Gibberellic Acid-Promoted Lignification and Phenylalanine Ammonia-lyase Activity in a Dwarf Pea (Pisum sativum)

The effects of gibberellic acid on lignification in seedlings of a dwarf and a tall cultivar of pea (Pisum sativum) grown under red or white light or in the darkness, were studied. Gibberellic acid (10⁻⁶-10⁻⁴ m) promoted stem elongation in both light and dark and increased the percentage of lignin in the stems of the light-grown dwarf pea. The gibberellin had no effect on the lignin content of the tall pea although high concentrations (10⁻⁴ m) promoted growth of the tall plants. Time course studies indicated that the enhanced lignification in the gibberellin-treated dwarf plants occurred only after a lag period of several days. It was concluded that gibberellic acid-enhanced ligmification had no direct relation to gibberellic acid-promoted growth. The activity of phenylalanine ammonia-lyase (E.C. 4.3.1.5) was higher in gibberellin-treated dwarf plants grown under white or red light than in untreated dwarf plants. Gibberellic acid had no detectable effect on the activity of this enzyme when the plants were grown in darkness, just as it had no effect on lignification under dark conditions. The data suggest that in gibberellin-deficient peas the activity of phenylalanine ammonia-lyase is one of the limiting factors in lignification.     

Isolation and identification of a new growth inhibitor, raphanusanin, from radish seedlings and its role in light inhibition of hypocotyl growth

A neutral growth inhibitor, for which the name raphanusanin is proposed, has been isolated in crystalline form from light-exposed Sakurajima radish (Raphanus sativus var. hortensis f. gigantissimus Makino) seedlings and identified as a new compound, 3-methoxy-4-methylthio-2-piperithione by spectrometric analyses.

Applied raphanusanin inhibited the hypocotyl growth of etiolated radish and lettuce seedlings at concentrations higher than 1.5 × 10⁻⁶ molar.

The endogenous raphanusanin contents in cotyledons and hypocotyls of radish seedlings increased more under red light, but decreased or maintained the initial level in the dark. Its content in roots showed almost no change between the light and dark materials.

     

Reduction of the Gibberellin Content of Pharbitis Seeds by CCC and After-Effects in the Progeny

Plants of Pharbitis nil were treated with the growth retardant (2-chloroethyl) trimethylammonium chloride (CCC) shortly before and after anthesis. Fresh and dry weight of immature seeds were not affected by the CCC treatment.

The level of gibberellin-like activity in Pharbitis seeds as compared to control seeds was strongly reduced by CCC application. The progenies of the treated plants also had a much reduced GA content in the seedling stage. These results are interpreted to indicate that CCC blocks gibberellin biosynthesis in higher plants, as it does in the fungus Fusarium.

CCC applied via the roots accumulated in the immature seeds and was carried over to the following generation. Consequently, growth of CCC progenies was dwarfed and flower formation inhibited. Both phenomena were overcome by application of gibberellin A₃.

Three gibberellin-like substances (called fractions I, II, and III) were present in Pharbitis seeds and could be separated by thin-layer chromatography. All 3 fractions were also present in seeds treated with CCC. Fractions II and III were present in much higher quantities than fraction I. Both fractions II and III promoted growth of d5 corn but only fraction II was active in dwarf peas grown under red light.

     

Control of Flowering of Xanthium pensylvanicum by Red and Far-red Light

A study was made of the effects of various durations, intensities and combinations of red and far-red light interruptions on the flowering responses of Xanthium pensylvanicum Wallr. A dual response to treatments of far-red light was observed. In short dark periods, far-red light alone did not greatly affect flowering but was able to overcome the inhibition of flowering caused by red light. In dark periods longer than 15 hours, far-red inhibited flowering and added to rather than overcame the inhibition by red light. The dark period length required for far-red inhibition remained the same whether far-red was given at the start or at the eighth hour of darkness.

In 48-hour dark periods Xanthium showed 3 responses to additions of red and far-red light breaks: A) response to red light; B) response to far-red light; and C) response to red followed by far-red light. Red light given any time in the first 30 hours of darkness overcame the inhibitory effect of far-red light given at either the start or the eighth hour of darkness. Red light given later than the thirtieth hour did not overcome the far-red effect.

