Phytochrome Synthesis Research Articles

PE-1, Encoding Heme Oxygenase 1, Impacts Heading Date and Chloroplast Development in Rice ( Oryza sativa L.).

The duration of the rice growth phase has always been an important target trait. The identification of mutations in rice that alter these processes and result in a shorter growth phase could have potential benefits for crop production. In this study, we isolated an early aging rice mutant, pe-1, with light green leaves, using γ-mutated indica rice cultivar and subsequent screening methods, which is known as the phytochrome synthesis factor Se5 that controls rice flowering. The pe-1 plant is accompanied by a decreased chlorophyll content, an enhanced photosynthesis, and a decreased pollen fertility. PE-1, a close homologue of HY1, is localized in the chloroplast. Expression pattern analysis indicated that PE-1 was mainly expressed in roots, stems, leaves, leaf sheaths, and young panicles. The knockout of PE-1 using the CRISPR/Cas9 system decreased the chlorophyll content and downregulated the expression of PE-1-related genes. Furthermore, the chloroplasts of pe-1 were filled with many large-sized starch grains, and the number of osmiophilic granules (a chloroplast lipid reservoir) was significantly decreased. Altogether, our findings suggest that PE-1 functions as a master regulator to mediate in chlorophyll biosynthesis and photosynthetic pathways.

Lack of influence of low temperature, light and growth substances on phytochrome resynthesis in coleoptiles of irradiated oat seedlings

The photoconversion of phytochrome P<sub>R</sub> into the P<sub>FR</sub> form causes at the same time the destruction of the initial large fraction of phytochrome found in the coleoptiles of etiolated oat seedlings. Factors such as low temperature, light of different wavelengths or growth substances are not capable of preventing the progressive destruction and restore the synthesis of phytochrome. Thus an abnormally high level of phytochrome is found only in etiolated seedlings. Such seedlings, on the other hand, are characterized by a very high rate of elongation growth. The role of phytochrome in the control of deetiolation of seedlings is discussed.

In vitro evaluation of mitochondrial–chloroplast subcellular localization of heme oxygenase1 (HO1) in Glycine max

Heme oxygenase1 (HO1) catalyzes the degradation of heme in to biliverdin, carbon monoxide, and ferrous ions. Its role in higher plants has been found as an antioxidant and precursor of phytochrome synthesis. The present study focuses on subcellular localization of HO1 in leaves of soybean has been investigated. Most activity appeared to be located within chloroplast due to its role in phytochrome synthesis but mitochondria also share its localization. Mitochondrial location of HO1 might be on its inner membranous space due to its role in the synthesis of electron donor species which facilitates HO1 catalyzed reaction. Study reports the co-localization of HO1 in both chloroplast and mitochondria.

Interaction between day length and phytohormones in the control of potato tuberization in the in vitro culture

We studied the interaction of the day length, cytokinins, and gibberellins in the control of tuberization in potato (Solanum tuberosum L, cv. Desire) plants and derived transgenic plants with the inserted PHYB gene from Arabidopsis encoding the synthesis of phytochrome B apoprotein and put under the control of the 35S CaMV promoter. Plantlets were cultured in vitro on hormone-free MS medium containing 5% sucrose and kinetin (1 mg/l) or/and GA (0.5 and 1.0 mg/l), at long day (LD, a 16-h photoperiod), short day (SD, a 10-h photoperiod), or continuous darkness conditions. The content of cytokinins (Ck, zeatin, and zeatin riboside) in various plant organs was determined by the immunoenzyme method, and GA activity was measured in bioassay with dwarf pea. Potato plant transformation with the PHYB gene enhanced substantially tuber initiation inhibition by LD. Kinetin addition to culture medium enhanced tuberization and reduced Ck content in aboveground shoots and Ck redistribution in the favor of underground organs. GA addition to the culture medium suppressed tuberization and induced Ck accumulation in aboveground organs. We concluded that Ck role in tuberization depends on their predominant localization in above- or underground potato organs. The involvement of Ck and GA in the competitive relations between growing tubers and shoots is considered.

