Changes in cotyledon mRNA during floral induction of Pharbitis nil CV. Violet

Floral induction in seedlings of Pharbitis nil Choisy cv. Violet, with one cotyledon removed, was manipulated by applying various photoperiodic treatments to the remaining cotyledon. Populations of polyadenylated RNA from treated cotyledons were examined to identify messages specifically involved in floral induction. The RNA was translated in vitro using a wheat-germ system, and the resulting translation products were analysed by two-dimensional polyacrylamide gel electrophoresis. Substantial qualitative and quantitative differences were found between mRNA from cotyledons of seedlings kept in continuous light (non-induced) and of seedlings given a 16-h dark period (induced). In contrast, inhibition of flowering with a night-break resulted only in one detectable, quantitative difference in mRNA.Abbreviations CL continuous light - kDa kilodalton - NB 16 h darkness+10 min red-light break, 8 h into the dark period - poly(A)⁺ RNA polyadenylated RNA (isolated by binding to a cellulose oligodeoxythymidine affinity column) - SD short day (16 h dark) - SDP short-day plant - SDS sodium dodecyl sulfate     

Preformed mRNA in Cotyledons of Ungerminated Seeds of Cicer arietinum L

Polyadenylated RNA was isolated from total RNA extracted from cotyledons of ungerminated or 18-hour-germinated chick-pea seeds by affinity chromatography on oligo(dT)-cellulose. Both poly(A)-containing RNA fractions exhibited a template activity when assayed in two cell-free translation systems, wheat germ extracts, and nuclease-treated reticulocyte lysates. Translation of preformed mRNA from cotyledons of dry seeds was completely abolished in the presence of several inhibitors of polypeptide chain initiation and also in the presence of the two “cap” analogues m⁷ GTP and m⁷ GMP. The patterns of polypeptides synthesized by translation of poly(A)-containing RNAs from cotyledons of ungerminated or 18-hour-germinated seeds, in the wheat germ system, analyzed by electrophoresis and autoradiography, were similar but not identical. It is concluded that cotyledons of dry Cicer arietinum L. seeds contain preformed mRNA.     

Accumulation of Phenylpropanoids in the Cotyledons of Morning Glory (Pharbitis nil) Seedlings during the Induction of Flowering by Low Temperature Treatment, and the Effect of Precedent Exposure to High-Intensity Light

Extracts from the cotyledons of seedlings of Pharbitis nil strain‘Violet’ cultured at low temperature, which inducestheir flowering even in continuous light, with or without precedentexposure to high-intensity light, which shortens the periodof low temperature required for flowering, were analyzed byHPLC for substances correlating with the flower-inducing process.The content of two phenylpropanoids were found to increase duringthe low-temperature, and were identified as 3-O-feruloylquinicacid and dehydrodiconiferyl alcohol-13-O-ß-D-glucoside.The increase was more rapid in the cotyledons exposed to high-intensitylight before the low-temperature. This suggests that the accumulationof these compounds is correlated to the promotive effect ofhigh-intensity light on the flower-induction by low temperature. (Received March 7, 1994; Accepted April 2, 1994)     

Effects of Elevated Carbon Dioxide Concentration in the Dark on the Growth of Soybean Seedlings

Previous work has shown that elevated carbon dioxide (CO₂) concentrationsin the dark reversibly reduce the rate of CO₂ efflux from soybeans.Experiments were performed exposing soybean plants continuallyto concentrations of 350 or 700 cm³ m^-3 for 24 h d^-1, or to350 during the day and 700 cm³ m^-3 at night, in order to determinethe importance of the reduced rate of dark CO₂ efflux for plantgrowth. High CO₂ applied only at night conserved carbon andincreased dry mass during initial growth compared with the constant350 cm³ m^-3 treatment. Long-term net assimilation rate was increasedby high CO₂ in the dark, without any increase in daytime leafphotosynthesis. However, leaf area ratio was reduced by thedark CO₂ treatment to values equal to those of plants continuallyexposed to the higher concentration. From days 14-21, leaf areawas less for the elevated night-time CO₂ treatment than foreither the constant 350 or 700 cm³ m^-3 treatments. For the days7-21-period, relative growth rate was significantly reducedby the high night CO₂ treatment compared with the 350 cm³ m^-3continuous treatment. The results indicate that some functionallysignificant component of respiration was reduced by the elevatedCO₂ concentration in the dark.Copyright 1995, 1999 AcademicPress Glycine max L. (Merr.), carbon dioxide, plant growth, respiration     

