INTERACTION BETWEEN PHYTOCHROME AND THE BLUE LIGHT PHOTORECEPTOR SYSTEM IN Mougeotia*

Abstract— Face-to-profile chloroplast movement in Mougeotia was induced by sequences of strong blue and red short irradiations. This type of response occured only when blue light was applied prior to or simultaneously with red light, and far-red irradiation was necessary after the sequence to cancel the remaining gradient of the far-red absorbing form of phytochrome P_fr. The dependence of the response magnitude on blue and red light sequences was studied for a wide range of light durations and dark intervals. The relationship between the response and the dark interval points to the lack of direct coupling between phytochrome and blue-absorbing “cryptochrome”. It was postulated that a photoproduct having a life-time of2–3 min is formed by the blue-light-mediated reaction. This photoproduct interacts with phytochrome during its transformation or with its final P_fr form.     

PHYTOCHROME MEDIATED CAROTENOID SYNTHESIS IN THE FUNGUS VERTICILLIUM AGARZCZNUM

Abstract— Ten minutes of red irradiation (R) increased carotenogenesis in Verticillium agoricinum and this effect was reversed by 10min of far-red (FR) irradiation indicating that phytochrome is involved. A far-red minus red difference spectrum of a crude extract shows a peak at 670 nm and a dip at 750 nm wavelength, values slightly larger than higher plant phytochrome. indicating the presence of phytochrome.     

PHYTOCHROME ACTION: A REAPPRAISAL

Stems of fully green plants show at least two types of response to light. In one, P_fr inhibits elongation. The second is a promotion of elongation which operates only in light; the effectiveness of red and far-red wavelengths indicates that this response is also mediated through phytochrome. Studies of the de-etiolation process also provide evidence for two modes of action of phytochrome; one is a P_fr-dependent reaction, and the second requires continuous light (or frequent short irradiations). It is proposed that, in addition to reactions which require P_fr and proceed in darkness, an important reaction of phytochrome in green plants occurs only in light. We have termed these two modes of action of phytochrome “static” and “dynamic”. The static mode operates after a brief exposure to light which establishes P_fr; the potential responses are largely reversible by far-red and exhibit reciprocity. The dynamic mode operates only in light and the responses do not show reciprocity. We consider that this mode operates through the transition from one bound form of phytochrome to another. The possible involvement of these two modes of action of phytochrome in photoperiodic mechanisms is discussed.     

PHYTOCHROME-MEDIATED PHOTOTROPISM AND DIFFERENT DICHROIC ORIENTATION OF Pr AND Pfr IN PROTONEMATA OF THE FERN ADIANTUM CAPILLUS-VENERIS L.

Abstract— Single-celled protonemata of Adiantum capillus-veneris were cultured under continuous red light for 6 days and then in the dark for 15 h. Brief local exposure of a flank (5 times 20 /mi) of the subapical region of a protonema to a microbeam of red light effectively induced a phototropic response toward the irradiated side. The degree of the response was dependent upon the fluence of the red light. Red/far-red reversibility was typically observed in this photoreaction, showing that phytochrome was the photo-receptive pigment. When the flank was irradiated with a microbeam of linearly polarized red and far-red light, red light with an electrical vector parallel to the cell surface was most effective. However, the far-red light effect was most prominent when its electrical vector was normal to the cell surface. These polarized light effects indicate the different dichroic orientation of P_r (red-light-absorbing form of phytochrome) and P_r (far-red-light-absorbing form of phytochrome) at the cell flank.     

SPECTRAL CHARACTERIZATION OF HIGH-MOLECULAR-WEIGHT PHYTOCHROME*

Abstract— Previous information about the spectral and photochemical properties of phytochrome in vitro has apparently been determined in large part with chromopeptides derived from the native molecule by proteolysis. Characterization of high-mol-wt phytochrome in vitro has led to the observation that the far-red-absorbing form (Pfr) may undergo relatively large and reversible changes in far-red extinction. Phytochrome preparations which exhibit reduced far-red extinction as Pfr also exhibit a rapid reappearance of red absorbance after discontinuing the red illumination used to establish photostationary equilibrium. This rapid change in the red spectral region is not accompanied by any equivalent absorbance change in the far-red. The molecular basis for these newly reported spectral properties is not known. However, both properties may be eliminated by the addition of either 2-mercaptoethanol or ethylenediaminetetraacetic acid.     

