Morphogenetic and diurnal variation of hypericin in some Hypericum species from Turkey during the course of ontogenesis

The genus Hypericum has received considerable interest from scientists, as it is a source of a variety of biologically active compounds including the hypericins. The present study was conducted to determine ontogenetic, morphogenetic and diurnal variation of the total hypericins content in some species of Hypericum growing in Turkey namely, Hypericum aviculariifolium subsp. depilatum var. depilatum (endemic), Hypericum perforatum and Hypericum pruinatum. The Hypericum plants were harvested from wild populations at vegetative, floral budding, full flowering, fresh fruiting and mature fruiting stages four times a day. Plants were dissected into stem, leaf and reproductive tissues, which were dried separately, and subsequently assayed for total hypericin content. The density of dark glands on leaves at full flowering plants was determined for each species. Floral parts had the highest hypericin content in all species tested. But diurnal fluctuation in the hypericin content of whole plant during the course of ontogenesis varied among the species. It reached the highest level at floral budding and tended to increase at night in H. aviculariifolium subsp. depilatum var. depilatum and H. pruinatum, whereas in H. perforatum hypericin content was the highest at full flowering and no diurnal fluctuation was observed. In general, hypericin content of leaves and whole plant was higher in H. aviculariifolium subsp. depilatum var. depilatum whose leaves had more numerous dark glands than those of the two other species.     

Stress enhances the gene expression and enzyme activity of phenylalanine ammonia-lyase and the endogenous content of salicylic acid to induce flowering in pharbitis

The involvement of salicylic acid (SA) in the regulation of stress-induced flowering in the short-day plant pharbitis (also called Japanese morning glory) Ipomoea nil (formerly Pharbitis nil) was studied. Pharbitis cv. Violet was induced to flower when grown in 1/100-strength mineral nutrient solution under non-inductive long-day conditions. All fully expanded true leaves were removed from seedlings, leaving only the cotyledons, and flowering was induced under poor-nutrition stress conditions. This indicates that cotyledons can play a role in the regulation of poor-nutrition stress-induced flowering. The expression of the pharbitis homolog of PHENYLALANINE AMMONIA-LYASE, the enzyme activity of phenylalanine ammonia-lyase (PAL; E.C. 4.3.1.5) and the content of SA in the cotyledons were all up-regulated by the stress treatment. The Violet was also induced to flower by low-temperature stress, DNA demethylation and short-day treatment. Low-temperature stress enhanced PAL activity, whereas non-stress factors such as DNA demethylation and short-day treatment decreased the activity. The PAL enzyme activity was also examined in another cultivar, Tendan, obtaining similar results to Violet. The exogenously applied SA did not induce flowering under non-stress conditions but did promote flowering under weak stress conditions in both cultivars. These results suggest that stress-induced flowering in pharbitis is induced, at least partly, by SA, and the synthesis of SA is promoted by PAL.     

Volatiles associated with different flower stages and leaves of Acacia cyclops and their potential role as host attractants for Dasineura dielsi (Diptera: Cecidomyiidae)

Acacia cyclops (Fabaceae) is an Australian species which was introduced into South Africa in the nineteenth century. Because of its invasive status in South Africa, a gall midge, Dasineura dielsi (Diptera: Cecidomyiidae), was released in 2001 in order to impact its reproduction by inducing galls on the flowers and thereby preventing seed set. Nothing is known about the cues used by D. dielsi for locating its host flowers. As part of an initial investigation into whether or not chemical cues might play a role in host finding, we analysed headspace samples of Acacia cyclops volatiles from leaves and reproductive parts at different stages (early bud, late bud, early flowering, and senescing flowering stages) using gas chromatography–mass spectrometry (GC–MS). In total, 72 different compounds were detected of which 62 were identified. The analyses showed that open flowers, the stage used by D. dielsi for oviposition, and yellow buds had similar odour compositions with (Z)-3-hexen-1-ol acetate, 4-oxoisophorone, (Z)-β-ocimene, an unknown aliphatic compound, heptadecane, and nonadecane dominating in open flowers. Leaf volatiles were distinct from those in the reproductive plant parts by their high relative amount of (Z)-β-ocimene. (Z)-3-Hexen-1-ol acetate had its maximum relative amount in the green bud samples and was much lower in the later floral stages. In contrast, 4-oxoisophorone peaked in yellow buds and open flowers with little or none of it found in younger or older stages. The volatile compounds of the different flower stages and leaves are discussed in relation to their potential role as attractants used by the biocontrol agent D. dielsi to locate its host plant.     

