Inhibition of flowering by hexahydrofluorene-9-carboxylic acids related to allogibberic acid

1,2,3,4,4a,9a-hexahydrofluorene-9-carboxylic acid isomers have been prepared by reduction of fluorene-9-carboxylic acid. Hexahydrofluorene-9-carboxylic acid (5) produce inhibition of flowering and vegetative frond development in Lemna perpusilla 6746 similar to that observed with allogibberic acid. The stereochemical requirements for this type of biological activity in allogibberic acids and hexahydrofluorene-9-carboxylic acids are considered.     

Effects of DCMU on long-day flowering of Lemna perpusilla 6746 and photosynthetic mutant strain 1073

Long-day flowering of wild-type Lemna perpusilla (strain 6746)on ammonium-free medium with sucrose occurred in continuouslight of low intensity (25 ft-c). In higher intensities of light,frond production was increased and flowering was reduced. Thephotosynthetic inhibitor DCMU inhibited frond production andpromoted flowering in the presence or absence of exogenous sucrose.In the photosynthetic mutant strain 1073, the higher intensitiesof light inhibited frond production, but did not reduce flowering.DCMU increased mutant frond production, thus leading to increasedflowering percents. The mechanism by which DCMU affects floweringand growth appears to differ from that of other flower-promotingsupplements reported by Takimoto and Tanaka. The results suggestthat inhibition of photosynthesis enhances flowering in longdays. (Received June 25, 1977; )     

Lack of flowering responses to DGMU in a mutant of Lemna perpusilla 6746

DCMU, in a sucrose supplemented medium, promoted short and longday flowering and inhibited long day frond production of wildtype Lemna perpusilla 6746, but not of mutant strain 1073. Resultssuggest a defect in the mutant that mimics DCMU poisoning. ¹ This work was supported by National Science Foundation GrantGB-12955. (Received December 11, 1972; )     

Gibberellin and CCC Effects on Flowering and Growth in the Long-day Plant Lemna gibba G3

The application of gibberellic acid (GA₃) to the non-rosette long-day plant Lemna gibba G3 at concentrations from 0.1 to 100 mg/l did not induce flowering on short days and inhibited flowering on long days at concentrations of 1 mg/l and higher. On both short and long days GA₃ concentrations above 1 mg/l caused a decrease in frond size and fresh and dry weight, but an increase in the rate of frond production and thus an increase in the # VF (number of vegetative fronds). Identical results were obtained when gibberellin A₇ was used instead of GA₃.     

Morphogenesis of Asymmetry and Symmetry in Lemna perpusilla Torr

The determination of which of the two reproductive pockets ofLemna perpusilla will produce the first daughter frond undernon-flowering conditions, or the flower under inductive conditions,is related to asymmetric gradients within the mother frond whichmay be the result of the asymmetric position of the axillaryfrond. Treatments that encourage vigorous growth of axillaryfronds may overcome the normal situation of asymmetry and causea morphological symmetry which is qualitatively different fromthe symmetry induced by treatments (TIBA) which prevent thedevelopment of axillary fronds. The origin and maintenance ofasymmetry in Lemna are discussed. Lemna perpusilla Torr., morphogenesis, asymmetry of fronds, tri-iodobenzoic acid     

TWO LIGHT REACTIONS IN THE PHOTOPERIODIC CONTROL OF FLOWERING OF LEMNA PERPUSILLA AND L. GIBBA

The effects of light quality on the photoperiodic control inthe flowering of a SD duckweed, Lemna perpusilla strain 6746,and a LD duckweed, L. gibba strain G3, were investigated withspecial reference to the interaction between R and B or FR lights. In the diurnal alternation of R or G light and dark periods,L. perpusilla responded as a SDP, but in that of B or FR lightit was almost daylength-indifferent. On the other hand, L. gibbaresponded as a LDP under B, R or FR light, although the criticallight length was altered by the light quality. In the diurnal alternation of R and B or FR light periods containingno dark period, L. perpusilla flowered with the shortening ofthe optimal and critical R light lengths, compared with theplant exposed to that of R light and dark period. The floweringresponse of L. gibba to the R light length showed double peaks,that is, the first peak at the R duration less than 9 hours,and the second at the R duration longer than 9 hours. The firstpeak corresponds to the optimal R light length in L. perpusilla. Under the CL with a mixture of R and B or FR lights, the floweringand frond production were influenced by the intensity ratioof two light given. In both plants, the optimal ratio of B toR or FR to R for the flowering was always greater than thatfor the frond production. It is suggested that the B or FR light interacts with the Rlight in the photoperiodic process in the plants and this interactionbetween the R and B or FR lights should be of importance forobtaining a better understanding of photoperiodism. (Received August 28, 1965; )     