Approximately the same energy of red light was required to overcome the inhibitory effect of far-red at the second hour of darkness as was required to produce maximum red light inhibition at the eighth hour. Although far-red light was most inhibitory when given early in a long dark period, approximately the same energy of far-red light was required to saturate the far-red response at the fourth, eighth and sixteenth hours.

The results are discussed in relation to other reports of far-red inhibition of flowering in short-day plants.

     

Isolation, Crystallization, and Partial Identification of Potato Factor II from Potato Tubers

The isolation, crystallization, and partial identification of potato factor II, a stimulator from the chemically neutral fraction of potato extract, is described. The compound was originally found to stimulate elongation of dwarf peas grown under red light, a gibberellin bioassay. It melts between 137° and 139°. In paper chromatography it migrates to R_F 0.62 in isopropyl alcohol: ammonium hydroxide: H₂O (10:1:1, v/v). Based on infrared and NMR data, it does not contain a lactone ring and possibly possesses an amide radical and an OH⁻ group, as well as many methylene radicals. Potato factor II may be similar to certain of the fatty acid derivatives previously reported to stimulate growth of excised sections, but it is unique in that it stimulates growth of intact plants. This effect points to the need for completely separating neutral from acid gibberellin-like substances when the latter are assayed on dwarf peas.     

Role of polyamines in gibberellin-induced internode growth in peas

To determine the requirement for polyamines in gibberellin (GA) induced internode growth polyamine content was measured in internodes of peas of various internode phenotypes (slender, tall, dwarf, nana) with and without applied gibberellin (GA₃) and polyamine synthesis inhibitors. Polyamines were assayed as dansyl derivatives which were separated by reverse phase high performance liquid chromatography and detected by fluorescence spectrophotometry. The amounts of polyamines in the different genetic lines of peas, which differed in internode lengths and extractable GA content, correlated with the extent of internode elongation. High polyamine concentrations were associated with young internodes and decreased with internode expansion. Extremely short internodes of nana plants without GA exhibited equal or higher amine concentrations relative to internodes of other lines of peas and GA-stimulated nana seedlings. The polyamine synthesis inhibitors, α-difluoromethylornithine and α-difluoromethylarginine, independently or in combination, inhibited polyamine accumulation and internode elongation of tall peas and GA-stimulated nana plants. Agmatine and putrescine restored growth and endogenous polyamine content to variable degrees. However, exogenous polyamines were not effective in promoting growth unless intracellular amines were partially depleted.

These results suggest that polyamines do not have a role in cell elongation, but may be required to support cell proliferation. Polyamines do not mediate the entire action of GA in internode growth of peas since GA induction of growth involves both cell division and cell elongation, whereas polyamines appear to affect cell division only.

     

Light regulation on growth, development, and secondary metabolism of marine-derived filamentous fungi

Effects of different light conditions on development, growth, and secondary metabolism of three marine-derived filamentous fungi were investigated. Darkness irritated sexual development of Aspergillus glaucus HB1-19, while white, red, and blue lights improved its asexual behavior. The red and blue lights improved asexual stroma formation of Xylaria sp. (no. 2508), but the darkness and white light inhibited it. Differently, development of Halorosellinia sp. (no. 1403) turned out to be insensitive to any tested light irradiation. Upon the experimental data, no regularity was observed linking development with secondary metabolism. However, fungal growth showed inversely correlation with productions of major bioactive compounds (aspergiolide A, 1403C, and xyloketal B) from various strains. The results indicated that aspergiolide A biosynthesis favored blue light illumination, while 1403C and xyloketal B preferred red light irradiation. With the favorite light sensing conditions, productions of aspergiolide A, 1403C, and xyloketal B were enhanced by 32.9, 21.9, and 30.8 % compared with those in the dark, respectively. The phylogenetic analysis comparing the light-responding proteins of A. glaucus HB 1-19 with those in other systems indicated that A. glaucus HB 1-19 was closely related to Aspergillus spp. especially A. nidulans in spite of its role of marine-derived fungus. It indicated that marine fungi might conserve its light response system when adapting the marine environment. This work also offers useful information for process optimization involving light regulation on growth and metabolism for drug candidate production from light-sensitive marine fungi.     

