Physiological Function of Night Interruption in Lemna paucicostata 6746: Action of Light as a Phaser on the Photoperiodic Clock

Flowering of Lemna paucicostata 6746 under short-day conditionsis completely inhibited by daily night interruption given tothe "inhibition zone" that starts at Zeitgeber time (ZT) 14,i.e., 14 h after dawn, and ends 14 h before the next dusk [Oota(1983a) Plant & Cell Physiol. 24: 327]. With a modifiedmin-SD method, most of these night interruptions were foundto signal the false dawn (false ZT 0) after one entraining cycle.Thus, on and after day 2 the interruption was associated withthe next main photoperiod to form a noninductive skeleton photoperiod.However, a light pulse applied at the start of the inhibitionzone, caused no phase shift in the photoperiodic clock, andformed a noninductive skeleton photoperiod in association withthe preceding main photoperiod. The complete floral inhibition due to the night interruptionwas ascribed to the illumination of both the LI-phase (realor false ZT 0) and L2-phase (real or false ZT 14), or the twolight-sensitive fractions of the original or shifted criticalphotoperiod, by the thus formed skeleton photoperiod, just aswas the case for the floral inhibition by complete photoperiodslonger than the critical daylength, 14 h [Oota (1983a), Oota(1983b) Plant & Cell Physiol. 24: 1503]. (Received October 20, 1983; Accepted January 7, 1984)     

Floral Induction in Short-Day Lemna paucicostata 6746 by 8-Hydroxyquinoline, under Long Days

Lemna paucicostata 6746 is a short-day duckweed and flowersin response to a single photoinductive cycle. Its critical darkperiod requirement is ca. 10 h. Flowering in this duckweed couldbe induced by 8-hydroxyquinoline (8-HQ) under an otherwise non-inductivelong-day regime of 16 h light and 8 h darkness; the criticaldark period requirement for initiation of flowering was thusreduced by at least 2 h. However, 8-HQ was ineffective in initiatingflowering in strain 6746 under continuous illumination. Atomicabsorption analysis of the plant material revealed that thecontent of both iron and copper is markedly higher in the plantstreated with 8-HQ. A comparison of the effects of 8-HQ and thoseof EDTA and ethylenediamine-di(o-hydroxyphenylacetic acid),on flowering of strain 6746, has also been made. (Received August 23, 1983; Accepted October 18, 1983)     

Measurement of the Critical Nyctoperiod by Lemna paucicostata 6746 Grown in Continuous Light

The short-day duckweed Lemna paucicostata 6746 could be inducedto flower in two days at 26C when continuous illumination forentrainment was followed by continuous darkness. This 48-h darkperiod or the minimum darkness requirement for floral inductionwas called the induction period. The length of the inductionperiod (IP) was routinely computed as the number of 24-h cyclesusing the equation of regression of flower number in logarithmon culture time. A light pulse given about 7 h after the startof the induction period increased the apparent IP value fromtwo to three, suggesting that the interrupted first day hadfunctioned as a noninductive day. A pulse given at any otherpart of the induction period did not modify the IP value. Thelight-sensitive part is probably the inducible phase, and thefirst 7-h period of darkness terminated by it seems to be thecritical nyctoperiod. These and relevant facts suggest thatthe light-off oscillator measures the critical night length,7 h. Either red or far-red irradiation at the inducible phase extendedthe IP value by one. No red/far-red photoreversibility was detected.As expected, however, red or far-red irradiation of any otherpart of the critical nyctoperiod could not modify the IP value. (Received February 8, 1985; Accepted May 14, 1985)     

Enhancement of long-day flowering by Mo-deficiency and application of some amino acids and asparagine in the short-day plant Lemna paucicostata 6746

The short-day plant Lemna paucicostata 6746 can be induced toflower on long days (continuous light) by the addition of copper,tungstate or ferricyanide to the medium, and in each case theeffect was greatly enhanced by deleting molybdate from the medium.Treatment with asparagine, aspartate, glutamate, -alanine, glycineorserine, all of which are known to increase the critical daylengthand almost nullify the light-break effect in L. paucicostata6746, enhanced the long-day flowering induced by copper, ferricyanideor Mo-deficiency. (Received October 13, 1978; )     

Effects of some metabolic inhibitors on flowering of Lemna gibba G3, a long-day duckweed

Flowering of Lemna gibba G₃, a long-day duckweed, was inhibitedby adding CuSO₄, AgNO₃, HgCl₂, Na₂WO₄ or iodoacetamide to themedium at the concentrations inducing long-day flowering inLemna paucicostata 6746, a short-day duckweed. This suggeststhat these metabolic inhibitors affected the photoperiodic sensitivityrather than directly affecting flower initiation. Ferricyanidepromoted flowering in both of these short-day and long-day duckweeds. (Received July 7, 1977; )     