Factors Affecting the Water-sensitive Phase of Flowering in the Short Day Plant Lemna perpusilla

The flowering of Lemna perpusilla strain 6746 is inhibited by daily transfers to water for short periods during a sensitive phase. Supplementing the water with Ca(NO₃)₂ partially reverses the inhibition of flowering while MgSO₄ increases the inhibition. The inhibition by MgSO₄ is overcome by low concentrations of Ca(NO₃)₂. Flower-promoting activity was detected in water and in MgSO₄ solutions that had been incubated with plants under dark but not light conditions. The prevention of this effect by light appears to be photosynthetic rather than to depend on phytochrome. The activity is destroyed by autoclaving but not by brief boiling. This loss of a flower-promoting material may explain the inhibiting effect on flowering by transfers to water.     

Influence of Giving Salicylic Acid for Different Time Periods on Flowering and Growth in the Long-Day Plant Lemna gibba G3

When the long-day plant Lemna gibba L., strain G3 is grown under continuous light on ammonium-free half-strength Hutner's medium (NH₄⁺-free 0.5 H medium) growth is excellent, but flowering is severely inhibited and often is zero. Addition of 10 micromolar salicylic acid (SA) to NH₄⁺-free 0.5 H medium quickly reverses this inhibition and leads to optimal flowering. The SA treatment also leads to a considerable reduction in the growth rate and increase in frond gibbosity. Removal of SA from the medium quickly leads to an increase in the growth rate and a large decrease in flowering. Thus, for maximal effectiveness SA must be present in the medium for the entire experiment, and the effect of SA is clearly not inductive.     

EXPERIMENTAL CONTROL OF FLOWERING IN LEMNA I. GENERAL METHODS. PHOTOPERIODISM IN L. PEPUSILLA 6746

Hillman , William S. (Yale U., New Haven, Conn.) Experimental control of flowering in Lemna. I. General methods. Photoperiodism in L. perpusilla 6746. Amer. Jour. Bot. 46(6): 466–473. Illus. 1959.—Lemna perpusilla strain 6746 flowers as a typical short-day plant when grown aseptically in Hutner's medium (containing ethylenediaminetetraacetic acid, [EDTA]) at 26–28°C. A method is described for quantitatively assaying the degree of flowering in a culture. Maximal flowering takes place under photoperiods of 6–11 hr., and none under photoperiods exceeding 15 hr. The flower-promoting effects of long nights are inhibited by brief interruptions with red light, such interruptions being most effective in the middle of the dark period. A single long night will cause the subsequent production of flowering fronds, but vegetative growth in the culture is resumed after a time. Only frond primordia at a very early stage of development appear to be sensitive to induction. Quantitative flowering experiments lasting a week or less can easily be performed with this plant; it is ideally suited for studies of the effects of light, darkness, temperature, organic compounds and other factors under highly controlled conditions.     

Response of Lemma perpusilla to Periodic Transfer to Distilled Water

The flowering of Lemma perpusilla grown on half-strength Hutner's medium with sucrose under inductive photo-periods is inhibited in a periodic manner by daily transfers to water for short periods of time. The phase of maximal inhibition of flowering caused by water treatment is about 1 to 2 hours after the time of maximal sensitivity to light pulses. The rhythm of sensitivity to water treatments does not persist under continuous blue light. Supplementing the water with either Ca(NO₃)₂ or K₂HPO₄ partially reverses the inhibition of flowering, with the first salt being more effective. Supplementation with NH₄NO₃ or MgSO₄ increases the inhibition. The water effect on flowering is not observed in plants grown on half-strength Hutner's medium without sucrose. The water treatments may act by removing or destroying a crucial precursor for photoperiodic induction, with the other conditions modifying permeability. The system provides a new technique for investigating the mechanism of photoperiodic induction.     

