We demonstrated the ability of salicylic acid (SA) to induce a compound in avocado roots that strengthens their defense against Phytophthora cinnamomi. The SA content of avocado roots, before and after the application of exogenous SA, was determined by High-Performance Liquid Chromatography (HPLC). After 4h of SA feeding, the endogenous level in the roots increased to 223μgg(-1) FW, which was 15 times the amount found in control roots. The methanolic extract obtained from SA-treated avocado roots inhibited the radial growth of P. cinnamomi. A thin layer chromatographic bioassay with the methanolic extract and spores of Aspergillus showed a distinct inhibition zone. The compound responsible for the inhibition was identified as phenol-2,4-bis (1,1-dimethylethyl) by gas chromatography and mass spectrometry. At a concentration of 100μg/mL, the substance reduced germinative tube length in Aspergillus and radial growth of P. cinnamomi. A commercial preparation of phenol-2,4-bis (1,1-dimethylethyl) caused the same effects on mycelium morphology and radial growth as our isolate, confirming the presence of this compound in the root extracts. This is the first report of the induction of this compound in plants by SA, and the results suggest that it plays an important role in the defense response of avocado.
A critical stage in pollen development is the dissolution of tetrads into free microspores. Tetrads are surrounded by a wall composed primarily of beta-1,3-glucan. At the completion of meiosis, tetrads are released into the anther locule after hydrolysis of the callose by a beta-1,3-glucanase complex. The cDNA corresponding to a beta-1,3-glucanase cloned from tobacco (Tag 1) represents a gene that is highly similar to other beta-1,3-glucanases and is expressed exclusively in anthers from the tetrad to free microspore stage of pollen development. Tag 1 protein was overexpressed in E. coli, accumulating in insoluble inclusion bodies. Polyclonal antibodies against Tag 1 recombinant protein identify a single 33 kD protein accumulating only in anthers at tetrad and free microspore stages where beta-1,3-glucanase activity is present. Transgenic plants expressing Tag 1 antisense RNA were produced. Although Tag 1 RNA and protein levels were greatly reduced, tetrad dissolution and pollen development were normal. These data indicate that under the conditions these tobacco plants were grown, wild type levels of Tag 1 protein are not necessary for male fertility.
We isolated from strawberry (Fragariae x ananassa Duch) a genomic clone of a beta-1,3-glucanase gene, designated as FaBG2-2. In addition, a related cDNA clone, designated as FaBG2-3, was also isolated. FaBG2-2 and FaBG2-3 are similar in their coding regions, except that FaBG2-2 does not appear to contain a signal peptide coding sequence. The 5' and 3' flanking regions of FaBG2-2 and FaBG2-3 are differentt. Using real-time PCR, the expression patterns of FaBG2-3 and a previously isolated beta-1,3-glucanase gene, FaBG2-1, in strawberry plants infected with Colletotrichum fragariae or Colletotrichum acutatum were analyzed at different time points post-infection. The results showed that expressions of both genes in the leaves of infected plants were induced by the two fungi, but the level of induction was several fold greater with C. fragariae. Comparison of the expression levels of the two genes revealed that the level of FaBG2-3 expression was several hundred to over a thousand fold higher than that of FaBG2-1. Furthermore, the expression levels of the two genes in the leaf, fruit, crown and root of uninfected strawberry plants were analyzed.
We isolated two rice endo-(1,3;1,4)-beta-glucanase genes, denoted OsEGL1 and OsEGL2, which encoded proteins that shared 64% amino acid sequence identity. Both the OsEGL1 and OsEGL2 genes were successfully expressed in Escherichia coli to produce functional proteins. Purified OsEGL1 and OsEGL2 proteins hydrolyzed (1,3;1,4)-beta-glucans, but not (1,3;1,6)-beta-linked or (1,3)-beta-linked glucopolysaccharides nor carboxymethyl cellulose, similar to previously characterized grass endo-(1,3;1,4)-beta-glucanases. RNA blot analysis revealed that the OsEGL1 gene is expressed constitutively not only in young roots of rice seedlings, but also in mature roots of adult rice plants. Little or no expression of the OsEGL2 gene was observed in all tissues or treatments tested, but database and RT-PCR analysis indicated it is expressed in ripening panicle. In rice seedling leaves, OsEGL1 gene expression significantly increased in response to methyl jasmonate, abscisic acid, ethephon and mechanical wounding. Mechanical wounding also increased the leaf elongation rate in rice seedlings by 16% relative to that of control seedlings at day 4 after treatment. The increase in the leaf elongation rate of rice seedlings treated under mechanical wounding was concomitant with an increase in OsEGL1 expression levels in seedling leaves.
Nitrogen fixation in legumes is dramatically inhibited by abiotic stresses, and this reduction is often associated with oxidative damage. Although ascorbate (ASC) has been firmly associated with antioxidant defence, recent studies have suggested that the functions of ASC are related primarily to developmental processes. This study examines the hypothesis that ASC is involved in alleviating the oxidative damage to nodules caused by an increase in reactive oxygen species (ROS) under water stress. The hypothesis was tested by supplying 5mM ASC to pea plants (Pisum sativum L.) experiencing moderate water stress (ca. -1 MPa) and monitoring plant responses in relation to those experiencing the same water stress without ASC. A supply of exogenous ASC increased the nodule ASC+dehydroascorbate (DHA) pool compared to water-stressed nodules without ASC, and significantly modulated the response to water stress of the unspecific guaiacol peroxidase (EC 126.96.36.199) in leaves and nodules. However, ASC supply did not produce recovery from water stress in other nodule antioxidant enzymes, nodule carbon and nitrogen enzymes, or nitrogen fixation. The supply of the immediate ASC precursor, galactono-1,4-lactone (GL), increased the nodule ASC+DHA pool, but also failed to prevent the decline of nitrogen fixation and the reduction of carbon flux in nodules. These results suggest that ASC has a limited role in preventing the negative effects of water stress on nodule metabolism and nitrogen fixation.