Fruit load modulates flowering-related gene expression in buds of alternate-bearing ‘Moncada’ mandarin

Background and Aims

Gene determination of flowering is the result of complex interactions involving both promoters and inhibitors. In this study, the expression of flowering-related genes at the meristem level in alternate-bearing citrus trees is analysed, together with the interplay between buds and leaves in the determination of flowering.

Methods

First defruiting experiments were performed to manipulate blossoming intensity in ‘Moncada’ mandarin, Citrus clementina. Further defoliation was performed to elucidate the role leaves play in the flowering process. In both cases, the activity of flowering-related genes was investigated at the flower induction (November) and differentiation (February) stages.

Key Results

Study of the expression pattern of flowering-genes in buds from on (fully loaded) and off (without fruits) trees revealed that homologues of FLOWERING LOCUS T (CiFT), TWIN SISTER OF FT (TSF), APETALA1 (CsAP1) and LEAFY (CsLFY) were negatively affected by fruit load. CiFT and TSF activities showed a marked increase in buds from off trees through the study period (ten-fold in November). By contrast, expression of the homologues of the flowering inhibitors of TERMINAL FLOWER 1 (CsTFL), TERMINAL FLOWER 2 (TFL2) and FLOWERING LOCUS C (FLC) was generally lower in off trees. Regarding floral identity genes, the increase in CsAP1 expression in off trees was much greater in buds than in leaves, and significant variations in CsLFY expression (approx. 20 %) were found only in February. Defoliation experiments further revealed that the absence of leaves completely abolished blossoming and severely affected the expression of most of the flowering-related genes, particularly decreasing the activity of floral promoters and of CsAP1 at the induction stage.

Conclusions

These results suggest that the presence of fruit affects flowering by greatly altering gene-expression not only at the leaf but also at the meristem level. Although leaves are required for flowering to occur, their absence strongly affects the activity of floral promoters and identity genes.     

SHB1 plays dual roles in photoperiodic and autonomous flowering

Flowering was initiated by the integration of environmental signals such as day-length with the internal development status in Arabidopsis, a facultative long-day plant. The photoperiodic flowering involves two key components, CONSTANS and FT, whereas the autonomous flowering is operated through a central quantitative floral repressor, FLC, and several other genes that act upstream of FLC. SOC1 acts downstream to integrate the flowering signals from the two pathways. Here, we report that SHB1 plays dual roles in both photoperiodic and autonomous flowering. shb1-D, a gain-of-function mutant, flowered early and shb1, a loss-of-function allele, flowered late under both long days and short days. The shb1-D mutation activated the expression of CO, FT, and SOC1 under both long and short days, and however, the co-2 mutation attenuated the shb1-D activated expression of FT and SOC1 only under long days but not short days. The shb1-D or shb1 mutations also reduced and increased, respectively, the expression of FLC under both long and short days. Transgenic remedy of FLC to wide-type level in shb1-D background also reverted shb1-D flowering and FT or SOC1 expression to wild type mostly under short days. Furthermore, the shb1-D suppression on FLC expression is likely operated through LD as ld-3 blocked this suppression and SHB1 appears to act upstream of LD. In summary, SHB1 represents signaling steps that regulate CO expression in leaves and LD or FLC expression in either leaves or shoot apical meristem, contributing to a threshold expression of SOC1 in shoot apical meristem for floral initiation.     

Fruit regulates seasonal expression of flowering genes in alternate-bearing 'Moncada' mandarin

Background and Aims

The presence of fruit has been widely reported to act as an inhibitor of flowering in fruit trees. This study is an investigation into the effect of fruit load on flowering of ‘Moncada’ mandarin and on the expression of putative orthologues of genes involved in flowering pathways to provide insight into the molecular mechanisms underlying alternate bearing in citrus.

Methods

The relationship between fruit load and flowering intensity was examined first. Defruiting experiments were further conducted to demonstrate the causal effect of fruit removal upon flowering. Finally, the activity of flowering-related genes was investigated to determine the extent to which their seasonal expression is affected by fruit yield.

Key Results

First observations and defruiting experiments indicated a significant inverse relationship between preceding fruit load and flowering intensity. Moreover, data indicated that when fruit remained on the tree from November onwards, a dramatic inhibition of flowering occurred the following spring. The study of the expression pattern of flowering-genes of on (fully loaded) and off (without fruits) trees revealed that homologues of FLOWERING LOCUS T (FT), SUPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1), APETALA1 (AP1) and LEAFY (LFY) were negatively affected by fruit load. Thus, CiFT expression showed a progressive increase in leaves from off trees through the study period, the highest differences found from December onwards (10-fold). Whereas differences in the relative expression of SOC1 only reached significance from September to mid-December, CsAP1 expression was constantly higher in those trees through the whole study period. Significant variations in CsLFY expression only were found in late February (close to 20 %). On the other hand, the expression of the homologues of TERMINAL FLOWER 1 (TFL1) and FLOWERING LOCUS C (FLC) did not appear to be related to fruit load.

