYLIC ACIDS IN AQUEOUS SOLUTION. AN EPR STUDY. FORMATION OF SEMIDIONE RADICALS BY DECARBOXYLATIVE SUBSTITUTION OF α-OXOCARBOXYLIC ACIDS BY ACYL RADICALS-I: GLYOXYLIC, PYRUVIC, α-OXOBUTYRIC, α-OXOGLUTARIC AND α-OXOISOCAPROlC ACIDS

Abstract— By means of in situ photolysis EPR of aqueous solutions of α-oxocarboxylic acids (RCO-CO₂H) at pH values above 5, semidione radical anions [RC(O^-)=C(O')R] and α-hydroxy-α-carboxy alkyl radicals [RC(OH)CO₂-] were detected. C0₂ was identified as a reaction product. On photolysis of mixtures of α-oxocarboxylic acids (RCOCO₂H and R'COCC₂H), "mixed" semidione radical anions [RC(O^->=C(O)R'] were observed in addition to RC(O^-)=C(O')R, R'C(O^-)=C(O')R', RC(OH)CO₂^- and R'C(OH)CO₂^-. The experimental results are explained in terms of photodecarboxylation (α-clea-vage) of electronically excited RCOCOJ to yield RCO and CO₂. The radicals RC(OH)CO₂^- are formed by reduction of RCOCO₂^- by CO₂^-. The semidione radicals are produced by addition of RCO to RCOCO₂^- followed by decarboxylation of the intermediate adduct. This mechanism was confirmed by generating acyl radicals independently and reacting them with α-oxocarboxylic acids. Selected product studies support the mechanism suggested.