The tiller of rice (Oryza sativa L.), which determines the panicle number per plant, is an important agronomic trait for grain production. Ascorbic acid (Asc) is a major plant antioxidant that serves many functions in plants. l-Galactono-1,4-lactone dehydrogenase (GLDH, EC 188.8.131.52) is an enzyme that catalyzes the last step of Asc biosynthesis in plants. Here we show that the GLDH-suppressed transgenic rices, GI-1 and GI-2, which have constitutively low (between 30% and 50%) leaf Asc content compared with the wild-type plants, exhibit a significantly reduced tiller number. Moreover, lower growth rate and plant height were observed in the Asc-deficient plants relative to the trait values of the wild-type plants at different tillering stages. Further examination showed that the deficiency of Asc resulted in a higher lipid peroxidation, a loss of chlorophyll, a loss of carotenoids, and a lower rate of CO(2) assimilation. In addition, the level of abscisic acid was higher in GI-1 plants, while the level of jasmonic acid was higher in GI-1 and GI-2 plants at different tillering stages. The results we presented here indicated that Asc deficiency was likely responsible for the promotion of premature senescence, which was accompanied by a marked decrease in photosynthesis. These observations support the conclusion that the deficiency of Asc alters the tiller number in the GLDH-suppressed transgenics through promoting premature senescence and changing phytohormones related to senescence.
This study investigated the effects of cinnamic acid (CA) on ribulose-1,5-bisphosphate carboxylase (RuBPC) activity and the endogenous polyamine levels of cowpea leaves. The results show that 0.1 mM CA treatment decreased photosynthetic rate (P(n)) and RuBPC activity, but it did not affect the maximal photochemical efficiency of PSII (F(v)/F(m)), the actual photochemical efficiency of PSII (PhiPSII), intercellular CO(2) concentration (C(i)), and relative chlorophyll content. These suggest that the decrease in P(n) is at least partially attributed to a lowered RuBPC activity. In addition, 0.1 mM CA treatment increased the putrescine (Put) level, but decreased spermidine (Spd) and spermine (Spm) levels, thereby reducing the (Spd+Spm)/Put (PAs) ratio in the leaves. The exogenous application of 1 mM Spd markedly reversed these CA-induced effects for polyamine and partially restored the PAs ratio and RuBPC activity in leaves. Methylglyoxal-bis (guanylhydrazone) (MGBG), which is an inhibitor of S-adenosylmethionine decarboxylase (SAMDC), results in the inability of activated cells to synthesize Spd and exacerbates the negative effects induced by CA. The exogenous application of 1 mM D-arginine (D-Arg), which is an inhibitor of Put biosynthesis, decreased the levels of Put, but increased the PAs ratio and RuBPC activity in leaves. These results suggest that 0.1 mM CA inhibits RuBPC activity by decreasing the levels of endogenous free and perchloric acid soluble (PS) conjugated Spm, as well as the PAs ratio.
Plastidial degradation of transitory starch yields mainly maltose and glucose. Following the export into the cytosol, maltose acts as donor for a glucosyl transfer to cytosolic heteroglycans as mediated by a cytosolic transglucosidase (DPE2; EC 184.108.40.206) and the second glucosyl residue is liberated as glucose. The cytosolic phosphorylase (Pho2/PHS2; EC 220.127.116.11) also interacts with heteroglycans using the same intramolecular sites as DPE2. Thus, the two glucosyl transferases interconnect the cytosolic pools of glucose and glucose 1-phosphate. Due to the complex monosaccharide pattern, other heteroglycan-interacting proteins (HIPs) are expected to exist.
Lateral roots (LRs) play important roles in increasing the absorptive capacity of roots as well as to anchor the plant in the soil. Therefore, understanding the regulation of LR development is of agronomic importance. In this study, we examined the effect of methyl jasmonate (MJ) on LR formation in rice. Treatment with MJ induced LR formation and heme oxygenase (HO) activity. As well, MJ could induce OsHO1 mRNA expression. Zinc protoporphyrin IX (the specific inhibitor of HO) and hemoglobin [the carbon monoxide/nitric oxide (NO) scavenger] reduced LR formation, HO activity and OsHO1 expression. LR formation and HO activity induced by MJ was reduced by the specific NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-oxide. The effects of Ca(2+) chelators, Ca(2+)-channel inhibitors, and calmodulin (CaM) antagonists on LR formation induced by MJ were also examined. All these inhibitors were effective in reducing the action of MJ. However, Ca(2+) chelators and Ca(2+) channel inhibitors induced HO activity when combining with MJ further. It is concluded that Ca(2+) may regulate MJ action mainly through CaM-dependent mechanism.