The role of low intensity red luminescent radiation in the control of Arabidopsis thaliana morphogenesis and hormonal balance

The effects of low intensity red luminescent radiation emitted by the polyethylene light-correcting film due to the conversion of UV-A radiation on Arabidopsis thaliana (L.) Heynh. morphogenesis and hormonal balance were studied. Wild-type Ler plants and two mutants, hy3 and hy4, displaying disturbances in the synthesis of phytochrome B and cryptochrome 1, respectively, were compared. In wild-type and hy4 plants grown under the light-correcting film, growth and development were substantially accelerated, whereas, in hy3 plants, they were retarded. These changes were correlated with changes in the levels of endogenous hormones, both growth activators and inhibitors. We concluded that low intensity red luminescent radiation affected the plant hormonal balance. In its turn, the changes in the hormone ratios, growth stimulators and inhibitors, affected the rate of plant growth and their productivity.

The photoreceptors in the high irradiance response of plants

Several studies show that the high irradiance response (HIR) of plants is probably due to two photoreceptors. One of the photoreceptors is phytochrome, and the other is an unidentified pigment provisionally named heliochrome. One of the functions of heliochrome is the synthesis of phytochrome, using far‐red and blue radiations of high intensities, to replace the phytochrome destroyed by light. Another possible function could be an interaction of heliochrome with a substance produced by phytochrome. The data presented show that heliochrome is a pigment with different properties from phytochrome. It shows a far‐red/green reversibility. Heliochrome has been shown to participate with phytochrome in such HIRs as leaf movement in Albizzia and flowering in a long‐day plant. The first event initiated by phytochrome and by heliochrome could be the generation of a strong positive, electrostatic charge in the cell membrane.

Studies on the Synthesis of Phytochrome and Related Tetrapyrroles. Dihydropyrromethenones by Photochemical Rearrangement of N-Pyrrolo Enamides

Dihydropyrromethenone 67b, a potential precursor for the synthesis of phytochrome 1, has been prepared in enantiomerically pure form beginning with N-aminopyrrole 64 and the acetylenic acid 62b. The key step involved a 3,5-sigmatropic rearrangement of N-pyrrolo enamide 66b.

Very low fluence and low fluence response in the induction and inhibition of seed germination in Celosia argentea

AbstractSeeds of Celosia argentea L. displayed an absolute requirement for light for germination. Germination could be induced by far-red light as well as red light, and therefore, the effect of red light was not completely reversible by far-red light. A considerable proportion of the seed population was sensitive to red light within the range 0.01–0.1 μmol m−2, which suggested that phytochrome was operating in the very low fluence response mode. Seeds showed a gradual increase in germination in response to red light with an increasing duration of previous dark imbibition. Interruption of the dark imbibition period with very low fluence (10−4−10–1 μmol m−2) or low fluence (1–103 μmol m−2) red light prevented germination subsequent to a terminal, saturating red light irradiation. Increasing responsiveness to red light with increase in imbibition temperature and inhibition of development of photosensitivity by cycloheximide suggested that phytochrome synthesis occurred during the dark imbibition period. An absolute requirement for light for seed germination, inhibition of seed germination on interrupting the dark imbibition period with very low fluence or low fluence red light, and the recovery from photoinhibition of seeds in darkness are consistent with the hypothesis that PhyA is the principal phytochrome involved in the photoregulation of seed germination in C. argentea.

An unequivocal synthesis of the ring-A,B dihydropyrromethenone of phytochrome

Dihydropyrromethenone 1b (R = p-methoxybenzyl), a potential precursor for the synthesis of phytochrome ( 3), has been prepared in enantiomerically pure form beginning with the homochiral acetylenic lactone 18.

The kinetics of type 1 phytochrome in green, light-grown wheat (Triticum aestivum L.)

