Sensitivity to Humidity and Temperature in Darkness of the Photoinduction of S-Adenosyl-L-Methionine Decarboxylase Activity in Leaves of Pharbitis nil

The activity of S-adenosyl-L-methionine decarboxylase (SAMD)in leaves of Pharbitis nil increased maximally at "lights-on"after growth in darkness for 16 h at 21 to 25°C and 30%to 40% relative humidity. Under other condition, less SAMD activitywas induced. The photoinduction of SAMD was also observed inleaves of many other plant species. (Received December 14, 1994; Accepted April 6, 1995)     

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     

Dark-Induced Accumulation of mRNA for a Homolog of Translationally Controlled Tumor Protein (TCTP) in Pharbitis

A cDNA encoding a homolog of human translationally controlledtumor protein (TCTP) was isolated from cotyledons of the short-dayplant Pharbitis nil cv. Violet. The level of the correspondingmRNA increased gradually during darkness. This increase wasinhibited by end-of-day exposure to far-red light. (Received October 30, 1997; Accepted January 7, 1998)     

Guanylyl cyclase activity during photoperiodic flower induction in <Emphasis Type="Italic">Pharbitis nil</Emphasis>

It is known that the level of cGMP is modulated in plant cells in response to a number of stimuli but intracellular events dependent on cGMP metabolism are not clear. Guanylyl cyclases (GCs) are enzymes which are responsible for synthesis of cGMP in eukaryotic and prokaryotic cells. To collect evidence for the participation of cGMP in light signal transduction we isolated enzyme with guanylyl cyclase activity from Pharbitis nil and analysed its level and activity during photoperiodic flower induction. Soluble proteins were isolated from seedlings of a model short-day plant P. nil, partly purified and identified by in vivo and in vitro enzyme assay. In green plants enzyme activity amounted to 484 nmol cGMP/min/mg protein, whereas in etiolated plants it was three times lower (158 nmol cGMP/min/mg protein). Analyse cyclase consists of a single polypeptide of Mr 40 kDa. In order to determine if changes in guanylyl cyclase activity occurred in response to a long, inductive night, we measured enzyme activity in 4-h intervals and observed its increase at 4, 8 and 16 h of darkness. This pattern also fits well with changes in the endogenous cGMP level during a 16 h long flower inductive night. Immunocytochemical analysis confirmed these observations and revealed that changes in the GC level during light/dark conditions appeared. During 16 h long inductive night the strongest signal was observed in cotyledons after 4 and 16 h of the darkness. A high level of fluorescence was generally distributed in mesophyll, however, it was also observed in guard cells. Staining was apparently absent in the veins and cotyledon body. Furthermore, the location inside the cell was analysed. The protein was immunolocalized preferentially in the cytosol, chloroplasts and peroxysomes. Taken together, these data demonstrate in Pharbitis nil the presence of an enzyme which is able to convert GTP to cGMP. Because its level and activity are affected by light we believe that GC/cGMP play a substantial role in light/dark dependent process in plants, such as photoperiodic flower induction.     

The Inhibition of Flowering by Non-Induced Cotyledons of Pharbitis nil

Inhibitory effects on flowering of a non-induced cotyledon havebeen examined in Pharbitis nil seedlings. The photoperiodicinduction of one cotyledon was accomplished by wrapping it inaluminium foil for 13 to 15 h while the seedling remained inthe light. The presence of the other cotyledon in the lightblocked this inductive stimulus. The timing of its inhibitoryeffect suggested that its action was to block the expressionof the inductive stimulus, presumably at the shoot apex. Byvarying the area of the non-induced cotyledon parallel inhibitoryeffects were shown on export of stimulus and of ¹⁴C-labelledassimilate to the apex from the induced cotyledon. Thus, partof the inhibition was by interference with assimilate/stimulusco-transport in the phloem. However, an additional inhibitoryeffect was also evident and for this second component therewas no relationship between assimilate and stimulus transport.This latter inhibition was generated by brief light interruptionsof darkness given to one cotyledon only whilst the other waswrapped. The control treatment, removal of the unwrapped cotyledon,did not alter flowering compared to seedlings with intact, darkenedcotyledons. Thus, these studies show that the brief night interruptionsacted to trigger a photoperiodically sensitive inhibitor notto block induction. The implications of these findings are discussedin relation to models of time measurement in the photoperiodiccontrol of flowering. (Received March 20, 1989; Accepted November 16, 1989)     