Environmental stresses trigger a wide variety of plant responses, ranging from altered gene expression and cellular metabolism to changes in growth rates and crop yields. A plethora of plant reactions exist to circumvent the potentially harmful effects caused by a wide range of both abiotic and biotic stresses, including light, drought, salinity, high temperatures, and pathogen infections. Among the environmental stresses, drought stress is one of the most adverse factors of plant growth and productivity. Understanding the biochemical and molecular responses to drought is essential for a holistic perception of plant resistance mechanisms to water-limited conditions. Drought stress progressively decreases CO2 assimilation rates due to reduced stomatal conductance. Drought stress also induces reduction in the contents and activities of photosynthetic carbon reduction cycle enzymes, including the key enzyme, ribulose-1,5-bisphosphate carboxylase/oxygenase. The critical roles of proline and glycine-betaine, as well as the role of abscisic acid (ABA), under drought stress conditions have been actively researched to understand the tolerance of plants to dehydration. In addition, drought stress-induced generation of active oxygen species is well recognized at the cellular level and is tightly controlled at both the production and consumption levels in vivo, through increased antioxidative systems. Knowledge of sensing and signaling pathways, including ABA-mediated changes in response to drought stress, is essential to improve crop management. This review focuses on the ability and strategies of higher plants to respond and adapt to drought stress.
The time course of glycinin hydrolysis by papain was followed using densitometry of SDS-PAGE patterns, quantification of the residual protein and determination of its molecular mass by gel filtration, and by other appropriate methods. The hydrolysis occurs in two steps. In the first step, a limited proteolysis was observed consisting of a gradual detachment of the α-chain C-terminal sequence region, leading to the formation of glycinin-P, a relatively stable proteolysis product retaining the primordial hexameric structure. Glycinin-P was found to be composed of the intact β-chains covalently bound with the C-terminally truncated α-chains lacking the helix domain, strand J', and the C-terminal disordered region. Glycinin-P is further hydrolyzed in the second step exclusively by a one-by-one mechanism. Comparison of the kinetics of the limited and one-by-one proteolyses analyzed separately indicated that the decrease of protein concentration by 24-25% in the first step occurs almost exclusively due to the decrease of the molecular mass of the residual protein. Thus, the onset of the one-by-one proteolysis is delayed, suggesting a regulatory role of the preceding limited proteolysis in the subsequent massive degradation of glycinin. Probable structural alterations of glycinin generated by this limited proteolysis are discussed.
Orthosiphon stamineus (Java tea) has been widely used as traditional herb and several bioactive compounds against animal cells have been isolated. However, no bioactive compound against plants has been reported. Therefore, we investigated possible allelopathic properties and substances in O. stamineus. Aqueous methanol extracts of O. stamineus inhibited root and hypocotyl growth of cress (Lepidium sativum) and lettuce (Lactuca sativa) seedlings. Increasing the extract concentration increased the inhibition, which suggests that O. stamineus may have allelopathic properties. When the extract was divided into an ethyl acetate and an aqueous fraction, the ethyl acetate fraction showed the stronger inhibitory effect. Thus, the ethyl acetate phase was further purified, and the main allelopathic substance was isolated and identified as 13-epi-orthosiphol N, a novel compound, by spectral data. 13-epi-Orthosiphol N inhibited root and hypocotyl growth of cress and lettuce at concentrations greater than 10μmol/L. The concentrations required for 50% inhibition ranged from 41 to 102μmol/L. These results suggest that 13-epi-orthosiphol N may be an allelochemical and main contributor to the growth inhibitory effect of O. stamineus and may have potential as a template for the development of new plant control substances.
Chlorophyll-oxidase in barley thylakoids was activated by linolenic acid. The reaction products from endogenous chlorophylls were investigated by thin layer and high performance liquid chromatography. Within 2-10 min a new pigment was formed which migrated identically with authentic 13(2)-hydroxychlorophyll a (1a). After reaction with methanolic acid, the reaction product was identified as 13(2)-hydroxymethylpheophorbide (1b) by high performance thin layer and liquid chromatography.
Fusarium verticillioides is the causal agent of ear rot in most maize-growing areas of southern Europe. F. verticillioides produces fumonisins commonly found at biologically significant concentrations in maize grain; the molecular interaction between the fungus and the plant is not well known, and little information is currently available about the defense response of maize against F. verticillioides infection. We attempted to identify genes that may be involved in Fusarium ear rot resistance using resistant and susceptible maize genotypes. Kernels of the resistant inbred showed significantly reduced incidence of infection by F. verticillioides, limited amounts of total fumonisin content and reduced fungal growth, as indicated by a lower copy number of β-tubulin 2 and FUM 21 genes of F. verticillioides. Gene expression data were obtained from microarray hybridizations using maize seeds infected with F. verticillioides, by comparing seeds at 0 and 48h after infection. Differentially expressed sequences were identified and classified into 11 functional categories. Most of the differentially expressed genes were assigned to the category "cell rescue, defense and virulence" in both resistant and susceptible maize lines. These genes encode for PR proteins, detoxification enzymes and β-glucosidases. Most of the pathogenesis-related genes were differentially activated after F. veticillioides infection, depending on the resistance level of the maize genotypes. In kernels of the resistant line, the defense-related genes assayed were transcribed at high levels before infection and provided basic defense against the fungus. In the susceptible kernels, the defense-related genes were induced from a basal level, responding specifically to pathogen infection. The qRT-PCR in infected silks showed that PR1, PR5, PRm6 and thaumatin genes had lower expression ratios in the resistant line compared to the susceptible one.