Changes in Nuclear DNA Content, Cell and Nuclear Size, and Frequency of Cell Division in the Cotyledons of Brassica napus L. during Embryogenesis

Cellular behaviour was examined during embryogenesis in Brassicanapus to test whether or not polyploidy occurs in the cotyledonsduring the phase of oil deposition. Nuclear DNA content, nuclearand cell size, and the mitotic index were measured in the cotyledonson various days post anthesis (dpa). In squashed monolayersfrom 15 dpa cotyledons, a polyploid (>5C) population wasdetected together with a substantial number of cells in G2 (4C).Nuclear volume was measured on sectioned tissues and, at 15dpa, the range of values from the cotyledons (40–500 *m³)contrasted with that in the vestigial suspensor and endosperm(50–> 600 µm³). At 15 dpa the nuclear volumedata suggest that whilst cells in the cotyledons were in Gland G2 many endosperm and suspensor cells were polyploid. Thus,polyploidy observed in the squashed monolayers was probablydue to contaminating endosperm/suspensor cells. At 25 and 35dpa, polyploidy was not detected; all cells were in Gl (2C)and cell area increased. The mitotic index peaked at 20 dpabefore declining and given the narrower distribution of nuclearvolumes at 25 and 35 dpa (50–300 µm³), these dataare consistent with cell arrest in Gl. Thus, polyploidy wasnot detected in the cotyledons of B. napus which differs fromwhat is known about cellular development in legume cotyledons. Key words: Brassica napus L., DNA, nuclear volume     

Photodynamic Effects of Rose Bengal on Senescent Flax Cotyledons

Senescent cotyledons, excised intact from 6-d-old Flax (Linumusitatissium) seedlings, were treated with Rose Bengal, a photo-sensitizingagent. In continuous light, chlorophyll breakdown was increasedfrom 44% in untreated cotyledons to 73% in Rose Bengal-treatedcotyledons. Depletion of carotenoid and linolenic acid was enhancedby 25% and 39% respectively over the same period. After 48 hthe decline in uncoupled photosynthetic electron transport wasincreased from 58% to 82% in isolated chloroplasts. Over thesame period CO₂ exchange was greatly inhibited in treated cotyledons.Rose Bengal had little effect on dark incubated cotyledons,suggesting that singlet oxygen generated by photo-sensitizedreactions was responsible for the enhanced deterioration ofstructure and function of senescent cotyledons and the primesite of action was the photosynthesis apparatus. These photodynamiceffects are discussed in relation to the possible involvementof toxic oxygen species in the senescence of leaf tissue. Key words: Senescence, Photodynamic damage, Rose Bengal, Singlet oxygen     

Some Effects of Potassium Deficiency on Seedling Development

Seedlings with leaf-like photosynthetic cotyledons such as Cucumissativus, Sinapis alba and Trifolium pratense have higher growthrates and a greater requirement for an external supply of potassiumthan hypogeal species (Pisum sativum, Vicia faba) or epigealspecies with very fleshy cotyledons (Phaseolus vulgaris, Lupinusalbus, Lupinus angustifolius). The initial stages of seedlingdevelopment in the latter two groups involve mainly transferof reserves from cotyledons to axis and a relatively small increasein total seedling dry weight takes place during the first 2to 3 weeks. Potassium may be drawn either from the supply already presentin the cotyledons or from an external source. Species with expandingphotosynthetic cotyledons utilize the reserve potassium duringcotyledon development and it is not transported to the epicotyl.Thus, in these species an external supply of potassium is essentialfor full development of the photosynthetic system and the roots.In species whose cotyledons are not major photosynthetic organsthe reserve potassium is transported from the cotyledons tothe developing axis. In these species culture in a nutrientsolution deficient in potassium or in distilled water increasesroot dry weight but reduces plumule weight (although to a lesserextent than in species with leaf-like cotyledons). In general,the response of seedlings to culture in a nutrient solutiondeficient in potassium was very similar to culture in distilledwater only. Thus, although all the species studied had similar levels ofpotassium in the seed (µg potassium/mg dry weight) itssite of utilization differs. It appears that seedlings withleaf-like cotyledons are more susceptible during early seedlinggrowth to deficiencies in external factors, such as potassium,than are species with fleshy cotyledons.     