The kinetics of type 1 phytochrome were investigated in green, light-grown wheat. Phytochrome was measured by a quantitative sandwich enzyme-linked immunosorbent assay using monoclonal antibodies. The assay was capable of detecting down to 150 pg of phytochrome. In red light, rapid first-order destruction of the far-red-light-absorbing form of phytochrome (Pfr) with a half-life of 15 min was observed. Following white light terminated by red, phytochrome synthesis was delayed in darkness by about 15 h compared to plants given a terminal far-red treatment. Synthesis of the red-light-absorbing form of phytochrome (Pr) was zero-order in these experiments. Phytochrome synthesis in far-red light was approximately equal to synthesis in darkness in wheat although net destruction occurred in light-grown Avena sativa tissues in continuous far-red light, as has been reported for other monocotyledons. In wheat, destruction of Pfr apparently did not occur below a certain threshold level of Pfr or Pfr/total phytochrome. These results are consistent with an involvement of type 1 phytochrome in the photoperiodic control of flowering in wheat and other long-day plants.

Spectrophotometric and Spectrofluorometric Methods in Weed Science

The utility, advantages, and disadvantages of several spectrofluorometric and in vivo spectrophotometric methods in plant science are reviewed, with emphasis on their use in weed science. Examples of the use of in vivo, dual-wavelength spectrophotometry to probe the effects of herbicides on cytochromefoxidation/reduction, the P515 chromatic shift, phytochrome synthesis, and other processes are discussed. Use of in vivo spectrophotometry to examine effects of herbicides on phytylation of chlorophyllide, protochlorophyllide photoconversion, porphyrin accumulation, and other processes that result in spectral changes is discussed and examples are provided. Spectrofluorometric methods for pigment identification and quantification and for enzyme assays are illustrated with examples. Spectrofluorometric methods used in conjunction with high-pressure liquid chromatography (HPLC) and thin-layer chromatography (TLC) are discussed.

LIGHT PROMOTION OF SEED GERMINATION IN Datura ferox IS MEDIATED BY A HIGHLY STABLE POOL OF PHYTOCHROME

Abstract— The duration of the far‐red light‐absorbing form of phytochrome (Pfr) of the photoreceptor pool involved in the control of seed germination was investigated for Datura ferox seeds. These seeds require both Pfr and alternating temperatures (20/30°C) to germinate. After 24 h imbibition (25°C), the seeds received pretreatment‐light pulses providing different phytochrome photoequilibria (Pfr/P), followed by a 24 h dark incubation (25°C), and test‐light pulses providing different Pfr/P immediately prior to transfer to alternating temperatures. Germination increased with increasing Pfr/P provided by the test‐light pulses, but was unaffected by the pretreatment‐light pulses. This suggests that phytochrome synthesis, phytochrome degradation and phytochrome‐mediated changes in response to phytochrome were negligible. In other experiments, red light‐pretreatment pulses were followed by dark incubations (25°C) of different duration before transfer to alternating temperatures. The proportion of Pfr remaining after the 25°C incubation period was estimated by comparing germination rates with those of seeds that received test‐light pulses of known calculated Pfr/P immediately prior to the start of the cycles of alternating temperatures. More than 80% of the Pfr established by a Pfr/P= 0.87 light pulse was present and active even after 48 h dark incubation at 25°C. Surprisingly, when a pretreatmentlight pulse providing a Pfr/P= 0.70 was given, the reduction in [Pfr] was significantly faster.Germination of Datura ferox seeds is under the control of a highly stable (type II like) phytochrome pool. Apparently, this pool follows Pfr dark reversion to the red light‐absorbing form, the times to reach half the original Pfr pool being > 96 h or <14 h after light pulses providing Pfr/P= 0.87 or 0.70, respectively.

Inhibition of Phytochrome Synthesis by the Transaminase Inhibitor, 4-Amino-5-Fluoropentanoic Acid

Gardner and Gorton (1985 Plant Physiol 77: 540-543) demonstrated that the transaminase inhibitor gabaculine (5-amino-1,3-cyclohexadienyl-carboxylic acid) inhibits the initial synthesis and resynthesis of spectrophotometrically detectable phytochrome in vivo. Another mechanism-based transaminase inhibitor, 4-amino-5-fluoropentanoic acid (AFPA), is examined in this report for its effects on phytochrome synthesis in developing etiolated seedlings. Preemergence treatment with AFPA was found to inhibit initial phytochrome synthesis in peas (Pisum sativum L.), corn (Zea mays L.), and oats (Avena sativa L.). In general, reduction in phytochrome correlated with reduction in chlorophyll. However, the extent of inhibition of phytochrome synthesis was not as great as that of chlorophyll synthesis. These results confirm those with gabaculine, indicating that both initial synthesis and resynthesis of phytochrome require de novo synthesis of chromophore as well as apoprotein. AFPA was a more effective inhibitor of both chlorophyll and phytochrome synthesis than was gabaculine, suggesting that AFPA may be the preferred tool with which to probe the physiological consequences of the inhibition of phytochrome biosynthesis.