Accumulation of fructans was confirmed in asparagus tissues that had been cultured for 2 days on media supplemented with glucose. It is very common that fructans are biosynthesized from sucrose. We hypothesized however that fructans could also be biosynthesized from glucose. Stem tissues of in vitro-cultured asparagus were subcultured for 72h on a medium containing 0.5M of [1-(13)C]glucose. A medium containing 0.5M of normal ((12)C) glucose was used as control. Carbohydrates were extracted from the tissues and analyzed using HPLC, MALDI-TOF MS and ESI-MS. HPLC results indicated that the accumulation of short-chain fructans was similar in both (13)C-labelled and control samples. Short-chain fructans of DP=3-7 were detected using MALDI-TOF MS. The molecular mass of each oligomer in the (13)C-labelled sample was higher than the mass of the natural sample by 1 m/z unit per sugar moiety. The results of ESI-MS on the HPLC fractions of neokestose and 1-kestose showed that these oligomers (DP=3) were biosynthesized from exogenous glucose added to the medium. We conclude that not only exogenous sucrose but glucose can induce fructan biosynthesis; fructans of both inulin type and inulin neoseries are also biosynthesized from glucose accumulated in asparagus tissues; the glucose molecules (or its metabolic products) were incorporated into fructans as structural monomers.
The objective was to investigate the expression of a lipid transfer protein gene (LTP) both in bromegrass (Bromus inermis) cells and seedlings after exposure to abiotic stresses, abscisic acid (ABA), anisomycin, and sphingosine. A full-length cDNA clone BG-14 isolated from bromegrass suspension cell culture encodes a polypeptide of 124 amino acids with typical LTP characteristics, such as a conserved arrangement of cysteine residues. During active stages of cold acclimation LTP expression was up-regulated, whereas at the final stage of cold acclimation LTP transcript level declined to pre-acclimation level. A severe drought stress induced the LTP gene; yet, LTP expression doubled 3 d after re-hydration. Both temperature and heat shock duration influence LTP induction; however temperature is the primary factor. Treatment with NaCl stimulated accumulation of LTP mRNA within 15 min and the transcripts remained at elevated levels for the duration of the salinity stress. Most interestingly, Northern blots showed LTP was rapidly induced not only by ABA, but also by anisomycin and sphingosine in suspension cell cultures. Of the three chemicals, ABA induced the most rapid and highest response in LTP expression as well as highest freezing tolerance, whereas sphingosine was the least active for both LTP expression and freezing tolerance.
Sucrose-6-phosphate synthase (SPS) is a target for 14-3-3 protein binding in plants. Because several isoforms of the 14-3-3 protein are expressed in plants, I investigated which isoforms have the ability to bind SPS. Two 14-3-3 isoforms (T14-3d and a novel isoform designated T14-3 g) were found to interact with SPS from tobacco (Nicotiana tabacum L.) in a two-hybrid screen. To further address the question of isoform specificity of 14-3-3s, four additional isoforms were tested for their ability to interact with SPS in the yeast two-hybrid system. The results clearly revealed large differences in affinity between individual 14-3-3 isoforms toward SPS. Deletion analysis suggested that these differences were mediated by the variable C-terminus of 14-3-3s. Site-directed mutagenesis of candidate 14-3-3 binding sites on SPS demonstrated that interaction could be independent of a phosphorylated serine residue within conserved binding motifs in the yeast system. These findings suggest that the large number of 14-3-3 isoforms present in plants reflects functional specificity.
The C to U editing event that converts an ACG codon to an AUG translation initiation codon in the chloroplast rps 14 transcript is unique to the moss Physcomitrella patens and has not been found in other species. The efficiency of RNA editing was 80% in the young protonemata and decreased to approximately 20% in old protonemata and fully developed leafy shoots. This indicates that RNA editing of this site is regulated in a tissue- and stage-specific manner. In this study, a novel C to U RNA editing site has been identified at the -1 position relative to the AUG. Because the editing site is localized in the mRNA 5' untranslated region, it may affect the efficiency of rps 14 mRNA translation.
In intact vegetative plants of N. tabacum grown with progressively rising supplies of potassium, the increase in concentration of the cation is greater in the basal region, and growth is promoted in leaves approaching their final weight and subtending basal nodes while it is restricted in the apical leaves and internodes. The restrained growth changes to promotion after the leaves have attained 25 % and 51 % of their maximum weight in plants supplied with 3 and 9 meq K+ dm-3 respectively. The rate and duration of expansion as well as the final weight of the leaves were affected by K-supply, but not their emergence.
(32)P and (14)C-glucose uptake were reduced under B deficiency in both Daucus cell suspensions and Helianthus roots. Similarly, efflux rates were found to be smaller in B deficient material. Efflux rates tended to be more affected than net uptake of both (32)P and glucose. This may explain why sunflower roots showed a higher glucose net uptake immediately after transferring from a B sufficient to a B deficient nutrient solution. The data confirm earlier findings of B as an essential element for membrane function and integrity. B-deficiency effects could be reversed by the addition of B within less than one hour. Cell suspensions reacted similarly to roots with respect to B deficiency and may thus be suitable for further research on B deficiency effects.