Translatable mRNAs for Chloroplast-Targeted Proteins in Detached Radish Cotyledons during Senescence in Darkness

A protein-import system prepared with isolated chloroplastswas used to monitor changes in levels of mRNAs for chloroplast-targetedproteins during dark-induced leaf senescence. Biologically activechloroplasts were isolated from young (9-day-old) and aged (14-day-old)radish cotyledons. Poly(A)⁺-RNA was prepared from radish cotyledonsthat had been detached from seedlings and placed in darknessto accelerate senescence. The RNA was translated in a wheatgerm system, and the products were added to an import systemprepared with chloroplasts from young cotyledons. Electrophoreticanalysis of the imported proteins suggested that most chloroplast-targeted proteins decreased in abundance during dark treatmentof cotyledons. However, the relative abundance of 38 stromaland three thylakoid proteins increased transiently or continuouslyamong the products of translation of RNA isolated during thecourse of senescence. The efficiency of the uptake of precursorproteins by chloroplasts isolated from aged cotyledons was lowerthan that by chloroplasts from young tissue. The chloroplastsfrom aged cotyledons more efficiently imported at least onestromal protein and one thylakoid protein than chloroplastsfrom the young tissue. The relative abundance of these two proteinsincreased among the products of translation of RNA from senescingcotyledons when tested in the uptake system with chloroplastsfrom young cotyledons. These results suggest that some nucleargenes for chloroplast-targeted proteins are expressed in senescingcotyledons more efficiently than in young tissue, and that themachinery for import of proteins into chloroplasts changes duringaging of the tissue to allow more efficient import of certainproteins that may be responsible for the senescence of the chloroplasts. ¹Present address: Kihara Institute for Biological Research,Yokohama City University, Mutsukawa 3-122-20, Minami-ku, Yokohama,232 Japan     

Effect of Sink-Source Manipulations on the Photosynthetic Rate and Carbohydrate Content of Cucumber Cotyledons

Mayoral, M. L., Plaut, Z. and Reinhold, L. 1985. Effect of sink-sourcemanipulations on the photosynthetic rate and carbohydrate contentof cucumber cotyledons.-J. exp. Bot. 36 1551–1558. The photosynthetic rate of cucumber cotyledons (Cucumis sativuscv. Dahla) reached a maximum value of 12 mg dm^–2 h^–1,10 d after emergence. In 12-d-old seedlings removal of one cotyledondoubled the CO₂ fixation rate of the other, as observed 3 dafter treatment. When the primary leaf was removed, the photosyntheticrate of the cotyledons was decreased by 33%. At this stage ofgrowth elimination of the roots as a sink for assimilates bygirdling the hypocotyl affected neither the photosynthetic ratenor the carbohydrate content of the cotyledons. By contrast,in 18-d-old seedlings removal of the first leaf brought abouta 42% increase in the photosynthetic rate of the cotyledons.The simultaneous removal of the first leaf and one cotyledondoubled the rate of CO₂ fixation of the remaining cotyledon.Girdling the hypocotyl lowered the photosynthetic rate of thecotyledons by 73%. In both 12- and 18-d-old seedlings a decreaseor increase in the sink-source ratio was correlated with anincrease or a decrease respectively in the carbohydrate contentof the cotyledons. The stomatal resistance of the cotyledonswas not affected by any of the treatments. The effect of sink-sourcemanipulations on photosynthesis and on the level of carbohydratespresent in the cotyledons was more evident in those seedlingsgrowing under high light intensity (580 µE m^–2 s^–1),than in those exposed to 300 µE m^–2 s^–1 Key words: Sink-source relationship, cotyledons, photosynthesis     