PHYTOCHROME CONTROL OF ITS OWN SYNTHESIS IN Sorghum vulgare AND Avena sativa

The accumulation of phytochrome in the dark was measured for Avena sativa seedlings after a white light pretreatment and for Sorghum vulgare seedlings after continuous red or far-red light treatments, using the herbicide Norflurazon to prevent greening under continuous irradiation. In both cases the accumulation of phytochrome depends on the state of the phytochrome at the light-dark transition: high P_fr levels (red light pulse) led to a slower rate of phytochrome accumulation than lower P_fr levels (long wavelength far-red (RG 9) light pulse). Poly-(A⁺)-RNA was isolated fromA. sativa seedlings grown for 48 h in darkness + 24 h WL + light pulse (5 min) (red, RG 9 light, red followed by RG 9 light or RG 9 followed by red light pulse) + 19 h darkness. The poly-(A⁺)-RNA was translated in a rabbit reticulocyte lysate system and the translation products were immunoprecipitated by specific anti-phytochrome antibodies. It was demonstrated that the activity of mRNA coding for phytochrome was under phytochrome control.     

EVIDENCE FOR PHYTOCHROME CONTROLLED ENDOMITOSIS AND CELL ELONGATION IN PISUM SATIVUM EPICOTYLS

Elongation and endomitosis were studied in the epicotyl cortex cells of germinating seeds of Pisum sativum cv. Rondo. One min of red light per 24 h is sufficient to fully inhibit endomitosis. Terminal far-red irradiation can reverse the red effect to the level established by far-red light alone. This justifies the conclusion that phytochrome is involved in the regulation of endomitotic DNA replication. Since far-red light alone inhibits endomitosis to about 50%, we conclude that very low levels of Pfr are required to influence the endomitotic cycle.     

LIGHT-INDUCED LINEAR ACTION DICHROISM IN PHOTOREVERSIBLY PHOTOCHROMIC SENSOR PIGMENTS—I. THEORY

Abstract— With a photoreversibly photochromic regulator pigment such as phytochrome, linear action dichroism could theoretically be obtained after photoselection even if the molecules are initially randomly oriented: If randomly oriented P_fr (fed-absorbing phytochrome) molecules are partially converted to P_fr (far-red absorbing phytochrome) molecules by plane-polarized red light, those molecules will preferentially be converted which have their 'red' transition moments nearly parallel to the electric vector of the red light. The effect of subsequent plane-polarized far-red light will depend on the plane of polarization. A general theory is developed for how this can be used to determine whether or not the transition moment changes direction during conversion. The pigment need not be isolated, since only physiological reactions (such as germination or chromatic adaptation) are measured.     

EFFECT OF PHYTOCHROME ON THE BIOSYNTHESIS OF ACYCLIC AND CYCLIC CAROTENOIDS IN HIGHER PLANTS

Radish plants were grown in the presence of three different herbicides that interfere with the formation of the normal range of cyclic carotenoids, leading to an accumulation of acyclic biosynthetic intermediates, mainly phytoene (SAN 6706 and amitrole) and zeta-carotene (3852). Plants were then irradiated by four different light programs in order to gain more insight into the first steps of carotenoid biosynthesis and their control by light and phytochrome. In all cases, herbicide-treated and control, carotenoid biosynthesis was greatly enhanced by red light consistent with an effect of phytochrome on the early steps of the pathway. However, similar enhancement was also obtained after treatment with far-red light. Indeed with SAN 6706-treated plants synthesis of phytoene was stimulated to a much greater extent by far-red light given alone, than by red light. The involvement of phytochrome in the regulation of carotenoid biosynthesis appears not to be as simple as previously supposed.     

LIGHT-GROWN PLANTS OF THE CUCUMBER LONG HYPOCOTYL MUTANT EXHIBIT BOTH LONG-TERM AND RAPID ELONGATION GROWTH RESPONSES TO IRRADIATION WITH SUPPLEMENTARY FAR-RED LIGHT

Seedlings of wild type (WT) and the lh mutant of Cucumis sativus were grown in white light and given supplementary far-red radiation either to the whole plant, or via fiber-optic probes directed at the apical region of the hypocotyl. In both WT and mutant seedlings, reducing the ratio of red to far-red radiation resulted in significant increases in extension growth. Direct measurement of extension rate by transducer techniques revealed that the mutant seedlings responded to additional far-red radiation by an increment in extension rate that was equivalent to that observed with the WT seedlings. Assuming that the lh mutant is deficient in phytochrome B (Kendrick and Nagatani, The Plant Journal 1: 133–139, 1991), these results indicate that although phytochrome B clearly has an important role in the induction of the shade avoidance syndrome, it is unlikely to be the sole responsible receptor.     