Inhibitory Effect of Carbohydrate on Flowering in Lemna perpusilla: II. Reversal by Glycine and l-Aspartate. Correlation with Reduced Levels of beta-Carotene and Chlorophyll

Flowering of Lemna perpusilla strain 6746 grown on 0.1 strength Hutner's medium in short days was inhibited by sucrose, glucose, fructose, and mannose, but not by various other sugars or metabolic intermediates. Only those sugars that inhibited flowering supported heterotrophic growth. Experiments with a single inductive long night indicated that an early stage in flowering was the sugar-sensitive process. Inhibition of flowering by carbohydrates was accompanied by reduced levels of chlorophyll and β-carotene. The inhibitory effects of carbohydrates on flowering were partially reversed by iminodiacetate, glycine, and l-aspartate but not by d-aspartate, ethylenediaminetetraacetate, acetate, δ-aminolevulinic acid, or mevalonic acid. The possibility is discussed that carbohydrate repression of flowering and of chloroplast pigments resulted from inadequate levels of amino acids.     

Influence of Temperature on the Flowering of Lemna perpusilla 6746 Grown under Skeleton Photoperiods

The flowering of Lemna perpusilla Torr. strain 6746 grown under 24-hour skeleton photoperiods consisting of 13- and 10.5-hour dark periods separated by 0.25-hr light pulses is strongly dependent on temperature. When plants are cultured in 50-ml Erlenmeyer flasks containing 20 ml of half-strength Hutner's medium supplemented with 1% (w/v) sucrose maximum, per cent, flowering occurs at 23 C. At temperatures above and below 23 C a marked decline in per cent flowering is seen.     

Seed and Embryo Germination in Syringa vulgaris and S. reflexa as Affected by Temperature during Seed Development

A nitrogen source was needed for the flowering of Lemna gibba L., a long-day plant, and L. perpusilla Torr., a shortday plant. The level of endogenous amino acids analyzed by an Amino Acid Analyzer, rose during the first few inductive cycles, but was reduced during later stages of the flowering process. Serine and threonine levels increased during the light period and decreased during the dark period in L. perpusilla. Exogenous serine and threonine added to the culture medium at 10^?6M increased the rate of flowering by more than 35% over the controls. Cysteine inhibited flowering, while other amino acids had little or no promotive effect on flowering. Serine and threonine increased flowering rate in L. perpusilla only when added during a dark period of the inductive cycle. The addition of amino acids during a light period not followed by a dark period had no effect on flowering.     

EXPERIMENTAL CONTROL OF FLOWERING IN LEMNA. III. A RELATIONSHIP BETWEEN MEDIUM COMPOSITION AND THE OPPOSITE PHOTOPERIODIC RESPONSES OF L. PERPUSILLA 6746 AND L. GIBBA G3

Hillman , William S. (Yale U., New Haven, Conn.) Experimental control of flowering in Lemna. III. A relationship between medium composition and the opposite photoperiodic responses of L. perpusilla 6746 and L. gibba G3. Amer. Jour. Bot. 48(5): 413–419. Illus. 1961.—In a simple Hoagland-type medium, L. perpusilla 6746 flowers, irrespective of daylength; L. gibba G3 does not flower under any daylength, as long as the medium is changed frequently. Ethylenediaminetetraacetic acid (EDTA) prevents the flowering of L. perpusilla in long days but not in short, and brings about the flowering of L. gibba in long days but not in short. The same results are obtained with medium “aged” by the growth in it of L. gibba for several weeks (in any photoperiod) as well as with tartaric acid. The effectiveness of both EDTA and “aged” medium is greater at pH levels near 5 than at 4 or below, probably reflecting action through metal-chelation. These results are most easily interpreted as effects on photoperiodic sensitivity, and suggest a central role of metals in photoperiodism.     