Jasmonates Inhibit Flowering in Short-Day Plant Pharbitis nil

The role of jasmonates in the photoperiodic flower induction of short-day plant Pharbitis nil was investigated. The plants were grown in a special cycle: 72 h of darkness, 24 h of white light with lowered intensity, 24-h long inductive night, 14 days of continuous light. At 4 h of inductive night the cotyledons of non-induced plants contained about two times the amount of endogenous jasmonates (JA/JA-Me) compared to those induced. A 15-min long pulse of far red light (FR) applied at the end of a 24-h long white light phase inhibited flowering of P. nil. The concentration of jasmonates at 2 and 4 h of inductive night in the cotyledons of the plants treated with FR was similar. Red light (R) could reverse the effect of FR. R light applied after FR light decreased the content of jasmonates by about 50%. Methyl jasmonate (JA-Me) applied to cotyledons, shoot apices and cotyledon petioles of P. nil inhibited the formation of flower buds during the first half of a 24-h long inductive or 14-h long subinductive night. Application of JA-Me to the cotyledons was the most effective. None of the plants treated with JA-Me on the cotyledons in the middle of the inductive night formed terminal flower buds. The aspirin, ibuprofen and phenidone, jasmonates biosynthesis inhibitors partially reversed the effect of FR, stimulating the formation of axillary and terminal flower buds. Thus, the results obtained suggests that phytochrome system control both the photoperiodic flower induction and jasmonates metabolism. Jasmonates inhibit flowering in P. nil.     

Flower-Inducing Activity of Water Extracts of Various Plant Species, in Particular Pharbitis nil

The crude water extracts of leaves of many plant species belongingto Spermatophyta and some belonging to Bryophyta induced floweringof Lemna paucicostata 151 (PI51) under continuous light, atthe concentrations equivalent to 0.1 to 10 mg fr wt leaf per10 ml culture medium (mg fr wt/10 ml). The extract of Salvinia(Pterydophyta) added together with the extract of Lemna at aconcentration lower than that necessary to cause flowering alsoinduced flowering. The activity of the water extracts of someplants varied considerably from experiment to experiment dueto unknown factors, but the extracts of Pharbitis nil strainViolet, a sensitive short-day plant, always showed a high activity,as did the extracts of Lemna paucicostata reported previously. The extract of Pharbitis cotyledons induced flowering of P151even at 0.3 mg fr wt/10 ml, and significantly promoted floweringof L. paucicostata 441 and 6746 at 1–3 mg fr wt/10 ml.Ex-udate from the cuttings of the seedlings was also active.However, neither the activity of the water extract nor thatof the exudate could be correlated with photoperiodic floralinduction. On the other hand, the extract of leaves or cotyledonshad higher activity (on a fr wt basis) than that of other organs,and the activity of the extract of cotyledons changed with theirage roughly in parallel with their photoperiodic sensitivity. (Received April 17, 1989; Accepted August 10, 1989)     