KINETICS OF PHYTOCHROME PELLETABILITY*

Abstract— The pelletability of P_r from maize coleoptiles was studied as a function of the delay time between a red and a far-red light pulse given in vivo. The obtained curve can be resolved into three parts. The two slowest reactions have half lives of 40 s and 3.6 min at 0°C. Furthermore, a break in the Arrhenius plot from the slowest reaction of the curve indicates that either the phytochrome “receptor” or the phytochrome molecule itself undergoes a jump in the Arrhenius activation energy at 20°C. These data are in good agreement with kinetic studies of phytochrome pelletability also discussed in this paper.     

THE FUNCTION OF PHYTOCHROME IN THE NATURAL ENVIRONMENT—I. CHARACTERIZATION OF DAYLIGHT FOR STUDIES IN PHOTOMORPHOGENESIS AND PHOTOPERIODISM

Abstract— The daily changes in spectral energy distribution of natural daylight, with special reference to the red and far-red wayelength bands, are outlined. The effects of solar elevation and sky condition are described. The changes in the proportions of red and far-red radiation in daylight which occur at sunrise and sunset are discussed in relation to the possible function of phytochrome in photoperiodic timing.     

PHYTOCHROME-MEDIATED THRESHOLD CONTROL OF HYPOCOTYL GROWTH IN PARTLY DE-ETIOLATED MUSTARD (Sinapis alba L.) SEEDLINGS

Abstract— Hypocotyl elongation in etiolated mustard ( Sinapis alba L.) seedlings is known to be controlled by phytochrome (Pfr) through a threshold mechanism. The Pfr threshold value required to suppress hypocotyl growth was low (3 times 10⁻²% Pfr, based on total phytochrome in the hypocotyl at 36 h after sowing = 100%). In the present study the question was addressed whether the threshold control by Pfr of hypocotyl elongation also operates in light-pretreated, partly de-etiolated seedlings after transfer to darkness. The experimental results show that this is the case. Calculation of the threshold level in far-red light pretreated seedlings led to a very low value (3 times 10⁻⁷%) compared to etiolated seedlings (3 times 10⁻²%). In red light pretreated seedlings the threshold level was calculated to be 9 times 10⁻⁷%. Since the light pretreatment affected the rate of degradation of phytochrome strongly (half-life of Ptot in continuous red light was found to be 35 min in far-red pretreated instead of 47 min in etiolated material), the difference in threshold level between far-red and red pretreated material cannot be interpreted unambiguously. However, the conclusion can be drawn that light nretreatment strongly increases the degradation rate of Pfr and decreases the threshold level.     

Light regulation of cyclic-AMP levels in the red macroalga Porphyra leucosticta.

Total cyclic-3'-5'-adenosine monophosphate (cAMP) levels were measured in the gametophyte of the red macroalga Porphyra leucosticta under different light conditions in order to study its regulation by phytochrome or photosynthesis. cAMP levels were relatively low when samples were incubated in darkness, or exposed to red or far-red light. Irradiation with red+far-red light induced a moderate increase in cAMP levels, while white light induced a pronounced increase in cAMP levels. When incubated under increasing white light irradiance, cAMP levels closely followed the increase in photosynthetic oxygen evolution rate, suggesting a direct relationship between photosynthesis and cAMP accumulation. cAMP levels were not dependent on cellular ATP concentration, as inhibitors of ATP synthesis did not significantly affect cAMP levels in light. We conclude that cAMP depends on photosynthetic activity regardless of ATP synthesis and concentration or phytochrome activity.     

PHOTOREVERSIBLE CHANGES IN pH OF PEA PHYTOCHROME SOLUTIONS

Abstract— Photoinduced pH changes in unbuffered solutions of undegraded pea phytochrome were studied at 10_oC by using a glass electrode. Red light irradiation caused alkalinization of the solutions in the pH range 5.2–xs7.5 and acidification in pH 7.5–8.9. The pH changes were fully reversed by a subsequent irradiation with far-red light. The red and far-red light effects were repeatedly reversible. The solution of tryptic peptide of phytochrome (mol. wt 60000) showed similar photoreversible pH changes.     