Flowering responses of Lemna perpusilla and L. gibba in relation to nitrate concentration in the culture medium

Flowering responses of Lemna perpusilla strain 6746, a short-dayplant, and L. gibba strain G3, a long-day plant, to nitrateconcentration in Hoagland's type medium with or without EDTA,were compared. Maximum flowering of L. perpusilla under SD occurredat higher nitrate concentrations than did colony proliferation.Even under CL, L. perpusilla grown at sub-optimal nitrate concentrationsfor colony proliferation, flowered irrespective of the presenceof EDTA which reduces flowering. Unlike L. perpusilla, L. gibba failed to flower under SD atany nitrate concentration whether or not EDTA was added. UnderCL, however, L. gibba flowered at almost any nitrate concentrationwith or without EDTA. Double optima for nitrate concentrationwas exhibited in the presence of EDTA; optimal concentrationfor colony proliferation came between the two optima for flowering. We concluded that the nitrogen level of the medium is importantin regulating flowering of duckweeds, and that the effect ofEDTA, if any, may primarily be on colony proliferation and onlysecondarily or antagonistically on flowering. ¹ Present address: Institute for Agricultural Research, TohokuUniversity, Sendai 980, Japan. (Received September 25, 1971; )     

Lipid metabolism in the fern Polypodium vulgare

The early stages of spore germination in Polypodium vulgare involve the catabolism of endogenous triglyceride which is accompanied by the de novo synthesis of several classes of neutral and polar lipid. These newly synthesized lipids include triglycerides which possess different fatty acid compositions from those in dormant spores and resemble the triglyceride fraction found in the sporophyte frond tissue. The C₂₀ acids which are present in the non-chloroplast lipids of the sporophyte frond tissue do not occur in the spore to an appreciable extent.     

Photoreversibility of flower initiation in Lemna perpusilla as affected by the light quality of the main light period

Reversible floral responses of Lemna perpusilla to red and far-redlights appeared only at the beginning of the inductive darkperiod when the 8 hr photoperiod consisted of white or red light.When blue or far-red light was given during the 8 hr photoperiod,the far-red given at the beginning of the dark period scarcelyinhibited flowering; red/far-red reversibility newly appearedat the middle of the dark period. This indicates that the photoregulationsystem in the flowering of L. perpusilla can be converted fromthe Pharbitis type to the Xanthium type by changing the lightquality of the main photoperiod from white or red to blue orto far-red, which is known to be effective for the so-calledhigh-energy photoreaction of photomorphogenesis. (Received July 2, 1975; )     

Photoperiodic Entrainment Patterns in the CO(2) Output of Lemna perpusilla 6746 and of Several Other Lemnaceae

The CO₂ output of Lemna perpusilla 6746 in “skeleton photoperiods” consisting of alternating 10½-hour and 13-hour dark periods separated by ¼-hour illuminations was recorded under stable high and low nitrate conditions. The phase relationship finally attained between light schedule and output is the same regardless of which dark period is given first, but entrainment is more rapid (as is flowering) with an initial 13-hour dark period. In all respects other than bistability—the assumption of two different stable phase relationships depending on the initial dark period—both flowering and the course of CO₂ output conform to Pittendrigh's model derived from Drosophila eclosion rhythms, confirming the view that an endogenous circadian rhythm, or biological clock, underlies the photoperiodic control of flowering in this plant. Experiments with rigorous temperature control show that earlier results with long light exposures were in part due to temperature changes; in consequence, it is clear that entrainment patterns with high nitrate differ even more from those in low nitrate than was previously evident, and not simply by the addition of a “nitrate peak.” Other Lemnaceae tested with a few simple light-dark schedules in both types of media show a variety of responses, with no obvious correlation to photoperiodic response type.     

Influence of Short Term Inhibitor Treatment on the Flowering of Lemna perpusilla 6746

Short term inhibitor treatment can be used to examine the processes that occur during an inductive dark period in the short day plant Lemna perpusilla Torr., strain 6746. Several inhibitors of protein biosynthesis are most effective in reducing per cent flowering when treatment occurs over the 2-hour intervals beginning at the 12th hour or the 14th hour of a 8 (16) photoperiodic cycle. The antimetabolite, 5-fluorouracil, is most effective when treatment occurs early in the dark period. Evidence is cited suggesting that distilled water incubation inhibits flowering by interfering with protein biosynthesis.