Effect of L-Pipecolic Acid on Flowering in Lemna paucicostata and Lemna gibba

L-Pipecolic acid was found to be effective in inducing floweringof Lemna paucicostata 151, 381, 441 and 6746, and of Lemna gibbaG3. When the plants were grown on half-strength Hutner's medium,L-pipecolic acid caused profuse flowering of L. paucicostata151 maintained under 9 and 10 h of light daily. In L. paucicostata441 and 6746, L-pipecolic acid had a strong flower-promotingeffect under a near critical photoperiod. In L. paucicostata381, by contrast, L-pipecolic acid had only a very small effecton flowering. In L. gibba G3 substantial promotion of floweringwas observed under continuous light. When one-twentieth-strengthHutner's medium was used as the basic medium, L-pipecolic acidstimulated flowering in all strains of Lemna examined, evenunder continuous light. When L. paucicostata 151 was grown on one-tenth-strength M mediumor one-twentieth-strength Hutner's medium, the flower-inducingactivity of L-pipecolic acid was greatly enhanced by cytokininunder continuous light. However, when this strain was grownwith 9 h of illumination daily, this synergistic effect of cytokininwas only slight. A short-term (even 1-h) treatment with L-pipecolicacid resulted in flowering, suggesting that L-pipecolic acidis involved in the induction of flowering, rather than its evocation.D-Pipecolic acid also had flower-inducing activity, but itsactivity was 50 times lower than that of the L-isomer. (Received January 23, 1992; Accepted March 9, 1992)     

Inhibition of Flower Induction by Some Inhibitors of Protease in the Short-day Plant Lemna paucicostata 6746 and the Long-day Plant Lemna gibba G3

Four inhibitors of proteases, namely, bestatin, diisopropylfluorophosphate, elastatinal and p-toluenesulfonyl-L-lysinechloromethyl ketone hydrochloride, were examined for their effectson flowering of a short-day plant Lemna paucicostata 6746 anda long-day plant Lemna gibba G3. Each of the inhibitors greatlyinhibited the flowering of Lemna paucicostata 6746 that is normallyinduced by nitrogen deficiency. Bestatin or elastatinal givenonly during the first half of the culture period inhibited theflowering more clearly than when each was given during the latterhalf, suggesting that they inhibited the inductive process(es)involved in flowering rather than development of flower buds.Bestatin or elastatinal greatly inhibited the flowering of Lemnapaucicostata 6746 induced by photoperiodic stimulus, ferricyanideand continuous far-red light. Simultaneous application of thesetwo inhibitors was more effective in the inhibition of photoperiodicallyinduced and ferricyanide-induced flowering than was each inhibitoralone. They also completely inhibited the photoperiodic floweringof Lemna gibba G3. These results suggest that the inductionor activation of some proteases, probably followed by the degradationof some protein(s), is necessary for the induction of floweringin both these plants. (Received November 21, 1989; Accepted February 19, 1990)     

Flower-Inducing Activity of Lysine in Lemna paucicostata 6746

Flowering of Lemna paucicostata 6746, a typical short-day plant,was induced by culture for 96 or 120 h in nitrogen-free mediumunder continuous illumination. To examine the effects of lysine,we homogenized entire plants of L. paucicostata 151 in a solutionof lysine and the supernatant obtained after centrifugationof the homogenate was added to the medium to give various concentrationsof lysine in the medium. Flowering of strain 6746 in nitrogen-freeor nitrogen-deficient culture medium was effectively promotedby the addition of a lysine-containing supernatant to the medium.The suppressive effect of elastatinal, a protease inhibitor,on the induction of flowering was almost completely reversedby the simultaneous application of a lysine-containing supernatantto the medium. During nitrogen-free culture, the level of endogenousfree lysine, expressed on the basis of the amount of total freeamino acids, increased. Lysine-containing supernatants alsoinduced flowering of plants in nitrogen-rich medium under continuousillumination. These findings suggest that endogenous lysineis involved in the induction of flowering in L. paucicostata6746 on nitrogen-free or nitrogen-deficient medium, as it isin the induction of flowering in L. paucicostata 151 (Received July 29, 1996; Accepted November 18, 1996)     

Absorption of Copper by Lemna as Influenced by Some Factors Which Nullify the Copper Effect on Flowering and Growth

The effect of copper on flowering and growth of Lemna paucicostata6746 and Lemna gibba G3 in a copper-containing medium is nullifiedby the addition of EDTA, ammonium ions or salicylic acid tothe medium or a decrease in its nitrate concentration. Thesefactors were examined for their effects on the absorption ofcopper by the plants. The addition of EDTA to the medium completelyinhibited the absorption of copper in both species, thus eliminatingthe copper effect. Ammonium ions also inhibited copper absorption,their effectiveness rising with their concentration. Loweringthe nitrate concentration in the medium nullified the coppereffect on flowering in L. paucicostata 6746, and the additionof salicylic acid to the medium also nullified the copper effectin L. gibba G3, both without affecting the absorption of copper. (Received June 7, 1982; Accepted August 27, 1982)     

