Far-red Light Pulse Research Articles

Are NADP-dependent isocitrate dehydrogenases and ferredoxin-dependent glutamate synthase co-regulated by the same photoreceptors?

Activities of NADP-dependent isocitrate dehydrogenases (cytosolic and plastidic isoforms, ICDH1 and ICDH2; EC 1.1.1.42) and ferredoxin-dependent glutamate synthase (Fd-GOGAT; EC 1.4.7.1) in turions of Spirodela polyrhiza were all stimulated by light. Single or repeated red light (R) pulses induced the activity of the enzymes and this effect was reverted by subsequent far-red light (FR) pulses. The enzymes are, therefore, co-regulated by the low-fluence response of phytochrome. For ICDH, this is reported here for the first time. Neither an effect of the very low-fluence response nor of the FR-mediated high-irradiance response was detectable. Irradiance with continuous R resulted in enhanced enzyme activities and protein levels (Western analysis using polyclonal antibodies against ICDH1 and Fd-GOGAT). These additional effects of continuous R (called a "non-induction effect") could be inhibited for ICDH1 and ICDH2 by the inhibitor of photosynthetic electron transport, 3-(3',4'-dichlorophenyl)-1,1-dimethylurea, and are therefore related to the effect of photosynthesis. In contrast, the non-induction effect of Fd-GOGAT was resistant against this inhibitor. Moreover, hourly R pulses did not replace the effect of continuous R. The non-induction effect of light on the activity and protein level of Fd-GOGAT was therefore tentatively classified as an R-mediated high-irradiance response. The activity of Fd-GOGAT but not that of ICDHs was additionally regulated by a specific blue-light receptor. It can be concluded that the levels of ICDHs and Fd-GOGAT were coordinated by light but were not co-regulated by the same photoreceptors. Nitrate is necessary for the light regulation of both enzymes, contributing to the coordinated expression of the relevant genes.

Characterization of a novel non-constitutive photomorphogenic cop1 allele.

A specific light program consisting of multiple treatments with alternating red and far-red light pulses was used to isolate mutants in phytochrome A-dependent signal transduction in Arabidopsis seedlings. Because of their phenotype, the mutants were called eid (empfindlicher im dunkelroten Licht, which means hypersensitive in far-red light). One of the isolated mutants, eid6, is a novel recessive allele of the COP1 gene (constitutive photomorphogenic 1) that carries an amino acid transition in a conserved histidine residue of the RING finger domain. Mutant seedlings exhibited an extreme hypersensitivity towards all tested light qualities, but in contrast to known cop1 alleles, no constitutive photomorphogenic phenotype was detectable in darkness. Thus, the novel cop1eid6 allele seems to encode for a protein whose remaining activity is sufficient for the suppression of photomorphogenesis in dark-grown plants. In adult cop1eid6 plants, the development of the Cop1 phenotype is dominated by phytochrome B. Comparison of the phenotype of the novel cop1eid6 and the weak cop1-4 allele under continuous far-red light indicates that the RING finger and coiled-coil domains of COP1 are sufficient for some specific regulatory function in phytochrome A-dependent high irradiance responses.

The Cape Verde Islands allele of cryptochrome 2 enhances cotyledon unfolding in the absence of blue light in Arabidopsis.

We analyzed the natural genetic variation between Landsburg erecta (Ler) and Cape Verde Islands (Cvi) accessions by studying 105 recombinant inbred lines to search for players in the regulation of sensitivity to light signals perceived by phytochromes in etiolated seedlings of Arabidopsis. In seedlings grown under hourly pulses of far-red (FR) light, we identified three quantitative trait loci (QTLs; VLF3, VLF4, and VLF5) for hypocotyl growth inhibition and three different QTLs (VLF6, VLF7, and VLF1) for cotyledon unfolding. This indicates that different physiological outputs have selective regulation of sensitivity during de-etiolation. Ler alleles, compared with Cvi alleles, of VLF3, VLF4, VLF5, VLF7, and VLF1 enhanced, whereas the Ler allele of VLF6 reduced, the response to pulses of FR. We confirmed and narrowed down the position of some QTLs by using near-isogenic lines. VLF6 mapped close to the CRY2 (cryptochrome 2) gene. Transgenic Ler seedlings expressing the Cvi allele of CRY2 showed enhanced cotyledon unfolding under hourly pulses of FR compared with the wild type or transgenics expressing the CRY2-Ler allele. This response required phytochrome A. The cry1 cry2 double mutant lacking both cryptochromes showed reduced cotyledon unfolding under FR pulses. Because the CRY2-Cvi is a gain-of-function allele compared with CRY2-Ler, cryptochrome activity correlates positively with cotyledon unfolding under FR pulses. We conclude that the blue light photoreceptor cryptochrome 2 can modulate seedling photomorphogenesis in the absence of blue light. In addition to the nuclear loci, we identified cytoplasmic effects on seedling de-etiolation.