Nitrate uptake and assimilation were examined in intact 18 days old wheat (Triticum aestivum, cv Capitole) seedlings either permanently grown on nitrate (high-N seedlings) or N-stressed by transfer to an 0 N-solution for the final 7 days (low-N seedlings). The N-stressed seedlings were characterized by a lower organic N content (2.5 mg instead of 4.9 mg per seedling) and an increased root dry weight. The seedlings received (15)NO(3)K for 7 h in the light. Nitrate uptake was 2.8 times higher in low-N than in high-N seedlings. The assimilation rate was 35 and 16 μmol NO(3)(-)·(h-1)· g(-1) dry weight respectively. Partitioning of NO(3)(-) to reduction and assimilation was the very same in both kinds of seedlings. The results support the view that 50 % of the nitrate reduction in Triticum aestivum, cv Capitole could be achieved in the roots. The present observations are interpreted as evidence that factors closely associated with the seedling N-status may have a major role in regulating NO(3)(-) uptake and assimilation. In low-N seedlings, the high amount of carbohydrates in roots may add its stimulus to the specific inducing effect of nitrate whereas in high-N seedlings, excess of nitrate or amino-acids may set the pace by negative feedback control.
Two irrigation systems were used to compare nitrogen uptake efficiency in citrus trees and to evaluate the NO3- runoff in "Navelina" orange trees [Citrus sinensis (L.) Osbeck] on Carrizo citrange rootstock (Citrus sinensis x Poncirus trifoliata Raf.). These were fertilized with 125 g N as labelled K15NO3 and grown outdoors in containers filled with a sand-loamy soil. Two groups of 3 trees received this N dose either in five equally split applications by a flooding irrigation system or in 66 applications by drip. Trees were harvested at the end of the vegetative cycle (December) and the isotopic ratios of 15N/14N were measured in the soil-plant system. The N uptake efficiency of the whole tree was higher with drip irrigation (75%) than with flooding system (64%). In the 0-90 cm soil profile, the N immobilized in the organic fraction was similar for both irrigation methods (around 13 %), whereas the N retained as NO3- was 1% of the N applied under drip and 10% under flooding. In the last case, most of NO3- remained under root system and it could be lost to leaching either by heavy rainfalls or excessive water applications. These results showed that a drip irrigation system was more efficient for improving water use and N uptake from fertilizer, in addition to potentially reduced leaching losses.
Although isotopic discrimination processes during nitrogen (N) transformations influence the outcome of (15)N based quantification of N2 fixation in legumes, little attention has been given to the effects of genotypic variability and environmental constraints such as phosphorus (P) deficiency, on discrimination against (15)N during N2 fixation. In this study, six Phaseolus vulgaris recombinant inbred lines (RILs), i.e. RILs 115, 104, 34 (P deficiency tolerant) and 147, 83, 70 (P deficiency sensitive), were inoculated with Rhizobium tropici CIAT899, and hydroaeroponically grown with P-sufficient (250μmol P plant(-1) week(-1)) versus P-deficient (75μmol P plant(-1) week(-1)) supply. Two harvests were done at 15 (before nodule functioning) and 42 (flowering stage) days after transplanting. Nodulation, plant biomass, P and N contents, and the ratios of (15)N over total N content ((15)N/Nt) for shoots, roots and nodules were determined. The results showed lower (15)N/Nt in shoots than in roots, both being much lower than in nodules. P deficiency caused a larger decrease in (15)N/Nt in shoots (-0.18%) than in nodules (-0.11%) for all of the genotypes, and the decrease in shoots was greatest for RILs 34 (-0.33%) and 104 (-0.25%). Nodule (15)N/Nt was significantly related to both the quantity of N2 fixed (R(2)=0.96***) and the P content of nodules (R(2)=0.66*). We conclude that the discrimination against (15)N in the legume N2-fixing symbiosis of common bean with R. tropici CIAT899 is affected by P nutrition and plant genotype, and that the (15)N/Nt in nodules may be used to screen for genotypic variation in P use efficiency for N2 fixation.
Receptor-like protein kinases (RLKs) play vital roles in sensing outside signals, yet little is known about RLKs functions and roles in stress signal perception and transduction in plants, especially in wild soybean. Through the microarray analysis, GsSRK was identified as an alkaline (NaHCO(3))-responsive gene, and was subsequently isolated from Glycine soja by homologous cloning. GsSRK encodes a 93.22kDa protein with a highly conserved serine/threonine protein kinase catalytic domain, a G-type lectin region, and an S-locus region. Real-time PCR results showed that the expression levels of GsSRK were largely induced by ABA, salt, and drought stresses. Over expression of GsSRK in Arabidopsis promoted seed germination, as well as primary root and rosette leaf growth during the early stages of salt stress. Compared to the wild type Arabidopsis, GsSRK overexpressors exhibited enhanced salt tolerance and higher yields under salt stress, with higher chlorophyll content, lower ion leakage, higher plant height, and more siliques at the adult developmental stage. Our studies suggest that GsSRK plays a crucial role in plant response to salt stress.
Selenium (Se) is an essential element for many organisms but also toxic at higher levels. The objective of this study was to identify accessions from the model species Arabidopsis thaliana that differ in Se tolerance and accumulation. Nineteen Arabidopsis accessions were grown from seed on agar medium with or without selenate (50 microM) or selenite (20 microM), followed by analysis of Se tolerance and accumulation. Tissue sulfur levels were also compared. The Se Tolerance Index (root length+Se/root length control) varied among the accessions from 0.11 to 0.44 for selenite and from 0.05 to 0.24 for selenate. When treated with selenite, the accessions differed by two-fold in shoot Se concentration (up to 250 mgkg(-1)) and three-fold in root Se concentration (up to 1000 mgkg(-1)). Selenium accumulation from selenate varied 1.7-fold in shoot (up to 1000 mgkg(-1)) and two-fold in root (up to 650 mgkg(-1)). Across all accessions, a strong correlation was observed between Se and S concentration in both shoot and root under selenate treatment, and in roots of selenite-treated plants. Shoot Se accumulation from selenate and selenite were also correlated. There was no correlation between Se tolerance and accumulation, either for selenate or selenite. The F(1) offspring from a cross between the extreme selenate-sensitive Dijon G and the extreme selenate-tolerant Estland accessions showed intermediate selenate tolerance. In contrast, the F(1) offspring from a cross between selenite-sensitive and -tolerant accessions (Dijon GxCol-PRL) were selenite tolerant. The results from this study give new insight into the mechanisms of plant selenium (Se) tolerance and accumulation, which may help develop better plants for selenium phytoremediation or as fortified foods.