Conclusions

These results suggest for the first time that fruit inhibits flowering by repressing CiFT and SOC1 expression in leaves of alternate-bearing citrus. Fruit also reduces CsAP1 expression in leaves, and the significant increase in leaf CsLFY expression from off trees in late February was associated with the onset of floral differentiation.     

A stress responsive alternative splicing mechanism in Citrus clementina leaves

Chitinases are often considered pathogenesis-related proteins since their activity can be induced by viral infections, fungal and bacterial cell wall components, and also by more general sources of stress such as wounding, salicylic acid, ethylene, auxins and cytokinins. In the present study, comparative proteomic analysis showed the defense-related acidic chitinase II to be specifically induced in Citrus clementina leaves infested by the two-spotted spider mite Tetranychus urticae or treated with MeJA. In parallel, changes in the mRNA profiles of two partially homologous chitinase forms were shown by RT-PCR. In particular, the appearance of an additional cDNA chitinase fragment in T. urticae-infested and MeJA-treated leaves was observed. This finding may indicate a specific regulatory mechanism of chitinase expression. We report evidence for alternative splicing in T. urticae-infested C. clementina, where a premature stop codon after the first 135 amino acids was introduced. We observed inducible chitinase activity after MeJA treatment, indicative of a rapid plant response to infestation. This work provides the first evidence of chitinase alternative splicing in C. clementina. In addition, the presence of the dual-band pattern for chitinase cDNA by RT-PCR may represent a suitable predictive marker for early diagnosis of plant biotic stress.     

Potential use in phytoremediation of three plant species growing on contaminated mine-tailing soils in Sardinia

We have analyzed the relationship between total Zn, Pb and Cu concentrations in the soil and the capacity of three plant species to accumulate these elements in their leaves. The study was carried out in a highly contaminated area at Sulcis-Iglesiente (SW-Sardinia, Italy). We took samples of the leaves of Dittrichia viscosa, Cistus salviifolius and Euphorbia pithyusa subsp. cupanii and samples of the soil beneath each of them at depths of 0-30 and 30-60 cm, both in contaminated mine tailings and surrounding areas. Due to the anthropic origin of the soil materials the results varied considerably. Bioavailability of trace elements was mainly related to the calcium-carbonate content and the crystalline and amorphous forms of iron in the soil. The concentration of Zn in the leaves of the three plant species studied was highest, followed by Pb and finally Cu. The leaves of Dittrichia viscosa contained the highest concentrations of trace elements and this species may be considered as being a “phytoextractor” in soils where the trace-element concentrations are not too high. Euphorbia pithyusa subsp. cupanii had low trace-element concentrations in its leaves despite growing in highly contaminated soils, and so might be used as a “phytostabilizer”. Although Cistus salviifolius does not grow in the most contaminated soils, could be considered as a contamination indicator up to a given level.     

Novel vitamin E forms in leaves of Kalanchoe daigremontiana and Phaseolus coccineus

In the present study, we isolated novel tocochromanols from green leaves of Kalanchoe daigremontiana and primary leaves of etiolated seedlings of Phaseolus coccineus that were identified as β-, γ-, and δ-tocomonoenols with unsaturation at the terminal isoprene unit of the side chain. The content of γ-tocomonoenol in leaves of etiolated bean increased gradually with the age of seedlings, reaching 50% of the γ-tocopherol level in 40-day-old plants. The content of this compound in leaves was increased by short illumination of etiolated plants and by addition of homogentisic acid, a biosynthetic precursor of tocopherols. These data indicated that γ-tocomonoenol is synthesized de novo from homogentisic acid and tetrahydro-geranylgeraniol diphosphate, a phytol precursor. Based on these results, a biosynthetic pathway of tocomonoenols is proposed.     

Flower inhibition in Kalanchoe blossfeldiana. Bioassay of an endogenous long-day inhibitor and inhibition by (±) abscisic acid and xanthoxin

Summary The inhibition of flowering in Kalanchoe by crude sap expressed from leaves held in non-inductive long-day conditions is described, using a bioassay technique of leaf injection, which confirms the existence of a transferable inhibitor.This technique has also revealed that ± abscisic acid and Xanthoxin are inhibitory to flowering at 50 and 100 ppm respectively. The previously known inhibitory effects of gibberellic acid on flowering have also been confirmed.     