Activities of growth inhibitors isolated from light-grown dwarf pea shoots

The growth inhibitory activities of 6 endogenous growth inhibitors isolated from light-grown dwarf peas (Pisum sativum cv. Progress No. 9) were examined in the epicotyl of dark-grown seedlings of the same cultivar in the dark in order to examine the possible contribution of these compounds to the growth inhibition brought about by red light. The activities of these natural inhibitors, including two A-2 and A-2 of as yet undetermined structure, were compared with those of synthetic growth retardants and benzyladenine. Samples were applied directly into the epicotyls via a glass capillary tube. In 24-h tests doses for a 25% inhibition (I₂₅) were: A-2, 4.3 × 10^-2: cis-xanthoxin, 1.2 × 10^-1 ; A-2, 1.6 × 10^-1; trans-xanthoxin, 1.2; R,S-dihydromaleimide, 3.5 × 10² and pisatin, 4.0 × 10² nmol plant^-1 . In 72-h tests, I₂₅'s were: benzyladenine, 1.5; AMO-1618 (ammonium-(5-hydroxycarvacryl)-trimethylchloride piperidine carboxylate), 2.4; R,S-dihydromaleimide, 4.0 × 10² and CCC (chlorocholine chloride), 1.1 × 10³ nmol plant^-1. -D-Glucosyl-R-dihydromaleimide had no activity at all. Benzyladenine caused the thickening as well as elongation inhibition of the epicotyls of intact plants. The possible involvement of A-2 and in the red light growth inhibition of dwarf peas is discussed.Abbreviations AMO-1618 ammonium-(5-hydroxycarvacryl)-trimethylchloride piperidine carboxylate - CCC chlorocholine chloride - G-DHMD -D-glucosyl-R-dihydromaleimide - I₂₅ dose required for a 25% growth inhibition - R red light author for correspondence     

Growth, Pigment Synthesis, and Ultrastructural Responses of Phaseolus vulgaris L. cv. Blue Lake to Intermittent and Flashing Light

Growing bean plants (Phaseolus vulgaris L. cv. Blue Lake) on cycles of 1 minute light-1 minute dark or 5 minutes light-5 minutes dark, providing an integrated 12 hours light-12 hours dark per day for each set of plants, led to production after 21 days of new leaves low or lacking in chloroplast pigments. Subsequently, dry weight increase was sharply cut. Leaf area was affected by the light regimes after the second week of growth. By the fourth week, plants on the 1 minute light-1 minute dark cycle showed about one-half the leaf area of the controls. Shoot growth was favored over root growth to the greatest degree on the 1 minute light-1 minute dark regimes. Chlorophyll a/b ratios were close to 3.0 in all of the intermittent light regimes, but the total amounts of chlorophyll in milligrams per primary leaf were higher from day 9 to day 23 for the 12 hour light-12 hours dark controls than for other plants.

Although they produced chlorophyll, the plants receiving 1 or 2 milliseconds per second of light continued to lose weight at the same rate as the dark controls; thus, it is assumed there was no net photosynthesis. Plants receiving flashing light allocated significantly more food reserves from the seed to roots than did dark controls. Total chlorophyll formation was significantly accelerated by 2 milliseconds per second light. With 1 millisecond per second light, it took 5 days longer to achieve the same level of chlorophyll. After the 18th day, there was a steady decline in chlorophyll, b degrading more rapidly than a.

It is thought that several light-driven reactions are involved in the observed pigment synthesis, photosynthesis, food allocation, and growth of bean. Some of these reactions may be cyclic and others linear. Collectively, they must reach a harmonic point for normal metabolism and development to occur. Because time courses for each of these reactions are different, the intermittent and flashing light technique offers the possibility of individually studying some of the key light-driven reactions.

     

Responses of Heterotrophic Cultures of Chlorella vulgaris Beyerinck to Darkness and Light. I. Pigment and pH Changes

Glucose cultures of Chlorella vulgaris were grown in white light, in monochromatic light, and in darkness. Difference spectra showed that all wavelengths resulted in increased pigmentation over the dark controls.