Light-regulated differential expression of pea chloroplast and cytosolic fructose-1,6-bisphosphatases.

The light-regulated differential expression of pea chloroplast and cytosolic fructose-1,6-bisphosphatases (FBPase) was investigated using enzyme activity assay, immunoblot, and Northern blot analyses. The enzyme activities of both chloroplast and cytosolic FBPases gradually increased under continuous white light illumination, although the increase in chloroplast FBPase was more drastic. Northern and immunoblot analyses also indicated that light stimulated the expression of both enzymes. Enzyme activity and the transcript levels of both enzymes gradually decreased under the dark treatment, although protein levels were unchanged for up to 24 h following the initiation of culture in the dark, indicating that reversible modifications of the enzymes may occur during the transition from light to dark (or the reverse). Light pulse experiments using blue (420 nm) and red/far-red (660/730 nm) light were carried out to analyze the photoreceptors related to the light-mediated expression of both enzymes. Expression of the chloroplast enzyme was very sensitive to red or far-red light pulses-it was induced by red light, but suppressed by far-red light pulses, as determined by enzyme activity, immunoblot, and Northern blot analyses, suggesting that red light signaling is involved in the control of chloroplast FBPase expression. However, cytosolic FBPase was virtually insensitive to blue or red/far-red light pulses in terms of enzyme activity, as determined by protein and transcript levels, indicating that cytosolic enzyme expression is not directly regulated by light signals. Instead, the expression of the cytosolic enzyme may be closely related to photosynthetic energy conversion accompanied by continuous white light irradiation.

Electron flow to photosystem I from stromal reductants in vivo: the size of the pool of stromal reductants controls the rate of electron donation to both rapidly and slowly reducing photosystem I units.

Electron donation from stromal reductants to photosystem I (PSI) was studied using the kinetics of P700(+) (the oxidized primary donor of PSI) reduction in the dark after irradiation of barley ( Hordeum vulgare L.) leaves. The leaves were treated with diuron and methyl viologen to abolish both the electron flow from PSII and PSI-driven cyclic electron transport. The redox state of P700 was monitored using the absorbance changes at 830 nm (Delta A(830)). Two exponentially decaying components with half-times of about 3 s (the slow component) and about 0.6 s (the fast one) were distinguished in the kinetic curves of Delta A(830) relaxation after a 1-s pulse of far-red light. The complex kinetics of P700(+) reduction thus manifested two types of PSI unit differing in the rate of electron input from stromal reductants. The rates of both kinetic components assayed after 1-s pulses were increased about 20-fold by a short (2-5 min) heat-pretreatment of leaves, indicating the accelerated input of electrons to both types of PSI unit. The increased rates of electron flow to P700(+) were even observed 1.5 h after the action of heat had been completed. Both kinetic components were dramatically slowed down upon irradiation of heat-treated leaves for 20-30 s. Their rates were restored after a short (20-30 s) period of darkness. A 5-min leaf exposure at 38 degrees C was sufficient to stimulate by severalfold the reduction of P700(+) pre-oxidized by a brief light pulse. In contrast, the acceleration of P700(+) reduction after a 1-min irradiation was observed only if leaves were subjected to temperatures above 40 degrees C. Neither heat treatment of leaves nor light-dark modulations in the rates of the fast and the slow components of P700(+) dark reduction influenced the relative magnitudes of the two kinetic components, providing strong additional evidence in favor of two distinct types of PSI existing per se in barley leaves. The key role in the control of the activity of electron donation to P700(+) in both rapidly and slowly reducing PSI units was attributed to the amount of stromal reductants available for P700(+) reduction. The latter was expected to be reduced under illumination in the presence of methyl viologen, while increased again in the dark. The regeneration of the pool of stromal reductants in the dark was likely provided by starch breakdown within the chloroplast stroma, but not by import of reducing equivalents from the cytosol. This was evidenced by much lower rates, compared with 1-h dark-adapted leaves, of dark reduction of both components of P700(+) in leaves stored for 24 h in the dark and thus depleted of starch but containing large amounts of glucose, the respiratory substrate.