Floral Inhibition in Lemna paucicostata 6746 Due to Night Interruption

The floral response to various 24-h photoperiodic cycles ofthe short-day plant, Lemna paucicostata 6746 was investigated.No day that had a main photoperiod longer than about 14 h wasable to induce flowers, evidence that the critical day lengthwas ca.14 h. Flowering in the 12-, 9- or 6-h day was inhibitedcompletely by a light pulse inserted daily in the ‘inhibitionzone’ that ranged from about 14 h after the precedingdawn to about 14 h before the next dusk. In the 3- and 1-h days,only the pulse applied 14 h after the dawn completely inhibitedflowering. These results suggest that the daily night interruption prohibitedflowering only when it was linked to either the preceding orthe subsequent main photoperiod to form a skeleton photoperiodwhose length was equal to, or longer than, the critical daylength. Analysis of the floral response to skeleton schedules11:13 and 13:11 on Pittendrigh's model of the photoperiodicclock indicated that light-on circadian oscillation probablyis involved in the day length measurement. ¹ Dedicated to the memory of Dr. Joji Ashida. (Received July 13, 1982; Accepted January 17, 1983)     

Effect of Ferricyanide, Ferrocyanide and KCN on Growth and Flowering in the Short-Day Plant Lemna paucicostata 6746

Flowering in the short-day plant Lemna paucicostata 6746 canbe induced under continuous light by the addition of ferricyanie,ferrocyanide or KCN to M-sucrose medium. Each substance is nearly10 times more effective when the flasks are covered by glassbeakers than when cotton plugs are used. By contrast, when floweringis induced under continuous light by copper or by short-daytreatment, neither flowering nor growth are affected by whetherglass beakers or cotton plugs are used. Ferricyanide, ferrocyanideand KCN are also able to induce long-day flowering when theplants are grown on Msucrose medium in small beakers that areplaced in a covered storage dish that also contains a solutionof one of these compounds. Addition of a KOH trap to the storagedish completely blocks the flowering induced by these compounds.If [¹⁴C]ferrocyanide is added to the storage dish both the M-sucrosemedium and the plants contain significant amounts of radioactivity,the amount of radioactivity being proportional to the floweringresponse. These results indicate that ferricyanide, ferrocyanideand KCN break down to release HCN and that it is the HCN whichis responsible for inducing flowering in L. paucicostata 6746under continuous light. ¹Present address: Department of Biology, Osaka Kyoiku University,Ikeda, Osaka 563, Japan. ²Present address: Institute of Horticulture, The Volcani Center,P. O. B. 6, Bet-Dagan, Israel. (Received January 17, 1983; Accepted March 24, 1983)     

Effect of nitrate concentration in the medium on the flowering of Lemna paucicostata 6746

Addition of copper or tungstate to or exclusion of molybdenumfrom M-sucrose medium induced long-day flowering in Lemna paucicostata6746 provided the medium contained sufficient nitrate. By contrast,ferricyanide, cyanide or silver induced long day flowering evenin nitrate-deficient, M-sucrose medium. (Received August 26, 1977; )     

Effects of Exogenous Amino Acids on Iron Uptake in Relation to their Effects on Photoperiodic Flowering in Lemna paucicostata 6746

The flowering of Lemna paucicostata 6746 grown on 14-h photoperiodwas enhanced by the addition of high concentrations of ironto the medium, which also increased the endogenous iron concentration.The addition of asparagine, aspartate, glutamate, -alanine,glycine or serine to the medium also increased the endogenousiron level, resulting in the promotion of flowering. In contrast,the addition of cysteine, cystine, glutamine, arginine, threonineor phenylalanine lowered the endogenous iron level, resultingin the inhibition of flowering. Glycine and asparagine added to the medium during an inductive96-h dark period did not promote iron uptake and had no effecton flowering, but when added during the subsequent 120-h lightperiod, they promoted both iron uptake and flowering response.The increase in the endogenous iron level seems to favor floraldevelopment rather than induction of photoperiodic floweringof Lemna paucicostata 6746. (Received September 8, 1986; Accepted March 31, 1987)     

Effects of Ferrous and Phosphate Ions on Flowering in Lemna paucicostata 6746 in Diluted Hutner's Medium