Phytochrome-regulated Expansion of Mustard (Sinapis alba L.) Cotyledons

In developing mustard (Sinapis alba L.) seedlings continuousred light acting via the agency of phytochrome stimulates therate of expansion of cotyledons. Although phytochrome actionon cotyledon expansion is evident only after 36 h from sowing,the photoresponse escapes from reversibility at about 15 h fromsowing. The time lag of 21 h between loss of photoreversibilityand the onset of photoregulated cotyledon expansion indicatesthe existence of long-lived components in the phytochrome-triggeredsignal chain. Phytochrome-regulated cotyledon expansion doesnot require the involvement of photosynthesis, as applicationof SAN 9789, an inhibitor of chloroplast biogenesis, did notaffect cotyledon expansion. The role of turgor pressure-relatedcellular parameters such as osmotic potential () cell wall extensibility(m), hydraulic conductivity (L) and yield threshold (Y) forcell expansion were examined during photoregulated cotyledonexpansion. Using the general equation of cell growth dv/dt =[Lm/(L+m)]( - Y), where dv/dt is the rate of volumetric growth,it was demonstrated that the light-mediated cotyledon expansionresults from an increase in cell wall extensibility (m). Theseresults are discussed in relation to the photoregulation ofcotyledon expansion. Key words: Cell wall extensibility, growth, Sinapis alba L., phytochrome     

Flowering of Pharbitis nil is controlled by an endogenous semidian rhythm

Abstract Flowering of Pharbitis nil after an inductive dark period is greatly influenced by far-red (FR) irradiation during the preceding light period. The response to FR is rhythmic in otherwise constant conditions, and the period of the oscillation is approximately 12 h (i.e. semidian). The rhythm also appears to operate under daily light-dark cycles. The expression of this novel rhythm depends on the time from the beginning of FR pretreatment to the onset of the inductive dark period. The cotyledons are the site of response to both the pretreatment and inductive darkness, and both these conditions must be perceived by the same cotyledon.     

Effects of Number of Photoperiodic Cycles on Induction and Development of Flowers and Fruits in Corchorus olitorius L. (var. oniyaya Epen.)

Plants of Corchorus olitorius, a short-day plant, were subjectedto varying numbers of short-day cycles before transfer to longdays. Treatments started after germination of the seeds at thetime of cotyledon release. Four short-day cycles (10 h naturaldaylight followed by 14 h darkness) were sufficient to induceflowering in all plants. The number of flowers and fruits producedon a plant increased as the number of short-day cycles was increasedfrom three to 30. Plants given three to seven short-day cyclesproduced flowers on the main stem only but when plants weremaintained in short days for longer periods, flowers were alsoproduced on the branches. The growth in d. wt of fruits wasgreatest when plants were maintained in short days throughout. Corchorus olitorius L., flowering, fruit development, photoperiodism     

Two calcium dependent protein kinases are differently regulated by light and have different activity patterns during seedling growth in <Emphasis Type="Italic">Pharbitis nil</Emphasis>

In this study using biochemical approaches we identified two calcium-dependent protein kinases (CDPKs) named PnCDPK52 and PnCDPK56 in soluble protein extracts from seedlings of Pharbitis nil. Both enzymes phosphorylated the specific substrate histone III-S in the presence of Ca²⁺ and cross-reacted with antibodies against the CDPK. PnCDPKs exhibited quite different activity and protein levels during germination and successive stages of seedling growth. PnCDPK52 protein level was high in seeds and during germination, whereas PnCDPK56 increased in the next stages of seedling growth, being the dominant enzymes in mature seedlings, of the light- and dark-grown plant. In all cases both activity and accumulation of protein PnCDPK56 was higher in dark grown plants whereas exposure to light reduced both factors. When etiolated cotyledons were exposed to light, the activity of PnCDPK56 was reduced to the basal level within 5 h. Conversely, increasing activity of PnCDPK56 in cotyledons of green plants shifted to darkness was extremely rapid, reaching the maximum level after just 1 h of darkness and then gradually decreased. Further lengthening of the darkness to 16 h resulted in a strong increase in activity at 12 h. These data indicate that at least two isoforms of CDPK are involved in germination and seedling growth of P. nil. The differences in PnCDPKs strongly argue for the pleiotropic role of these isoforms. It seems that PnCDPK52 is associated with the germination process and PnCDPK56 with seedling growth. Moreover it suggests that activity of PnCDPK56 is controlled by light via the photoreceptor-dependent pathway.     