After several years of close contacts and extensive discussion between various plant physiologists of different European countries, the Federation of European Societies of Plant Physiology (FESPP) was established in 1978 in Edinburgh. The aim of the FESPP was and remains to promote up-to-date plant physiology research in all European countries and to stimulate scientific cooperation and the exchange of scientists between the different member societies by organizing congresses and workshops as well as editing four (recently five) Federation-affiliated journals. The short History of FESPP presented here covers the preparatory years of the 1970s that led to its actual foundation in 1978, and then its further development up to and following the Federation's reconstitution in 2002 as the Federation of European Societies of Plant Biology (FESPB).
Grain protein content (GPC) is a key quality factor for malting and brewing process. In wheat, a QTL explaining a large part of GPC variation was identified, which co-localizes with a gene encoding a NAC transcription factor (TtNAM-B1). NAC transcription factors influence GPC by their role in the regulation of senescence and in protein remobilization. An orthologous gene was discovered on barley chromosome 6H where a GPC QTL was mapped. In this study, we identify allelic variation of the NAM-1 gene for three species of Hordeum representing wild and cultivated barley and we investigate the possible link with GPC. Three haplotypes were identified, one corresponds to the sequences of 11 European varieties representing H. vulgare, one corresponds to the sequence found in H. spontaneum and one represents the sequence of H. bulbosum. Three SNPs were identified between H. spontaneum sequence and H. vulgare sequence. One of the H. bulbosum polymorphisms leads to the introduction of a stop codon and a non-functional protein. Differences in GPC between the 11 varieties were found but no polymorphism in the NAM-1 gene was observed, suggesting that differences in expression of the HvNAM-1 gene or other genes should play a role in GPC regulation. Nevertheless based on published values for GPC of H. bulbosum and H. spontaneum compared to GPC measured here in H. vulgare, the non-functional protein is associated with the lower GPC, suggesting that loss of functionality of the NAM-1 gene in Hordeum is related to lower GPC. Moreover H. spontaneum GPC seems to be higher than H. vulgare GPC, suggesting also that allelic variation of the functional NAM-1 gene could be associated with GPC variation within the genus Hordeum.
The transmembrane transport of indole-3-acetic acidd (IAA) and 2,4-dichlorophenoxyacetic acid (2,4-D) by 2 mm segments of dark-grown Cucurbita pepo L. hypocotyls involves nonme-diated diffusion of the neutral protonated auxin species as well as carrier-mediated components for influx and efflux. The efflux carrier is blocked by 2,3,5-triiodobenzoic acid (TIBA), a noncompetitive inhibitor of polar auxin transport which is itself transported in a polar fashion. The transmembrane transport of TIBA includes diffusion of the protonated species (common to lipophilic weak acids) and also a carrier-mediated efflux component which can be blocked by IAA, 2,4-D, 1-naphthylacetic acid (NAA), and by 1-naphthylphthalamic acid (NPA) which, like TIBA, noncompetitively inhibits polar auxin transport but is not itself polarly transported. The non-auxin benzoic acid has no effect on auxin or TIBA transport. No significant metabolism of IAA, 2,4-D, or TIBA occurred during the experimental periods. A model is proposed for the auxin efflux carrier in which auxins and TIBA have separate specific binding sites; occupation of either one of the sites allows ligand translocation whereas occupation of both sites prevents any transport. A third specific site may be necessary to account for the inhibitory effects of NPA. The consequences of cytoplasmic acidification which can accompany lipophilic weak acid uptake are discussed.
Live cells can reduce colorless 2,3,5-triphenyltetrazolium chloride (TTC) to a red insoluble compound, formazan. Maize (Zea mays) callus, when osmotically stressed by 0.53 mol/L mannitol, produced 7-times or more formazan than untreated control callus. This result was seen with all osmotica tested and could not be attributed to differences in TTC uptake rate or accumulation, increased respiration rate as measured by O2 uptake, or to de novo protein synthesis. Increased formazan production could be detected after 2.5 h of exposure to osmotic stress and leveled off after 48 h of exposure. The increased formazan production was only detected when callus was moved from high osmotic medium to low osmotic, TTC-containing medium. Abscisic acid increased TTC reduction only when added in combination with 0.53 mol/L mannitol. Incubation of maize seedling roots with 0.53 mol/L mannitol also increased formazan production as seen visually. Further studies are needed to determine the cause of the increased formazan production. These results show that TTC viability measurements must be carefully evaluated with appropriate controls to confirm their validity.