A Comparative Study of Germination Responses to High Irradiance Light

Seeds of 19 native British herbaceous species (14 grasses andfive forbs) were exposed to white light at three photon fluencerates: high (19–2 mol m^–2 d^–1), medium (9·6mol m^–1 d^–1) and low (2·3 mol m^–2 d^–2) These photon doses have been found by previous workers to inhibitgermination in several species. High and low photon doses wereapplied only as continuous light, but the medium dose was appliedas both continuous light and as a 12 h light/12 h dark photoperiod.All four treatments, plus a dark control, were carried out at15 °C; the high and low doses were also applied with a dailyalternation of 10/20 °C. The majority of species (15) fell into one of two groups. Inseven species germination was relatively high and consistentacross all treatments, including darkness; in the other eightspp germination was inhibited only in darkness. Mostly thesedata confirmed published results for the same species In contrast in Agrostis capillaris and A. stolonifera germinationwas high only at alternating temperatures, irrespective of photondose, but was also slightly promoted by a constant temperaturecombined with light/dark alternations. Only in Bromus sterilisand B. ereclus was germination inhibited by light, in B. erectusat all photon doses and in B. sterilis only at the highest photondose These results suggest that inhibition of germination by highirradiance light is not widespread among native British species Aira caryophyllea, Arrenatherum elatius, Festuca ovina, F. rubra, Hordeum murinum, Milhim effusum, Silene dioica, Achillea millefolium, Brachypodium syhaticum, Digitalis purpurea, Holcus lanatus, Leucanlhemum vulgare, Phleum pratense, Poa trivialis, Taraxacum officinale, Agrostis capillaris, A. stolonifera, Bromus erectus, B. sterilis, seed, dormancy, germination, light, High irradiance reaction, alternating temperatures, photoperiod     

The Effects of Temperature and Light Intensity on Embryo Numbers in Wheat Doubled Haploid Production through WheatxMaize Crosses

Triticum aestivumxZea mayscrosses are now widely used in theproduction of wheat doubled haploids to produce homozygous lines.Seasonal effects are known to influence the number of haploidembryos produced through wheatxmaize crosses, but the effectsof temperature and light have not been quantified. This studyinvestigated the effect of temperature and light intensity onhaploid embryo production. New Zealand wheat cultivars weregrown in a glasshouse until booting when they were transferredto growth cabinets at three temperatures (day/night; 17/12,22/17 or 27/22 °C at an irradiance of 250 µmol m^-2s^-1PAR).In another experiment, wheat lines were transferred to a growthcabinet at one of three light intensities (300, 500 or 1000µmol m^-2s^-1PAR at 22/17 °C day/night, with a photoperiodof 16 h). The temperature and light intensity at which pollinationswere made and subsequent fertilisation and embryo developmentoccurred, significantly (P<0.01) influenced the frequencyof haploid embryo production. The optimal temperature for embryorecovery was 22/17 °C. The greatest number of embryos wasproduced at a light intensity of 1000 µmol m^-2s^-1. Thesefindings will result in improvements in the overall efficiencyof the wheatxmaize system for wheat doubled haploid production.Copyright1998 Annals of Botany Company Intergeneric crossing, temperature, light intensity,Triticum aestivum,wheat,Zea mays,maize.     

The recalcitrant plant species, Castanospermum australe and Trichilia dregeana, differ in their ability to produce dehydrin-related polypeptides during seed maturation and in response to ABA or water-deficit-related stresses

In constrast to seeds of orthodox species, those of recalcitrantspecies do not acquire desiccation tolerance during their developmentand are shed from the parent plant at high water contents. Dehydrinproduction in seeds of recalcitrant species was examined duringdevelopment and germination, in response to abscisic acid (ABA),and following the imposition of various water-deficit-relatedstresses, including desiccation, water stress, high salt, highosmolarity, and low temperature. Two tropical species exhibiteda differential capacity to produce dehydrin-related proteinsduring seed maturation. Dehydrins were present in axes and cotyledonsof Castanospermum australe seeds during mid-maturation and atmaturity. In Trichilia dregeana, no dehydrin-related polypeptideswere detected in the mature seed. During the development ofC. australe seeds, the nature of the dehydrin related polypeptidesaccumulated in the cotyledons and axis changed and new polypeptideswere detected in the mature seeds that were not present duringmid-maturation. The dehydrins present in cotyledons of matureseeds (31, 37 and 40 kDa) were still detectable after germination(i.e. in untreated seedlings). These dehydrins became less abundantin the cotyledons of C. australe seedlings following ABA andall stress treatments except cold, although most of the dehydrinswere still detectable. An exception was the desiccation-treatedseedlings, in which no dehydrins were detected. In the rootsof C. australe seedlings, no dehydrins were found after germinationnor were they induced in the root by ABA or any of the stresstreatments imposed on seedlings. Seedlings of Trichilia dregeanadid not produce dehydrins in the roots or cotyledons when exposedto ABA or water-deficit-related stresses. Key words: Dehydrin, ABA, desiccation, recalcitrant, seed     