The Relationship between Sugar and Amino Acid Export to the Phloem in Young Wheat Plants

The relationship between amino acid and sugar export to thephloem was studied in young wheat plants (Triticum aestivumL. ‘Pro-INTA, Isla Verde’) using the EDTA-phloemcollection technique. Plants grown with a 16 h photoperiod showeda rapid decrease in the concentration of sugars and amino acidsin the phloem exudate from the beginning of the dark period.When plants grown with a 16 h photoperiod were kept in the darkfor longer than 8 h the free amino acid content in leaves andexudate (on a dry weight basis) increased continually throughoutthe 72 h of darkness. During the first 24 h of darkness thesugars in the phloem exudate decreased to 30% of the initialvalue, and returned to the control level when plants were returnedto light. When plants grown under low light intensity for 10d were transferred to high light intensity, they showed an increasein leaf sugar content (dry weight basis) after 3 d but therewere no differences in leaf free amino acid content (dry weightbasis) compared to low-light plants. The sugar concentrationin the phloem exudate was increased by higher light intensities,but there was no difference in the amino acid concentrationof the phloem exudate, and thus the amino acid:sugar ratio inthe phloem decreased in the high-light plants. The present resultssuggest that amino acids can be exported to the phloem independentlyof the export of sugars. Copyright 1999 Annals of Botany Company Sugar exudation, amino acid transport, nitrogen, phloem, transport, wheat, Triticum aestivum L.     

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.     

Methyl jasmonate-dependent senescence of cotyledons in <Emphasis Type="Italic">Ipomoea nil</Emphasis>

Jasmonic acid methyl ester (JAMe) has been recently shown to play a crucial role in many physiological processes. In this paper, we focused on cotyledon senescence in Ipomoea nil and revealed that JAMe and darkness are the main factors stimulating the process examined. What is more, we showed that mefenamic acid (a jasmonate biosynthesis inhibitor) reverses the stimulatory effect of darkness on senescence. In plants growing under dark conditions, stimulation of JASMONIC ACID CARBOXYL METHYLTRANSFERASE (InJMT) expression and, consequently, an increase in JAMe content, have been observed. In turn, the level of jasmonic acid (JA) gradually decreased. Moreover, dark-grown seedlings demonstrated a lower PSII functional activity and a reduced chlorophyll content and autofluorescence. All of these data suggest that JAMe is a signal molecule controlling the senescence of cotyledons in I. nil.     

The Interactive Effects of Atmospheric Carbon Dioxide and Light on Stem Elongation in Seedlings of Four Species

Four species,Sinapis albaL.,Medicago sativaL.,Gypsophila paniculataL.andPicea abies(L.) Karsten, were grown in three light regimes:darkness, low light (25 µmol m^-2s^-1for 10 min d^-1) andhigh light (120 µmol m^-2s^-1for 12 h  d^-1) and fourlevels of carbon dioxide: 0, 350, 700 and 1400±50 µll^-1. Germination was not affected by any of the treatments.The effects of carbon dioxide on stem elongation were identicalin low and high light: stem length increased at a decreasingrate with level of carbon dioxide in all species. Level of carbondioxide also affected stem elongation in complete darkness,but the pattern was more complex and varied among species. Totalweight did not vary with level of carbon dioxide to any significantextent in either darkness or low light, but increased with levelof carbon dioxide at high light in all four species. Due tothe absence of any effect of carbon dioxide on growth in darknessand low light, we suggest the effects of carbon dioxide on stemelongation are independent of effects on growth and may be dueto a direct interaction with developmental processes. In contrast,level of carbon dioxide had little effect on allocation patternsin the dark and low light experiments, but had marked effectsin high light. Therefore, the effect of carbon dioxide on allocationwas probably due to the effects of carbon dioxide on growthrather than to any direct interaction between carbon dioxideand development. An understanding of the mechanisms by whichcarbon dioxide affects development may help us understand theoften variable effects of carbon dioxide upon plants.Copyright1998 Annals of Botany Company Sinapis albaL.;Medicago sativaL.;Gypsophila paniculataL. andPicea abies(L.) Karsten; elevated carbon dioxide; stem elongation; germination; allocation; phytochrome.     