Trap diversity and evolution in the family Droseraceae

We review trapping mechanisms in the carnivorous flowering plant family Droseraceae (order Caryophyllales). Its members are generally known to attract, capture, retain and digest prey animals (mainly arthropods) with active snap-traps (Aldrovanda, Dionaea) or with active sticky flypaper traps (Drosera) and to absorb the resulting nutrients. Recent investigations revealed how the snap-traps of Aldrovanda vesiculosa (waterwheel plant) and Dionaea muscipula (Venus’ flytrap) work mechanically and how these apparently similar devices differ as to their functional morphology and shutting mechanics. The Sundews (Drosera spp.) are generally known to possess leaves covered with glue-tentacles that both can bend toward and around stuck prey. Recently, it was shown that there exists in this genus a higher diversity of different tentacle types and trap configurations than previously known which presumably reflect adaptations to different prey spectra. Based on these recent findings, we finally comment on possible ways for intrafamiliar trap evolution.     

Using a starch-rich mutant of Arabidopsis thaliana as feedstock for fermentative hydrogen production

A mutant plant (Arabidopsis thaliana), sex1-1 (starch excess 1-1), accumulating high starch content in leaves was created to serve as better biomass feedstock for a H₂-producing strain Clostridium butyricum CGS2, which efficiently utilizes starch for H₂ production but cannot assimilate cellulosic materials. The starch content of the mutant plant increased to 10.67 mg/fresh weight, which is four times higher than that of wild type plant. Using sex1-1 mutant plant as feedstock, C. butyricum CGS2 could produce 490.4 ml/l of H₂ with a H₂ production rate of 32.9 ml/h/l. The H₂ production performance appeared to increase with the increase in the concentration of mutant plant from 2.5 to 10 g/l. The highest H₂ to plant biomass yield was nearly 49 ml/g for the mutant plant. This study successfully demonstrated the feasibility of using a starch-rich mutant plant for more effective bioH₂ production with C. butyricum CGS2.     

Al-hyperaccumulator Vochysiaceae from the Brazilian Cerrado store aluminum in their chloroplasts without apparent damage

We investigated the pattern of aluminum (Al) accumulation in leaf tissues of native hyperaccumulator Vochysiaceae species Qualea grandiflora,Callisthene major, and Vochysia pyramidalis, from the Brazilian Cerrado. Non-accumulator Sclerolobium paniculatum was used as a control species. We expected a strong compartmentalization of Al in non-active leaf cell compartments such as cell walls and vacuoles in Al-accumulating species and the absence of Al in critical metabolic sites such as the chloroplasts. Plant leaves were harvested in the field and cut in small segments for histological analysis; hematoxylin dye was used for Al localization in tissues. Results of soil analysis of the three sites and the concentration of Al in leaves indicated that there is no direct relationship between Al availability in soils and Al hyperaccumulation among the Vochysiaceae species evaluated. The cross-sections of leaf tissues showed hematoxylin color in the palisade and spongy parenchyma cells (chloroplast) of Q. grandiflora and C. major. The vascular system of Q. grandiflora was not colored, but some cells from the xylem region of C. major were stained. In contrast, the adaxial and abaxial epidermal cells of V. pyramidalis were colored by hematoxylin, as were some cells from the vascular bundle, but color formation was not observed in the cells of palisade parenchyma. Al was not detected in leaves of S. paniculatum. We concluded that, although hyperaccumulation of Al is a common trait in the Vochysiaceae family, the processes of storage and detoxification in leaf tissues differ among the species. Two of the three hyperaccumulator species use chloroplasts as a sink for Al, with no apparent signs of toxicity. Therefore, the physiological role of Al in plant tissues remains to be elucidated.     

The complex interactions between host immunity and non-biotrophic fungal pathogens of wheat leaves

Significant progress has been made in elucidating the mechanisms used by plants to recognize pathogens and activate “immune” responses. A “first line” of defense can be triggered through recognition of conserved Pathogen or Microbe Associated Molecular Patterns (PAMPs or MAMPs), resulting in activation of basal (or non-host) plant defenses, referred to as PAMP-triggered immunity (PTI). Disease resistance responses can also subsequently be triggered via gene-for-gene type interactions between pathogen avirulence effector genes and plant disease resistance genes (Avr-R), giving rise to effector triggered immunity (ETI). The majority of the conceptual advances in understanding these systems have been made using model systems, such as Arabidopsis, tobacco, or tomato in combination with biotrophic pathogens that colonize living plant tissues. In contrast, how these disease resistance mechanisms interact with non-biotrophic (hemibiotrophic or necrotrophic) fungal pathogens that thrive on dying host tissue during successful infection, is less clear. Several lines of recent evidence have begun to suggest that these organisms may actually exploit components of plant immunity in order to infect, successfully colonize and reproduce within host tissues. One underlying mechanism for this strategy has been proposed, which has been referred to as effector triggered susceptibility (ETS). This review aims to highlight the complexity of interactions between plant recognition and defense activation towards non-biotrophic pathogens, with particular emphasis on three important fungal diseases of wheat (Triticum aestivum) leaves.     