Cells irradiated with the 600 mμ beam showed a much higher absorption in the blue end of the spectrum with respect to the red end than is normally found in absorption spectra of white-light grown Chlorella cells.

Dry weight comparisons between monochromatic light and dark controls showed the controls to be somewhat higher. This demonstrated that the monochromatic irradiation produced pigment synthesis but no increase in growth. Dark growth experiments suggested that cultures incubated in darkness on glucose excreted an acidic product.

     

Über den Gibberellingehalt von Zwerg- und Normalerbsen im Rotlicht und die Wirkung von Chlorcholinchlorid auf das Wachstum der Erbsen

Zusammenfassung Keimlinge von Zwerg- und Normalerbsen wurden auf ihren Gibberellingehalt getestet. Die Normalerbsen enthalten etwa achtmal soviel wie die Zwerge von der Gibberellinfraktion I nach Kende und Lang, die auf Normalerbsen stark, auf Zwerge kaum wirkt.Beide Rassen wurden mit Chlorcholinchlorid, einem Hemmstoff der Gibberellinsynthese bei Fusarium, behandelt. Im Rotlicht werden die normalen Pflanzen verzwergt, doch im Dunkeln hat die Substanz nur einen geringen Effekt. Die Dunkelhemmung ist so groß wie die Hemmung der Zwerge in Rotlicht. Eine mögliche Erklärung für die Unwirksamkeit von CCC im Dunkeln ist, daß die Erbsen im Dunkeln zum Wachstum kein oder nur wenig Gibberellin benötigen.

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The content of gibberellin-like substances in dwarf and normal peas growing in red light, and the effect of chlorocholinchloride on growth of peas

Summary Dwarf and normal pea seedlings were extracted and their gibberellinlike substances bioassayed. The normal peas yield about 8 times as much as the dwarfs of gibberellin-fraction I after Kende and Lang, which is highly effective on normal peas but nearly without effect on dwarfs.Both cultivars were treated with the inhibitor of gibberellin-synthesis in the fungus Fusarium, chlorocholinchloride (CCC). In red light the normal plants are dwarfed, but in darkness this substance has only a weak inhibiting effect on growth. The inhibition in darkness is of the same magnitude as the effect of CCC on dwarfs growing in red light. A possible explanation for the ineffectiveness of CCC in darkness is that peas need there no gibberellin at all or only very minute amounts for growth.

     

Internode length in pisum: do the internode length genes effect growth in dark-grown plants?

Internode length in light-grown peas (Pisum sativum L.) is controlled by the interaction of genes occupying at least five major loci, Le, La, Cry, Na, and Lm. The present work shows that the genes at all of the loci examined (Le, Cry, and Na) also exert an effect on internode length in plants grown in complete darkness. Preliminary results using pure lines were verified using either segregating progenies or near isogenic lines. The major cause of the differences was due to a change in the number of cells per internode rather than to an alteration of the cell length. Since the genes occupying at least two of these loci, Le and Na, have been shown to be directly involved with gibberellin metabolism, it appears that gibberellins are not only essential for elongation in the dark but are limiting for elongation in the nana (extremely short, na), dwarf (Na le), and tall (Na Le) phenotypes. These results are supported by the large inhibitory effects of AMO 1618 treatments on stem elongation in dwarf and tall lines grown in the dark and the fact that applied gibberellic acid could overcome this inhibition and greatly promote elongation in a gibberellin-deficient na line. It is clear that the internode length genes, and in particular the alleles at the Le locus, are not acting by simply controlling the sensitivity of the plant to light.     