Inhibition of photoperiodic floral induction in Pharbitis nil by ethylene

Application of ethylene at 100 ppm or higher completely inhibitedflowering in Pharbitis nil when made during an inductive darkperiod. Exposing plants to ethylene before or after the inductivedark period produced only slight or almost no inhibition. Ethylenewas effective when it was applied only to a cotyledon, but wasineffective when applied only to a receptor bud. Ethylene hadno effect on translocation of the floral stimulus. Ethylene-treatedcotyledon did not transfer any flower inhibiting entity. Thus,ethylene is considered to inhibit the induction process(es)in cotyledons. Except for an initial temporary cotyledon epinasty, ethylenetreatment had no effect on the subsequent growth and vigor ofplants. This temporary cotyledon epinasty disappeared withinthe next 24 hr. (Received May 4, 1972; )     

Changes in the Free Amino Compounds in Young Tomato Plants in Light and Darkness with particular Reference to {gamma}-Aminobutyric Acid

Tomato plants were grown to the five-leaf stage under uniformconditions in a growth room with a daily light period of 15h. Plants were sampled at intervals through 24 h periods andthe free ninhydrin-positive compounds determined in roots, bleedingsap, stems and shoots (mainly leaves), using ion-exchange columnchromatography and a lithium-buffer separation system. The compoundspresent and their range of concentrations are given for twooccasions: after illumination for 8 hand after 5 h of darkness. Data for -aminobutyric acid (GAB), glutamic acid, glutamine,alanine, aspartic acid and ammonia are summarized graphicallyfor all occasions and for all parts of the plant; asparaginefor sap only. The data were examined for correlations betweenthese substances for both light and dark conditions. Relative amounts of free acids were: root glutamine> glutamicand GAB > aspartic > alanine; bleeding sap glutamine >asparagine > GAB > aspartic> alanine; stem glutamine> glutamic > GAB and aspartic > alanine; shoot (leaf)GAB and glutamine > aspartic > alanine and glutamic. Patternsof change were as follows: in the root GAB and glutamic weresimilar and unlike glutamine; alanine did not change;sap ammonia,GAB and alanine were parallel, glutamine was similar to theseonly in light; in the stem glutamine and glutamic tended toaccumulate in parallel in light, but GAB did not; in the shoot(leaf) GAB and glutamine were similar except that the formeraccumulated more rapidly in the initial light period; glutamicacid and alanine were similar to each other but distinct fromGAB and glutamine. The relatively large amounts of GAB in tomato plants and themagnitude of the changes occurring in light and darkness seemindicative of its importance as a temporary storage productfor protein amino acids, but the factors controlling accumulationand utilization in different parts of the plant are unknown.     

不同催青方式对二化性家蚕过氧化氢酶基因表达的影响

25℃明催青和15℃暗催青分别诱导二化性家蚕Bombyx mori 产滞育性卵和非滞育性卵。此前我们的研究表明, 上述催青处理的二化性家蚕H₂O₂水平存在显著差异。过氧化氢酶（catalase, CAT）是昆虫清除H₂O₂的关键酶。为了进一步明确家蚕滞育过程中H₂O₂代谢的调控机制, 用RT-PCR测定了上述两种催青处理对二化性家蚕CAT基因表达的影响。结果表明：25℃明催青显著提高了滞育诱导和决定阶段的CAT mRNA 水平和CAT活性。滞育性卵的CAT mRNA水平在产后24 h形成峰值, 在72 h后消失; CAT活性在96 h前上升, 120 h后保持于低水平。非滞育性卵的CAT mRNA水平和CAT活性都随着胚胎发育而上升。可见, 25℃明催青诱导二化性家蚕子代滞育可能是通过影响CAT基因表达来调节H₂O₂水平。     

Molecular and cytological characterization of ZTL in <Emphasis Type="Italic">Ipomoea nil</Emphasis>

The ZEITLUPE (ZTL) protein is involved in the control of circadian period, hypocotyl elongation and flowering time in Arabidopsis thaliana. The aim of the present work was the identification of the InZTL gene and localization of its mRNA in the model short-day plant Ipomoea nil. The deduced InZTL protein of 622 amino acid residues contained a LOV domain at the N-terminal part, followed by an F-box domain and six carboxy terminal kelch repeats. Amino acid sequence of InZTL showed 84 % homology with Mesembryanthemum crystallinum ZTL (McZTL) and 83 % with Arabidopsis thaliana ZTL (AtZTL). Fluorescence in situ hybridization (FISH) to InZTL mRNA showed its high accumulation in the vascular bundles as well in the guard cells of the cotyledon. Immunolocalization of ZTL protein indicated a similar distribution pattern of ZTL protein as InZTL mRNAs.