SPA1, a component of phytochrome A signal transduction, regulates the light signaling current.

Mutations in a component of phytochrome A (phyA)-specific light signal transduction, SPA1, result in enhanced responsiveness of Arabidopsis seedlings to red and far-red light. Here, we have examined the effects of spa1 mutations on the two known modes of phyA function, the high-irradiance responses (HIRs) to continuous irradiation with far-red light and the very-low-fluence responses (VLFRs) to inductive pulses of light that establish only a small proportion of active phyA. spa1 mutants exhibited an enhanced VLFR under hourly pulses of far-red light for hypocotyl growth inhibition, cotyledon unfolding, anthocyanin accumulation, block of greening in subsequent white light and negative regulation of phyB signaling. We provide evidence that the phenotype of spa1 mutants in red light is also caused by an increase in the VLFR. Taken together, our results indicate that light-induced hypocotyl growth inhibition in spa1 mutants is primarily due to a VLFR. While wild-type seedlings required hourly pulses of far-red light to induce a VLFR, infrequent irradiation with far-red pulses (every 12 h) was sufficient to induce a strong VLFR of hypocotyl elongation in spa1 mutants. This shows that the effect of the VLFR was more persistent in spa1 mutants than in the wild type. We, therefore, propose that SPA1 has an important function in reducing the persistence of phyA signaling. spa1 mutations also enhanced the HIRs of anthocyanin accumulation and of phyA-mediated responsivity amplification towards phyB. Thus, our results suggest that spa1 mutations amplify both the phyA-mediated VLFR and the HIR.

Brassinosteroid Mutants Uncover Fine Tuning of Phytochrome Signaling

Phytochromes (phy) A and B provide higher plants the ability to perceive divergent light signals. phyB mediates red/far-red light reversible, low fluence responses (LFR). phyA mediates both very-low-fluence responses (VLFR), which saturate with single or infrequent light pulses of very low fluence, and high irradiance responses (HIR), which require sustained activation with far-red light. We investigated whether VLFR, LFR, and HIR are genetically coregulated. The Arabidopsis enhanced very-low-fluence response1 mutant, obtained in a novel screening under hourly far-red light pulses, showed enhanced VLFR of hypocotyl growth inhibition, cotyledon unfolding, blocking of greening, and anthocyanin synthesis. However, eve1 showed reduced LFR and HIR. eve1 was found allelic to the brassinosteroid biosynthesis mutant dim/dwarf1. The analysis of both the brassinosteroid mutant det2 in the Columbia background (where VLFR are repressed) and the phyA eve1 double mutant indicates that the negative effect of brassinosteroid mutations on LFR requires phyA signaling in the VLFR mode but not the expression of the VLFR. Under sunlight, hypocotyl growth of eve1 showed little difference with the wild type but failed to respond to canopy shadelight. We propose that the opposite regulation of VLFR versus LFR and HIR could be part of a context-dependent mechanism of adjustment of sensitivity to light signals.