High levan accumulation in transgenic tobacco plants expressing the Gluconacetobacter diazotrophicus levansucrase gene

Bacterial levansucrase (EC 2.4.1.10) converts sucrose into non-linear levan consisting of long β(2,6)-linked fructosyl chains with β(2,1) branches. Bacterial levan has wide food and non-food applications, but its production in industrial reactors is costly and low yielding. Here, we report the constitutive expression of Gluconacetobacter diazotrophicus levansucrase (LsdA) fused to the vacuolar targeting pre-pro-peptide of onion sucrose:sucrose 1-fructosyltransferase (1-SST) in tobacco, a crop that does not naturally produce fructans. In the transgenic plants, levan with degree of polymerization above 10⁴ fructosyl units was detected in leaves, stem, root, and flowers, but not in seeds. High levan accumulation in leaves led to gradual phenotypic alterations that increased with plant age through the flowering stage. In the transgenic lines, the fructan content in mature leaves varied from 10 to 70% of total dry weight. No oligofructans were stored in the plant organs, although the in vitro reaction of transgenic LsdA with sucrose yielded β(2,1)-linked FOS and levan. Transgenic lines with levan representing up to 30 mg g⁻¹ of fresh leaf weight produced viable seeds and the polymer accumulation remained stable in the tested T1 and T2 progenies. The lsdA-expressing tobacco represents an alternative source of highly polymerized levan.     

模拟酸雨对西洋杜鹃生理生态特性的影响

采用盆栽方法,研究了不同pH值条件下的模拟酸雨对西洋杜鹃生长及其叶片主要生理生化特征的影响。结果表明:在酸雨胁迫下,随着酸雨胁迫的增强,西洋杜鹃叶片受害程度逐渐加重;叶片丙二醛含量逐渐升高;可溶性蛋白含量先升后降;过氧化氢酶(CAT)、过氧化物酶(POD)和超氧化物歧化酶(SOD)活性均呈先升后降的单峰曲线变化,其中pH值为4.3的处理下西洋杜鹃叶片CAT和SOD活性最高,pH值为3.0的处理下其POD活性最高;其叶绿素含量逐渐下降;叶片净光合速率(Pn)、蒸腾速率(Tr)、气孔导度(Gs)及水分利用率(WUE)均呈先升后降的趋势,而胞间CO2浓度(Ci)则持续下降,pH值为4.3的处理下其Pn、Tr、Gs和WUE均达到最高。研究表明,大致可以认为pH值≤3.0是酸雨对西洋杜鹃造成隐形伤害的阀值,而酸雨灾害严重地区的降水pH值为2.0—4.0左右,说明西洋杜鹃可以在酸雨灾害较重的地区生长,可作为酸雨灾害严重地区园林绿化及植被构建的物种之一。     

Reproductive phenology of a northeast Brazilian mangrove community: Environmental and biotic constraints

Brazil has the third largest area of mangrove in the world, which is widely threatened by anthropogenic pressures. We carried out the first long-term phenological study investigating whether environment and competition for pollinators shape the reproduction of a western mangrove community in Brazil, and provide new information for mangrove conservation. We monitored monthly the flowering and fruiting of Avicennia schaueriana, Conocarpus erectus, Laguncularia racemosa and Rhizophora mangle, the only species composing this mangrove community. We applied circular statistics to detect seasonal trends, null models to test for aggregated, staggered or random flowering patterns, performed correlations between phenophases and climate, and calculated intra-specific phenological synchrony. Each species presented a different flowering pattern, from brief annual to continuous and from regular to irregular, resulting in a bimodal pattern at community level. Fruiting was annual or continuous and seasonally unimodal at community level. Precipitation showed the strongest correlation with reproduction for all species, except L. racemosa. Flowering was randomly distributed among species sharing pollinators and each species presented high intra-specific synchrony. The studied mangrove showed a diversity of flowering patterns despite the low number of species. Annual to sub-annual sequential flowering were prevalent, sustaining the pollinators of species all the year long, while the wind-pollinated species flowered continuously. We provide strong evidence that daylength, rainfall and temperature are driving the flowering and fruiting rhythm of these mangrove species.