REGULATION OF PROTEIN PHOSPHORYLATION BY PHYTOCHROME IN Sorghum bicolor

Abstract— In vitro phosphorylation of some polypeptides was affected in extracts obtained from 5-and 6-day-old plants irradiated with 5 min of red light. The phosphorylation of 55 kDa polypeptide in both 5- and 6-day-old plants, a 60 kDa, and 76 kDa polypeptide in 6-day-old plants and 70 kDa, 67 kDa polypeptide in 5-day-old plants was stimulated by red light. This effect was reversible by far-red light. The extent of stimulation by red light and reversal by far-red light varied for different polypeptides. No differential effect of red and far-red light was seen on the phosphorylation of 94 and 40 kDa polypeptides. In fact, phosphorylation of 94 kDa polypeptide in 6-day-old plants decreased on red light irradiation. These results show that the phosphorylation or dephosphorylation of some proteins is affected by phytochrome and the effect of light is also dependent on the age of the plant.     

Phytochrome-dependent photomovement responses mediated by phototropin family proteins in cryptogam plants

In this review, we describe the regulation of photomovement responses by phototropin and phytochrome photoreceptors. The blue light receptor phototropin mediates various photomovement responses such as phototropism, chloroplast movement and stomatal opening. In cryptogamic plants including ferns, mosses and green alga, red as well as blue light mediates phototropism and chloroplast movement. The red/far-red light reversibility suggests the involvement of phytochrome in these responses. Thereby, plant growth is presumably promoted by coordinating these photomovements to capture efficiently light for photosynthesis.     

Light Regulation of Phytochrome Content in Wild-type and Aphototropic Mutants of the Moss Ceratodon purpureus

In filaments of the moss Ceratodon purpureus , phototropism is controlled by the photoreceptor phytochrome. Thirty-three aphototropic mutants with a proposed defect in phytochrome-chromophore biosynthesis were isolated and analyzed. The phototropic response of those mutants was rescued with the precursor of the phytochrome chromophore, biliverdin. Phytochrome spectral activity was measured in 19 arbitrarily chosen mutants. All contained low but still measurable quantities of photoactive phytochrome; the highest level was around 15% of the wild-type. The level of total phytochrome (apophytochrome and holophytochrome) as assayed by immunoblotting was indistinguishable from wild-type. The content of photoactive phytochrome in Ceratodon is light-regulated. Phytochrome of wild-type kept for 24 h in red light was reduced to 50% as compared to dark controls but was unaffected by blue. The red-light-induced decrease was partially reversible by far-red light, indicating that phytochrome itself is the photoreceptor for this response. This regulation was further analyzed with the mutant ptr114 , which contains 15% photoactive phytochrome as compared to the wild-type. In this mutant, continuous red light given for 6 days decreased the level of spectrally active phytochrome down to 25% of dark controls, whereas the amount of phytochrome found on immunoblots was hardly reduced. This indicates that the loss of phytochrome affects only the holoprotein and implies that Ceratodon phytochrome is specifically degraded as a far-red-absorbing phytochrome.     

PHYTOCHROME INDUCTION OF PHOTOREACTIVATING ENZYME IN Phaseolus vulgaris L. SEEDLINGS*

Abstract— Photoreactivating enzyme (PRE) activity was measured in hypocotyls of Phaseolus vulgaris L. seedlings using a radioimmunoassay for thymine dimers. In dark-grown seedlings a five-fold increase in PRE activity was observed after 6 h of irradiation with blue or far-red light. Short time irradiations with red light were also effective. Reversibility of this red-light-effect by a subsequent short term irradiation with far-red light and also the high effectiveness of continuous far-red light indicate that PRE activity is under phytochrome control. This observation points to PRE induction via gene activation.     

PHYTOCHROME REGULATION OF STEM GROWTH AND INDOLE-3-ACETIC ACID LEVELS IN THE lv AND Lv GENOTYPES OF Pisum

Phytochrome influences stem elongation and the mechanism for this is not understood. The levels of indole-3-acetic acid (IAA) were analyzed in an leLv genotype of Pisum sativum L. which responded to end-of-day far-red light by doubling growth rate. The IAA levels in epidermal peels increased 40% after far-red light whereas IAA levels of the entire stem tissue changed insignificantly. This increase was reversible by red light. Under light-grown conditions, the lv mutation increases stem elongation rates by 2–3-fold and is thought to block the transduction of a phytochrome signal. Analysis of the short-term stem elongation kinetics of dark- and light-grown Lv and lv seedlings suggests that lv blocks the action of the light-stable form of phytochrome. The higher growth rate of lv plants was found to be associated with abnormally high epidermal IAA levels typical of far-red treated Lv plants. End-of-day far-red treatments did not substantially increase epidermal IAA levels in lv plants. These observations support the view that phytochrome regulation of stem elongation may occur in part through modulation of epidermal IAA levels. The lv mutation may result in increased internode growth in part by blocking the ability of phytochrome to decrease epidermal IAA levels.