The SMALL ACIDIC PROTEIN 2 (SMAP2) gene is a paralogue of the SMAP1 gene that mediates the response to the synthetic auxin 2,4-dichlorophenoxyacetic acid (2,4-D) in the root of Arabidopsis thaliana. Their encoded proteins, SMAP1 and SMAP2, are similar in calculated molecular weight and isoelectric point, and in having a highly conserved phenylalanine and aspartic acid-rich domain. RNA expression analysis showed that SMAP1 mRNA is present throughout the plant body while SMAP2 mRNA is restricted to siliques and anthers. Over-expression of the SMAP2 gene, as well as SMAP1, by 35S cauliflower mosaic virus promoter restored sensitivity to 2,4-D in the 2,4-D-resistant mutant, aar1, which is defective in SMAP1 function. The results suggest that SMAP2 has an ability to mediate the 2,4-D response and is expressed only in restricted tissues.
The effects of the herbicides 1,1'-dimethyl-4,4'-bipyridylium dichloride (paraquat), 3,6-dichloro-2-metoxybenzoic acid (dicamba) and 2,4-dichlorophenoxyacetic acid (2,4-D) on cell growth of non-green potato tuber calli are described. We attempted to relate the effects with toxicity, in particular the enzymes committed to the cellular antioxidant system. Cell cultures were exposed to the herbicides for a period of 4 weeks. Cellular integrity on the basis of fluorescein release was strongly affected by 2,4-D, followed by dicamba, and was not affected by paraquat. However, the three herbicides decreased the energy charge, with paraquat and 2,4-D being very efficient. Paraquat induced catalase (CAT) activity at low concentrations (1 microM), whereas at higher concentrations, inhibition was observed. Dicamba and 2,4-D stimulated CAT as a function of concentration. Superoxide dismutase (SOD) activity was strongly stimulated by paraquat, whereas dicamba and 2,4-D were efficient only at higher concentrations. Glutathione reductase (GR) activity was induced by all the herbicides, suggesting that glutathione and glutathione-dependent enzymes are putatively involved in the detoxification of these herbicides. Paraquat slightly inhibited glutathione S-transferase (GST), whereas 2,4-D and dicamba promoted significant activation. These results indicate that the detoxifying mechanisms for 2,4-D and dicamba may be different from the mechanisms of paraquat detoxification. However, the main cause of cell death induced by paraquat and 2,4-D is putatively related with the cell energy charge decrease.
To investigate the involvement of cytochrome P450s in the metabolism of plants treated with xenobiotic agrochemicals, bean leaves were treated with 3,5-dichlorosalicylic acid (DC-SA), a priming agent of plant defense and 2,6-dichloroisonicotinic acid (DC-INA), a chemical inducer of systemic acquired resistance. Through the use of directed differential display reverse transcription polymerase chain reactions, a differentially expressed cDNA amplicon, found to be up-regulated by both DC-SA and DC-INA treatment, was identified as a cytochrome P450 cDNA, CYP98A5. The nucleotide sequence indicates extensive homology to 3'-hydroxylases of p-coumaroyl esters. Dot blot analysis of leaves treated with various SA and isonicotinic acid derivatives showed enhanced expression of CYP98A5 due to DC-SA and DC-INA. Northern blot analysis of a time-dependent induction study of CYP98A5 in treated bean leaves indicated that DC-SA induces CYP98A5 mRNA transcripts earlier than DC-INA. Both inducers resulted in high transcript levels 24-48 h after treatment. The up-regulation of CYP98A5 is supportive of the conditioning and sensitizing effects of DC-SA and DC-INA to elicit a more rapid and effective defense response.
In the highly vacuolated epidermis cells of leaf explants of Nautilocalyx we investigated whether inhibition of cytokinesis by 2,6-dichlorobenzonitrile (DCB) could be caused by a suppression of the formation of phragmosome and band of microtubules (BMT), two structures which are probably involved in cytokinesis and in the determination of the plane of cell division. Interference contrast microscopy showed that DCB (116 μM) did not interfere with the formation of phragmosome and BMT in the expected plane of cell division. Also the positioning of the nucleus and the nuclear division proceeded normally. The phragmoplast was formed, and cell plate formation started, but in most cases the cell plate was not completed. After some time the part of the cell plate already formed shrank: folds appeared, and sometimes tears. The phragmoplast remained present for a long time after this premature end of cell plate growth. Electron microscopical studies showed a shortage of small Golgi vesicles with electron-dense contents in the plane of cell division where very large vesicles with little electron-dense material were present. Furthermore a dilatation of the endoplasmic reticulum in the microtubule zone of the phragmoplast was observed. These results indicate that the DCB-inhibition of cytokinesis does not result from interference with phragmosome and BMT. It seems likely that cytokinesis stops because a weak cell plate is formed that does not mature to a firm cell wall.
The duckweed Lemna minor L. clone St was used to investigate the effect of 10 heavy metals under the standardised test conditions of the ISO protocol 20079. By using growth rates derived from frond number (FN), fresh weight (FW), dry weight (DW), chlorophyll and carotenoid (Car) contents, concentration-response curves for all heavy metals and all growth parameters were classified. In addition, all data were fitted to obtain the inhibitions of growth rates (E(r)C(x)) at the level of 10%, 20% and 50% (E(r)C(10), E(r)C(20) and E(r)C(50), respectively) then used to evaluate the phytotoxicity of the different heavy metals. On the basis of the E(r)C(50) values (average ranking of all five growth parameters), the following series of phytotoxicity was detected by using molar concentrations: Ag(+)>Cd(2+)>Hg(2+)>Tl(+)>Cu(2+)>Ni(2+)>Zn(2+)>Co(2+)>Cr(VI)>As(III)>As(V).