Photosynthetic apparatus of pea thylakoid membranes : response to growth light intensity

We investigated the effect of growth light intensity on the photosynthetic apparatus of pea (Pisum sativum) thylakoid membranes. Plants were grown either in a growth chamber at light intensities that ranged from 8 to 1050 microeinsteins per square meter per second, or outside under natural sunlight. In thylakoid membranes we determined: the amounts of active and inactive photosystem II, photosystem I, cytochrome b/f, and high potential cytochrome b₅₅₉, the rate of uncoupled electron transport, and the ratio of chlorophyll a to b. In leaves we determined: the amounts of the photosynthetic components per leaf area, the fresh weight per leaf area, the rate of electron transport, and the light compensation point. To minimize factors other than growth light intensity that may alter the photosynthetic apparatus, we focused on peas grown above the light compensation point (20-40 microeinsteins per square meter per second), and harvested only the unshaded leaves at the top of the plant. The maximum difference in the concentrations of the photosynthetic components was about 30% in thylakoids isolated from plants grown over a 10-fold range in light intensity, 100 to 1050 microeinsteins per square meter per second. Plants grown under natural sunlight were virtually indistinguishable from plants grown in growth chambers at the higher light intensities. On a leaf area basis, over the same growth light regime, the maximum difference in the concentration of the photosynthetic components was also about 30%. For peas grown at 1050 microeinsteins per square meter per second we found the concentrations of active photosystem II, photosystem I, and cytochrome b/f were about 2.1 millimoles per mol chlorophyll. There were an additional 20 to 33% of photosystem II complexes that were inactive. Over 90% of the heme-containing cytochrome f detected in the thylakoid membranes was active in linear electron transport. Based on these data, we do not find convincing evidence that the stoichiometries of the electron transport components in the thylakoid membrane, the size of the light-harvesting system serving the reaction centers, or the concentration of the photosynthetic components per leaf area, are regulated in response to different growth light intensities. The concept that emerges from this work is of a relatively fixed photosynthetic apparatus in thylakoid membranes of peas grown above the light compensation point.     

Exposing broiler eggs to green,red and white light during incubation

Previous work has shown that exposing broiler eggs to white light during incubation can improve hatchability and post-hatch animal welfare. It was hypothesized that due to how different wavelengths of light can affect avian physiology differently, and how pigmented eggshells filter light that different monochromatic wavelengths would have differential effects on hatchability and post-hatch animal welfare indicators. To determine, we incubated chicken eggs (n=6912) under either no light (dark), green light, red light or white light; the light level was 250 lux. White and red light were observed to increase hatch of fertile (P<0.05) over dark and green light incubated eggs. White, red and green light exposure during incubation improved (P<0.05) the proportion of non-defect chicks over dark incubated eggs. Post-hatch 45-day weight and feed conversion was not affected by light exposure of any wavelength (P>0.05). Fear response of during isolation and tonic immobility was reduced (P<0.05) in broilers incubated under white or red light when compared with either green or dark broilers. Broilers incubated with white or red light had lower (P<0.05) composite asymmetry scores and higher (P<0.05) humoral immunity titers than dark incubated broilers, however, green light broilers did not differ (P>0.05) from dark incubated broilers. All light incubated broilers had lower (P<0.05) plasma corticosterone and higher (P<0.05) plasma serotonin concentrations than dark incubated broilers. These results indicate that white light and red light that is a component of it are possibly the key spectrum to improving hatchability and lower fear and stress susceptibility, whereas green light is not as effective. Incubating broiler eggs under these spectrums could be used to improve hatchery efficiency and post-hatch animal welfare at the same time.     

Gibberellin-auxin interaction in pea stem elongation

Joint application of gibberellic acid and indole-3-acetic acid to excised stem sections, terminal cuttings, and decapitated plants of a green dwarf pea results in a markedly synergistic growth response to these hormones. Synergism in green tall pea stem sections is comparatively small, although growth is kinetically indistinguishable from similarly treated dwarf sections.

Gibberellin-induced growth does not appear to be mediated through its effect on auxin synthesis, since gibberellin pretreatment of dwarf cuttings fails to elicit an enhanced tryptophan-induced growth response of sections, whereas auxin-induced growth is strongly enhanced. Also, tryptophan-gibberellin synergism is not significant in sections and cuttings of green dwarf peas, while auxin-gibberellin synergism is.

Administration of gibberellic acid prior to indole-3-acetic acid results in greatly increased growth. In reversed order, the application fails to produce any synergistic interaction. This indicates that gibberellin action must precede auxin action in growth regulation.