Involvement of phytochrome in Ca 2+-dependent, calmodulin independent L-leucine uptake in Brassica

Uptake of l-leucine in etiolated shoot tips of Brassica oleracea could be enhanced by 50% over control by red-light irradiation, which was reversed by far-red light. Red/far-red reversibility and the loss of red-light effect after treatment with inhibitor of phytochrome synthesis, gabaculine, indicates phytochrome control of this uptake. Maximum uptake was reached after 20 min incubation both in red-light-treated and etiolated shoot tips. Metabolic inhibitors e.g., azide, iodoacetate, 2,4-dinitrophenol and ouabain, a Na +/K + ATPase inhibitor, impeded uptake. Presence of l-glycine inhibited the uptake whereas that of d-leucine did not. Ca 2+ promoted the uptake, Na + and K + had no effect and Li + inhibited it. EGTA and verapamil alone had no effect but could inhibit Ca 2+-enhanced uptake. Calmodulin inhibitors, trifluoperazine and chloropromazine and specific calmodulin antagonists, compound 48/80 and calmidozolium (R24571), showed no inhibitory effect on Ca 2+-dependent l-leucine uptake. Ca 2+ ionophore A23187 proved stimulatory, increasing uptake by more than three fold. Results indicate a phytochrome regulated, Ca 2+-dependent, calmodulin-independent mechanism for l-leucine uptake.

Photoregulation of Phytochrome Synthesis in Germinating Embryos ofAvena sativaL.

Hilton, J. R. and Thomas, B. 1987. Photoregulation of phytochrome synthesis in germinating embryos of Avena sativa L.—J. exp. Bot. 38: 1704–1712. The effect of light on the accumulation of phytochrome in germinating Avena embryos was determined. A quantitative ELISA using monoclonal antibody AFRC MAC 56 was used to measure specifically type 1 (or dark) phytochrome. A pulse of red light given after 14 h imbibition but prior to the onset of type 1 phytochrome synthesis, strongly inhibited subsequent type 1 phytochrome accumulation. This effect of red light at 14 h was reversible by far-red light indicating the involvement of phytochrome. Red light also inhibited phytochrome synthesis after 18 h and 24 h imbibition but after 24 h, far-red light did not reverse the effect. The effect of red light treatment at 18 h was reversed by giving a pulse of far-red light at any time up to 30 h. Seed germination was not influenced by light under the conditions of these experiments. It is proposed that type 2 (or light) phytochrome may be responsible for photoregulation of type 1 phytochrome synthesis in germinating Avena embryos.

5' proximal sequences of a soybean ribulose-1,5-bisphosphate carboxylase small subunit gene direct light and phytochrome controlled transcription.

Two closely related ribulose-1,5-bisphosphate carboxylase small subunit (SSU) genes, SRS1 and SRS4, are transcribed at high levels in soybean plants in response to light. Transgenic petunia plants containing 5' sequences from SRS1 or SRS4 fused to the polypeptide encoding region of a neomycin phosphotransferase (NPTII) gene exhibit selectable kanamycin resistance. Deletion of three ATG codons from the region preceding the normal NPTII translation start site has little effect on the levels of kanamycin resistance in transformed plants. Run-on transcription assays in isolated nuclei demonstrate that transcription of the SRS1/NPTII chimera and the native petunia SSU11A gene subfamily is light regulated and under phytochrome control in leaves of transgenic plants. In young expanding leaves of fully light grown plants, transcription of these genes is markedly reduced within minutes of far-red treatment, while ribosomal DNA and actin gene transcription remains unchanged. This is analogous to the transcriptional response we observed for SRS1 and SRS4 in soybean seedlings. These data suggest (1) that transcription of SSU genes in both soybean and petunia require the continued presence or synthesis of phytochrome in the Pfr form and (2) that 5' sequences are sufficient to direct the phytochrome controlled transcriptional response of the SRS1 gene. In fully expanded mature leaves we found the transcription rates of the native SSU11A gene subfamily, the chimeric SRS1/NPTII gene, the rDNA genes, and several other control genes to be reduced markedly after far-red treatment or after extended periods of darkness. The contrast between results in young and mature leaves is discussed.