Brassinosteroid Mutants Uncover Fine Tuning of Phytochrome Signaling

Phytochromes (phy) A and B provide higher plants the ability to perceive divergent light signals. phyB mediates red/far-red light reversible, low fluence responses (LFR). phyA mediates both very-low-fluence responses (VLFR), which saturate with single or infrequent light pulses of very low fluence, and high irradiance responses (HIR), which require sustained activation with far-red light. We investigated whether VLFR, LFR, and HIR are genetically coregulated. The Arabidopsis enhanced very-low-fluence response1mutant, obtained in a novel screening under hourly far-red light pulses, showed enhanced VLFR of hypocotyl growth inhibition, cotyledon unfolding, blocking of greening, and anthocyanin synthesis. However,eve1 showed reduced LFR and HIR. eve1 was found allelic to the brassinosteroid biosynthesis mutantdim/dwarf1. The analysis of both the brassinosteroid mutant det2 in the Columbia background (where VLFR are repressed) and the phyA eve1 double mutant indicates that the negative effect of brassinosteroid mutations on LFR requires phyA signaling in the VLFR mode but not the expression of the VLFR. Under sunlight, hypocotyl growth of eve1 showed little difference with the wild type but failed to respond to canopy shadelight. We propose that the opposite regulation of VLFR versus LFR and HIR could be part of a context-dependent mechanism of adjustment of sensitivity to light signals.

Light-induced growth promotion by SPA1 counteracts phytochrome-mediated growth inhibition during de-etiolation.

Previous evidence has suggested that SPA1 is a signal transduction component that appears to require phytochrome A for function in seedling photomorphogenesis. Using digital image analysis, we examined the time course of growth inhibition induced by red light in spa1 mutants to test the interpretation that SPA1 functions early in a phyA-specific signaling pathway. By comparing wild-type and mutant responses, we found that SPA1 caused an increase in hypocotyl growth rate after approximately 2 h of continuous red light, whereas the onset of phyA-mediated inhibition was detected within several minutes. Thus, SPA1-dependent growth promotion began after phyA started to inhibit growth. The action of SPA1 persisted for approximately 2 d of red light, a period well beyond the time when the phyA photoreceptor and its influence on growth have both decayed to undetectable levels. Also, SPA1 promoted growth for many hours in the complete absence of a light stimulus when red-light-grown seedlings were shifted to darkness. We propose that SPA1 functions in a light-induced mechanism that promotes growth and thereby counteracts growth inhibition mediated by phyA and phyB. Our finding that spa1 seedlings do not display growth promotion in response to end-of-day pulses of far-red light, even in a phyA-null background, supports this interpretation. Combined, these results lead us to the view that the rate of hypocotyl elongation in light is determined by at least two independent, opposing processes; an inhibition of growth by the phytochromes and a promotion of growth by light-activated SPA1.

Light-induced starch degradation in non-dormant turions of Spirodela polyrhiza.

Continuous red light controls starch degradation in turions of Spirodela polyrhiza [Dölger, K., U. K. Tirlapur and K.-J. Appenroth [1997] Photochem. Photobiol. 66, 126-127 (1997)]. This light could be replaced by repeated red light pulses with the reciprocity law fulfilled over a large range of fluence rates. The effect of red light pulses repeated every 24 or 12 h for 6 days was reversible by subsequent far-red light pulses. In contrast, hourly applied red pulses were irreversible by far-red light. This discrepancy was explained by showing the starch degradation activity of far-red pulses themselves. The investigated process was categorized as a phytochrome low fluence response with an unusual property: requirement of light treatment for several days. A partial fulfillment of this requirement was obtained with a red pulse followed by a dark period and a 24 h continuous irradiation. These results suggest the existence of two separate steps in the process of starch degradation in turions: formation of a sprout (= sink) during the pulse-induced germination, and starch degradation in the storage tissue (= source) induced by the second light treatment.

Antagonistic action of low-fluence and high-irradiance modes of response of phytochrome on germination and beta-mannanase activity in Datura ferox seeds.