Further studies on the diurnal rhythm of uridine incorporation into RNA in the long-day duckweed, Lemna gibba G3

Using the long-day duckweed Lemna gibba G3, the changes in theactivities of RNA synthesis in isolated nuclei and chloroplastsand of the reaction prerequisite for the incorporation of exogenousuridine into RNA were examined. When the duckweed was exposedto either a light-dark cycle or to continuous light, the activityof RNA synthesis in the nuclear and chloroplast fractions changeddiurnally and reached its highest levels during the night phase.The changes coincided with uridine incorporation into RNA invivo. However, the amount of radioactive uridine taken up intothe acid-soluble fraction remained unchanged during the wholeday. The proportion of radioactivity incorporated into phosphorylateduridine compounds as well as UTP+UDP to the radioactivity inthis fraction remained constant. Thus, the diurnal rhythm ofuridine incorporation into RNA was related to the diurnal rhythmof RNA synthesis in isolated nuclei and chloroplasts. The loweractivity of uridine incorporation into RNA under continuousdarkness may be determined by the activity of RNA synthesisin nuclei and chloroplasts as well as the uptake rate of uridineinto the duckweed cells, not by the activity of its phosphorylation. (Received August 30, 1977; )     

Circadian Timekeeping for the Photoperiodic Control of Budset in Picea abies (Norway Spruce) Seedlings

Romanian populations of Norway spruce are induced to set terminal buds by four inductive cycles of 8 h light/16 h darkness. To distinguish between circadian and hourglass timekeeping for the photoperiodic control of budset, seedlings were raised in continuous light at 300 µmol m^-2s^-1 at 20°C for 10 weeks. They were then exposed to an extended night regime consisting of three cycles of 8 h light/40 h dark with 4-h or 1-h nightbreaks (120 µmol m^-2s^-1) applied to groups of plants at intervals during the extended night. Following a final cycle of 8 h light/16 h dark to maximize budset, the plants were transferred to continuous light. Budset was delayed when the night-break was applied close to the critical nightlength (CNL) of 6–7 h or about 22–23 h later in the extended night, consistent with circadian rather than hourglass timekeeping. Confidence intervals were calculated for the times to maximum effect of the night-breaks.     

Inhibition of Zeatin-Induced Growth of Cucumber Cotyledons by Sulfite Ions

The inhibitory effects of sulfite ions on zeatin-induced cellexpansion in cotyledons excised from dark-grown seedlings ofcucumber (Cucumis sativus L.) were examined. With 50 µMzeatin the growth rate in white light was about twice that ofthe control. Addition of Na₂SO₃ in the growth medium inhibitedthe zeatin-induced growth of cotyledons. Zeatin-treatment increasedthe osmotic potential in cell sap of cotyledons, while sulfitedecreased it. These treatments had no significant effect onpotassium concentration. Sulfite inhibited the zeatin-inducedincrease in contents of fructose and glucose, but did not affectsucrose content. The relative contents of non-cellulosic constituentsof cell walls fell with the advance of culture. This decreasewas repressed by sulfite, indicating that inhibition of expansiongrowth in cucumber cotyledons by sulfite ions was the resultof alterations in the cell wall structure due to changes inthe cell wall metabolism. (Received June 12, 1984; Accepted October 24, 1984)     

Effect of seed germination on levels of tRNA aminoacylation

In ungerminated seeds of Lupinus luteustRNAs are aminoacylated 10% or less depending on species of tRNA. The levels of tRNA aminoacylation for specific tRNAs increase steadily during seed germination. Specific tRNAs in cotyledons and axes of 3-day-old seedlings are aminoacylated to a similar extent. No significant changes are observed in the tRNA population during germination.