Effects of Gabaculine on Phytochrome Synthesis during Imbibition in Embryonic Axes of <italic>Pisum sativum</italic> L.

Effects of Gabaculine on Phytochrome Synthesis during Imbibition in Embryonic Axes of <italic>Pisum sativum</italic> L.

Phytochrome in Norflurazon‐treated seedlings of Pharbitis nil

The properties of phytochrome have been measured by dual‐wavelength spectropho‐tometry in the cotyledons of the short‐day plant Pharbitis nil Choisy cv. Violet, where it is known to play a role in flower induction. In plants de‐etiolated by a single white light period (4 h or longer), destruction of the far‐red absorbing form of phytochrome (Pfr) was twice as rapid as after 10 min red light. A small fraction of Pfr was stable. After de‐etiolation by a period of white light (6 h or longer) the rapid decrease of Pfr during the first 30 min was accompanied by a rapid increase of the red absorbing form of phytochrome (Pfr). This rapid increase of Pfr is probably due to dark reversion. Long term synthesis of phytochrome was inhibited by the presence of Pfr. Phytochrome synthesised in darkness showed the etiolated‐plant type characteristics and underwent rapid destruction upon photoconversion to Pfr. The stable Pfr identified here is possibly that pool of phytochrome associated with the long term promotive process in flower induction, and the rapidly reverting Pfr is that pool associated with the night break inhibition of flowering.

Inhibition of Phytochrome Synthesis by Gabaculine

Gabaculine (5-amino-1,3-cyclohexadienylcarboxylic acid), a transaminase inhibitor, also inhibits chlorophyll formation in plants, and the effect of this compound can be counteracted by 5-aminolevulinic acid (ALA) (Flint, personal communication, 1984). Since it is probable that ALA also serves as a precursor to phytochrome, the effects of gabaculine on phytochrome synthesis in developing etiolated seedlings were examined using in vivo spectrophotometry. Preemergence treatment with gabaculine was found to inhibit initial phytochrome synthesis in peas (Pisum sativum L.), corn (Zea mays L.), and oats (Avena sativa L.). In general, reduction in phytochrome correlated with reduction in chlorophyll. However, the extent of inhibition of phytochrome synthesis was not as great as that of chlorophyll synthesis, perhaps due to preexisting phytochrome in the seed. Foliar treatment of etiolated pea seedlings prior to light-induced destruction of phytochrome inhibited subsequent phytochrome resynthesis in the dark. These results suggest that both initial synthesis and resynthesis of phytochrome require de novo synthesis of chromophore as well as apoprotein.

Analysis of phytochrome kinetics in light-grown Avena sativa L. seedlings

The phytochrome content, the rate of phytochrome accumulation after a light/dark transition and the rate of phytochrome destruction after a 1.5 d reaccumulation period in darkness were measured in light grown Avena sativa L. seedlings. The results using spectrophotometrical methods (Norflurazon treated seedlings) and the radio-immunoassay (RIA) (green seedlings) were almost identical. The rate of phytochrome synthesis was analysed by measuring the activity of poly(A(+))-RNA coding for the phytochrome apoprotein. It was demonstrated that the rate of phytochrome synthesis is different in light and in dark. These results were confirmed by measuring the incorporation of radioactive label in vivo. Five minutes red (and 5 min far-red) light strongly reduces the rate of phytochrome synthesis. Even after prolonged dark periods only 50% of the initial rate of phytochrome synthesis is recovered for light and dark grown seedlings which received one red light pulse.