Seed germination is often induced by a pulse of red light perceived by phytochrome and cancelled by a subsequent pulse of far-red light. When the pulse of red light is followed by several hours of darkness, a pulse of far-red light is no longer effective and prolonged far-red is necessary to block germination. The aim was to investigate whether the red light pulse and prolonged far-red light act on the same or different processes during germination of Datura ferox seeds. Forty-five hours after the inductive red light pulse, germination could not be blocked by one pulse or six hourly pulses of far-red light, but was significantly reduced by 6 h of continuous far-red light. The pulse of red light increased embryo growth potential and the activities of beta-mannanase and beta-mannosidase extracted from the micropylar region of the endosperm. Continuous far-red light had no effect on embryo growth potential or beta-mannosidase activity, but severely reduced the activity of beta-mannanase. The effect of far-red light had the features of a high-irradiance response of phytochrome. Both germination and beta-mannanase activity were restored by a pulse of red light given after the end of the continuous far-red treatment. It is concluded that the low-fluence response and the high-irradiance response modes of phytochrome have antagonistic effects on seed germination and that the control of beta-mannanase activity is one process where this antagonism is established.

Fhy3-1 Retains Inductive Responses of Phytochrome A

The fhy3 mutation of Arabidopsis impairs phytochrome A (phyA)-mediated inhibition of hypocotyl growth without affecting the levels of phyA measured spectrophotometrically or immunochemically. We investigated whether the fhy3-1 mutation has similar effects on very low fluence responses (VLFR) and high irradiance responses (HIR) of phyA. When exposed to hourly pulses of far-red light, etiolated seedlings of the wild type or of the fhy3-1 mutant showed similar inhibition of hypocotyl growth, unfolding of the cotyledons, anthocyanin synthesis, and greening upon transfer to white light. In the wild type, continuous far-red light was significantly more effective than hourly far-red pulses (at equal total fluence). In the fhy3-1 mutant, hourly pulses were as effective as continuous far-red light, i.e. the failure of reciprocity typical of HIR was not observed. Germination was similarly promoted by continuous or pulsed far-red in wild-type and fhy3-1 seeds. Thus, for hypocotyl growth, cotyledon unfolding, greening, and seed germination, the fhy3-1 mutant retains VLFR but is severely impaired in HIR. These data are consistent with the idea that VLFR and HIR involve divergent signaling pathways of phyA.

A new type of mutation in the plant photoreceptor phytochrome B causes loss of photoreversibility and an extremely enhanced light sensitivity.

Photoreversibility, reversion of the inductive effect of a brief red light pulse by a subsequent far-red light pulse, is a property of photoresponses regulated by the plant photoreceptor phytochrome B (phyB). We screened for mutants with impaired photoreversibility to gain better insight into the phyB-specific signalling cascade. The phenotype of the mutant described is caused by a single amino acid exchange in a phyB subdomain that is highly conserved in all phytochromes but whose functional significance was unknown. The mutated phyB showed a slower dark reversion but no major alterations in its spectral properties. In addition to its loss of photoreversibility, the mutant also exhibited a hypersensitivity towards continuous red-light irradiation and an altered phenotype of adult plants under short-day conditions.

Light-induced nuclear import of phytochrome-A:GFP fusion proteins is differentially regulated in transgenic tobacco and Arabidopsis.

Phytochromes (phy) are a family of photoreceptors that control various aspects of light-dependent plant development. Phytochrome A (phyA) is responsible for the very low fluence response (VLFR) under inductive light conditions and for the high irradiance response (HIR) under continuous far-red light. We have recently shown that nuclear import of rice phyA:GFP is regulated by VLFR in transgenic tobacco. The import is preceded by very fast, light-induced formation of sequestered areas of phyA:GFP in the cytosol. Here we report that expression of the Arabidopsis phyA:GFP fusion protein in phyA-deficient Arabidopsis plants complements the mutant phenotype. In these transgenic Arabidopsis lines, both light-dependent cytosolic formation of sequestered areas of the phyA:GFP as well as VLFR or HIR-mediated nuclear import of the fusion protein was observed. By contrast, light-dependent nuclear import of the same fusion protein was induced only by continuous far-red light (HIR) but not by pulses of far-red light (VLFR) in transgenic tobacco. These results demonstrate that photoregulation of intracellular partitioning of the Arabidopsis phyA:GFP differs significantly in different genetic backgrounds.