In Lemna paucicostata 6746, P₂ (flower induction period) incontinuous darkness was largely extended by dilution of 1/2H1S medium (Hutner's medium supplemented with 1% sucrose) to1/10 strength. The dilution of 1/2 H1S medium to 1/10 strengthremoved completely the extension of P₁ (pre-flower inductionperiod) and P₂ due to a red light pulse given at the 7th h ofthe dark period, which was observed in 1/2 H1S medium. When iron and phosphate ions were added to 1/10 H1S medium upto the same concentration as in 1/2 H1S medium, the extensionof P₂ was removed completely. The red light pulse-induced extensionof P₁ was observed in 1/10 H1S medium only when iron ions wereadded. It is suggested that iron (Fe^{$ $}) and phosphate ionsare important in determining the rate of photoperiodic flowerinduction in L. paucicostata 6746. (Received December 23, 1983; Accepted June 6, 1984)     

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; )     

Phototactic Chloroplast Displacement in the Photosynthetic Mutant, Lemna paucicostata Strain 1073

The photosynthetic mutant, strain 1073, of Lemna paucicostataTorr. (L. perpusilla Hegelm.) which has a block in the electrontransport chain between plastoquinone and cytochrome f is capableof light-induced chloroplast displacement movements. At 8000–14000 lx, chloroplasts of the mutant move from their positionadjacent to the inner periclinal wall of the mesophyll cellsto the anticlinal walls, i.e. along those walls parallel tothe direction of the light. Light does not appear to enhancerespiration of the photosynthetic mutant or of the wild typestrain (6746). These and other results support the idea thatchloroplast displacement in light is not solely the result oflight effects on photosynthesis and respiration. Lemna paucicostata Torr., photosynthetic mutant, phototaxis, chloroplast displacement     

Promotion of Flowering in Lemna paucicostata 6746 (a Short-Day Duckweed) by Cytokinins

Lemna paucicostata 6746 is a short-day plant and flowers inresponse to a single photoinductive cycle. Flowering in thisduckweed could be promoted, under short days, in the presenceof a cytokinin in the nutrient medium. 6-Benzyladenine (BA)was the most effective for promotion of flowering, followedby zeatin and kinetin. Since EDTA itself is promotive for floweringin this plant, the cytokinin effect is best observed in mediumdevoid of EDTA. Nevertheless, in their combined presence floweringwas more as compared to when either of these was individuallypresent. This additive effect on flowering was especially prominentwhen the plants were kept in near-critical photoperiods. Besidesthe increase in percentage of the flowering fronds, floweringis also sustained a little longer in the cytokinin-treated plants.BA, however, did not alter the critical dark period requirement,either in the presence or absence of EDTA. (Received March 26, 1983; Accepted May 6, 1983)     

Effect of 8-Hydroxyquinoline on Flowering and Endogenous Levels of Iron and Copper in Lemna paucicostata, Strain LP6

Lemna paucicostata LP₆, a strain of duckweed isolated locally,does not flower under any photoperiodic schedule when grownin Bonner and Devirian or other media routinely employed invarious laboratories for studies on flowering in Lemnaceae.Flowering in this strain could be induced, however, by 8-hydroxyquinoline(8-HQ)—a well-known copper chelating agent—irrespectiveof the length of the photoperiod. To our knowledge, this isthe first report where a direct induction of flowering in aduckweed by 8-HQ has been observed. Atomic absorption analysisof the plant material revealed that the endogenous level ofcopper is significantly higher in the plants treated with 8-HQ.This is contrary to the general assumption that chelating agentsinfluence flowering of duckweeds by causing a reduction in theuptake of copper ions and making them less available to theplants. (Received May 23, 1983; Accepted July 22, 1983)     

Flower-inducing Activity of Vitamin K in Lemna paucicostata

Vitamins K₁ K₃ and K₅ induced flowering in Lemna paucicostata151, a short-day plant, cultured in 1/10 strength M medium (1/10M medium) under continuous light, and their activity was greatlyintensified by simultaneous application of benzyladenine. Themost active of these was vitamin K₅ L. paucicostata 6746 ismore sensitive to vitamin K₅ than strain 151, but the effectof vitamin K₅ on strain 6746 was not intensified by benzyladenine.The flower-inducing activity of vitamin K₅ was intensified bythe addition of benzoic acid in both strains and by the additionof copper or ferricyanide in Strain 6746, when these chemicalswere added at such low concentrations that they would scarcelyinduce flowering. In strain 6746, vitamin K₅ added to 1/10 M had little effecton flowering under a subcritical photoperiod, while it clearlyinduced flowering under continuous light. In this strain, vitaminK₅ added to full strength M medium, in which this plant wasmore sensitive to short photoperiods than in 1/10 M medium,did not induce flowering even under continuous light, and wasrather inhibitory under short photoperiods. (Received August 14, 1984; Accepted October 16, 1984)