     

Evidence for Phytochrome Regulation of Gibberellin A(20) 3beta-Hydroxylation in Shoots of Dwarf (lele) Pisum sativum L

The effect of light on the dwarfing allele, le, in Pisum sativum L. was tested as the growth response to gibberellins prior to or beyond the presumed block in the gibberellin biosynthetic pathway. The response to the substrate (GA₂₀), the product (GA₁), and a nonendogenous early precursor (steviol) was compared in plants bearing the normal Le and the deficient lele genotypes in plants made low in gibberellin content genetically (nana lines) or by paclobutrazol treatment to tall (cv Alaska) and dwarf (cv Progress) peas. Both genotypes responded to GA₁ under red irradiation and in darkness. The lele plants grew in response to GA₂₀ and steviol in darkness but showed a much smaller response when red irradiated. The Le plants responded to GA₂₀ and steviol in both light and darkness. The red effects on lele plants were largely reversible by far-red irradiation. It is concluded that the deficiency in 3β-hydroxylation of GA₂₀ to GA₁ in genotype lele is due to a Pfr-induced blockage in the expression of that activity.     

Metarhizium robertsii illuminated during mycelial growth produces conidia with increased germination speed and virulence

Light conditions during fungal growth are well known to cause several physiological adaptations in the conidia produced. In this study, conidia of the entomopathogenic fungi Metarhizium robertsii were produced on: 1) potato dextrose agar (PDA) medium in the dark; 2) PDA medium under white light (4.98 W m^?2); 3) PDA medium under blue light (4.8 W m^?2); 4) PDA medium under red light (2.8 W m^?2); and 5) minimum medium (Czapek medium without sucrose) supplemented with 3 % lactose (MML) in the dark. The conidial production, the speed of conidial germination, and the virulence to the insect Tenebrio molitor (Coleoptera: Tenebrionidae) were evaluated. Conidia produced on MML or PDA medium under white or blue light germinated faster than conidia produced on PDA medium in the dark. Conidia produced under red light germinated slower than conidia produced on PDA medium in the dark. Conidia produced on MML were the most virulent, followed by conidia produced on PDA medium under white light. The fungus grown under blue light produced more conidia than the fungus grown in the dark. The quantity of conidia produced for the fungus grown in the dark, under white, and red light was similar. The MML afforded the least conidial production. In conclusion, white light produced conidia that germinated faster and killed the insects faster; in addition, blue light afforded the highest conidial production.     

The early time course of the inhibition of stem growth of etiolated pea seedlings by fluorescent light

The stem elongation responses of etiolated peas (Pisum sativum L.) to fluorescent light (35–45 mol.m^t-2.s^-1) were recorded using high resolution position transducers. Continuous fluorescent light decreased growth by 70% within 9 min. The growth rate declined to 5% of the control over the next 2 h and remained at this level for 7 h. Pulses of fluorescent light ranging from 8 s to 34 min led to partial suppression of growth and resulted in a complex kinetic response. The distinctive kinetics of blue and red light inhibition were apparent as components of the responses to non-saturating levels of fluorescent light. The rapid suppression of growth by blue light was not affected by concomitant red light. The lag time for the onset of red light inhibition was not affected by concomitant blue, but the rate of inhibition appeared accelerated.     

Growth, greening, and phytochrome in etiolated spirodela (lemnaceae)

Two species of Spirodela were grown aseptically in a simple mineral medium containing sucrose. Weak red light (15 erg cm⁻² sec⁻¹) enhanced dark growth of S. oligorrhiza, whereas weak far red light (15 erg cm⁻² sec⁻¹) when given after the red light reduced this effect.     

The role of xanthoxin in the inhibition of pea seedling growth by red light

Intermittent periods of red light illumination inhibit the growth of dwarf pea seedlings within 1 day and stop length increase completely in 3 days. Cis,trans-xanthoxin is synthesized within the plant during the 1st day of illumination and reaches a maximum level on the 3rd day. The red light treatment also causes an increase in the levels of violaxanthin, linoleic acid and peroxidase, lipoxygenase and carotene-bleaching activities in the plant. The possible control of xanthoxin production is discussed.