Eid1: A New Arabidopsis Mutant Hypersensitive in Phytochrome A–Dependent High-Irradiance Responses

To identify specific mutants for components of phytochrome A (phyA) signaling in Arabidopsis, we established a light program consisting of multiple treatments with alternating red and far-red light. In wild-type seedlings, irradiation with multiple red light pulses can reduce the amount of phyA, which in turn decreases the high-irradiance responses (HIRs) mediated by the subsequent treatments with far-red light. Our mutants were able to avoid this red light-dependent reduction of the HIR. Here, we describe eid1, a new recessive mutant with increased sensitivity to far-red light. The eid1 mutation maps to the top of chromosome 4. The mutants showed no change in phenotype in darkness or under continuous white light, but they exhibited an increased sensitivity to red light and an increased persistence of HIR during prolonged dark phases after multiple short pulses of far-red light. The eid1 seedlings accumulated normal amounts of phytochrome and showed no alterations in the degradation or de novo synthesis of phyA. The expression of the Eid1 phenotype requires the presence of phyA. Our data provide evidence that EID1 is a negatively acting component in the phyA-dependent HIR-signaling pathway.

Photocontrol of subcellular partitioning of phytochrome-B:GFP fusion protein in tobacco seedlings.

Photomorphogenesis of higher plants is regulated by photoreceptors including the red/far-red light-absorbing phytochromes, blue-UV/A sensing cryptochromes and as yet uncharacterized UV/B receptors. Specific phototransduction pathways that are controlled by either individual or interacting photoreceptors mediate regulation. Phytochrome B (phyB) is the major red light-sensing photoreceptor. Phototransduction mediated by this light sensor has been shown to include light-dependent nuclear import and interaction of phyB with transcription factor-like proteins in the nucleus. Here we report that nuclear import of phyB and physiological responses regulated by this photoreceptor exhibit very similar wavelength- and fluence rate-dependence. Nuclear import of phyB is insensitive to single red, blue and far-red light pulses. It is induced by continuous red light and to a lesser extent by continuous blue light, whereas far-red light is completely ineffective. The data presented indicate that light-dependent partitioning of phyB exhibits features characteristic of blue light responsiveness amplification, a phenomenon that is thought to be mediated by interaction of phyB with CRY1.

Nuclear and cytosolic events of light-induced, phytochrome-regulated signaling in higher plants.

Plants as sessile organisms cannot walk away but have to adapt to changes in their natural environments. Light is obviously the most important external factor, being the energy source. To regulate their fitness, the photosynthetic capacity of plants must be adapted to changes in the ambient light environment. Therefore, plants have evolved a large set of photoreceptors to monitor light quality and quantity, ranging from the UV to the infrared part of the spectrum. At least three different photoreceptor classes have been identified and analyzed: (i) UV‐B receptors, primarily characterized by action spectroscopy (Wellmann, 1983); (ii) the blue UV‐A photoreceptors, cry1 and cry2 and phototropine (Ahmad and Cashmore, 1993; Christie et al ., 1998; Lin et al ., 1998); and (iii) the red/far‐red reversible phytochromes. These photoreceptors control different aspects of plant growth and development, and in this review we focus on the physiological and molecular events regulated by the most characterized plant photoreceptors, the phytochromes. In higher plants, phytochromes are encoded by a small multigene family; five genes ( PHYA–PHYE , Sharrock and Quail, 1989; Clack et al ., 1994) have been identified in Arabidopsis . Phytochromes exist as dimers composed of two 125 kDa polypeptides, each carrying a covalently linked tetrapyrrol chromophore in the N‐terminal domain and dimerization domains in the C‐terminal domain. The photosensory function of the molecule is based on its capacity for reversible interconversion between the red light‐absorbing Pr form and the far‐red light absorbing Pfr form following sequential absorption of red and far‐red light. Photosignal perception by the receptor is followed by conformational changes, and activates, through an as yet poorly understood mechanism, signaling pathways leading to changes in the expression of genes that underlie developmental responses to light (Quail et al ., 1995). In order to finely tune their …

Elementary processes of photoperception by phytochrome A for high-irradiance response of hypocotyl elongation in Arabidopsis.

Elementary processes of photoperception by phytochrome A (PhyA) for the high-irradiance response (HIR) of hypocotyl elongation in Arabidopsis were examined using a newly designed irradiator with LED. The effect of continuous irradiation with far-red (FR) light could be replaced by intermittent irradiation with FR light pulses if given at intervals of 3 min or less for 24 h. In this response, the Bunsen-Roscoe law of reciprocity held in each FR light pulse. Therefore, we determined the action spectrum for the response by intermittent irradiation using phyB and phyAphyB double mutants. The resultant action spectrum correlated well with the absorption spectrum of PhyA in far-red-absorbing phytochrome (Pfr). Intermittent irradiation with 550 to 667 nm of light alone had no significant effect on the response. In contrast, intermittent irradiation with red light immediately after each FR light pulse completely reversed the effect of FR light in each cycle. The results indicate that neither red-absorbing phytochrome synthesized in darkness nor photoconverted Pfr are physiologically active, and that a short-lived signal is induced during photoconversion from Pfr to red-absorbing phytochrome. The mode of photoperception by PhyA for HIR is essentially different from that by PhyA for very-low-fluence responses and phytochrome B for low-fluence responses.

Light-dependent regulation of cyanobacterial phytochrome expression.

A histidine kinase protein (Cph1) with sequence homology and spectral characteristics very similar to those of the plant phytochrome has been recently identified in the cyanobacterium Synechocystis sp. strain PCC 6803. Cph1 together with Rcp1 (a protein homologue to the response regulator CheY) forms a light-regulated two-component system whose function is presently unknown. Levels of cph1 rcp1 mRNA increase in the dark and decrease upon reillumination. A dark-mediated increase in cph1 rcp1 mRNA levels was inhibited by the presence of glucose, but not by inhibition of the photosynthetic electron flow. The half-life of cph1 rcp1 transcript in the light was about fourfold shorter than in the dark, indicating that control of cph1 rcp1 transcript stability is one of the mechanisms by which light regulates expression of the cyanobacterial phytochrome. After 15 min of darkness, 3-min pulses of red, blue, green, and far-red light were equally efficient in decreasing the cph1 rcp1 mRNA levels. Red light downregulation was not reversed by far-red light, suggesting that cph1 rcp1 mRNA levels are not controlled by a phytochrome-like photoreceptor. Furthermore, a Synechocystis strain containing an H538R Cph1 point mutation, unable to phosphorylate Rcp1, shows normal light-dark regulation of the cph1 rcp1 transcript levels. Our data suggest a role of cyanobacterial phytochrome in the control of processes required for adaptation in light-dark and dark-light transitions.

The effect of light on the level of acetylcholine in seedlings of the wild-type and phytochrome mutants of tomato (Lycopersicon esculentum Mill.)

Applying the method of pyrolysis coupled with gas chromatography (PYR-GC) the content of endogenous acetylcholine (ACh) was investigated in the extracts obtained from tomato (Lycopersicon esculentum Mill.). Seven-day-old seedlings of wild type (WT) and phytochrome mutants au (aurea), hp (high pigment), fri (far-red light insensitive) and tri (temporarily red light insensitive) were studied. In the analyzed material the presence of choline and acetylcholine was discovered. The highest content of ACh (381 mmole/g of fresh weight) was found in tomato cotyledons, whereas the lowest amount (162 nmole/g of fresh weight) in roots. The level of ACh in the plants grown under the continuous light was higher than in etiolated ones. However, no considerable differences in the concentrations of ACh in au and tri seedlings grown under the continuous light and in darkness were observed. The irradiation of etiolated seedlings of wild type with red light was accompanied by the increase of endogenous level of ACh. The pulse of far-red light applied directly after red light reversed this stimulating effect. A similar effect of both light wavelengths on the content of ACh was also found in the case of the tri mutant. On the other hand, in the case of fri mutant, pulse of red light caused the drop in the content of ACh, whereas far-red applied after red light caused visible increase in the level of the investigated substance. In tissues of au mutant no effect of red and far-red lights on the concentration of ACh was established.