Trichoderma harzianum is a fungus used as biocontrol agent using its antagonistic abilities against phytopathogenic fungi, although it has also direct effects on plants, increasing or accelerating their growth and resistance to diseases and the tolerance to abiotic stresses. We analyzed Arabidopsis thaliana gene expression changes after 24 h of incubation in the presence of T. harzianum T34 using the Affymetrix GeneChip Arabidopsis ATH1. Because this microarray contains more than 22,500 probe sets representing approximately 24,000 genes, we were able to construct a global picture of the molecular physiology of the plant at 24 h of T. harzianum-Arabidopsis interaction. We identified several differentially expressed genes that are involved in plant responses to stress, regulation of transcription, signal transduction or plant metabolism. Our data support the hypothesis that salicylic acid- and jasmonic acid-related genes were down-regulated in A. thaliana after 24 h of incubation in the presence of T. harzianum T34, while several genes related to abiotic stress responses were up-regulated. These systemic changes elicited by T. harzianum in Arabidopsis are discussed.
Brassinosteroids (BRs) are endogenous plant hormones essential for plant growth and development. Brassinosteroid insensitive1 (BRI1)-assocaiated receptor kinase (BAK1) is one of the key components in the BR signal transduction pathway due to its direct association with the BR receptor, BRI1. Although BRI1 and its orthologs have been identified from both dicotyledonous and monocotyledonous plants, less is known about BAK1 and its orthologs in higher plants other than Arabidopsis. This article provides the first piece of evidence that AtBAK1 can greatly affect growth and development of rice plants when ectopically expressed, suggesting that rice may share similar BR perception mechanism via BRI1/BAK1 complex. Interestingly, transgenic rice plants displayed semi-dwarfism and shortened primary roots. Physiological analysis and cell morphology assay demonstrated that the observed phenotypes in transgenic plants were presumably caused by hypersensitivity to endogenous levels of BRs, different from BR insensitive and deficient rice mutants. Consistently, several known BR inducible genes were also upregulated in transgenic rice plants, further suggesting that BAK1 was able to affect BR signaling in rice. On the other hand, the transgenic plants generated by overproducing AtBAK1 may potentially have agricultural applications because the dwarfed phenotype is generally resistant to lodging, while the fertility remains unaffected.
Green algae of the genus Dunaliella can adapt to hypersaline environments and are considered model organisms for salinity tolerance. In an EST analysis in Dunaliella viridis under salt stress, we isolated a salt-inducible cDNA coding for the 26S proteasome subunit RPN10, designated DvRPN10. The DvRPN10 cDNA is 1472 bp and encodes a polypeptide of 377 amino acids. The DvRPN10 protein shares a high similarity to orthologs from other species. The function of DvRPN10 was confirmed by complementation of the yeast Deltarpn10 mutant. Q-PCR analysis of D. viridis cells grown in different salinities revealed that the transcript level of DvRPN10 increased in proportion to the external salinity within a range of 0.5-3 M NaCl, but decreased significantly at extremely high salinities (4-5 M NaCl). When a salinity shock of 1-3 M NaCl was applied to D. viridis cells, DvRPN10 mRNA levels remained steady during the first 36 h, and then gradually elevated to the level observed at 3 M NaCl. The gene structure of DvRPN10 was revealed by sequencing of a BAC clone containing this gene. Possible transcription factor binding sites related to stress tolerance were found in the promoter region of DvRPN10. The expression of DvRPN10 in response to the external salinity suggests that RPN10-mediated protein degradation plays a role in the salinity tolerance of D. viridis.
The marine diatoms Bellerochea yucatanensis, Biddulphia sinensis, Ditylum brightwellii, Lauderia annulata and Thalassiosira rotula were grown for 2 days under different levels of UV-B radiation (439, 717, and 1230J · m (-2) ·d(-1)). UV-B stress depressed the growth rates of all species. A low UV-B dose (439 J · m (-2) · d (-1), weighted), usually caused a slight increase in biomass production (dry weight} in comparison to nontreated cells. Enhanced UV-B reduced the dry matter productivity of all diatoms. All marine diatoms exposed to UV-B showed a diminution of protein and pigment content (chlorophyll a, chlorophyll c(1) + C(2), and carotenoids). Algae grown in 20 %o or 35 %o salt concentrations were more sensitive to UV-B radiation than those grown in 45%o. S. The effect of higher UV-B dose (717J · m (-2) · d(-1), weighted) upon the pools of free amino acids was species-dependent. Aspartate and asparagine levels were reduced in all diatoms. A marked increase in glutamine levels was found in Bellerochea, Biddulphia, Ditylum and Lauderia. Thalassiosira cells exposed to UV-B showed a significant increase in glutamate levels and a reduction of glutamine levels. (15)N and (14)C incorporation into several amino acids was reduced by exposure to UV-B. The results are discussed with reference to the inhibition of the enzymes for carbon and nitrogen metabolism.
Cowpea (Vigna unguiculata) alternative oxidase is encoded by a small multigene family (Aox1, 2a and 2b) that is orthologous to the soybean Aox family. Like most of the identified Aox genes in plants, VuAox1 and VuAox2 consist of 4 exons interrupted by 3 introns. Alignment of the orthologous Aox genes revealed high identity of exons and intron variability, which is more prevalent in Aox1. In order to determine Aox gene expression in V. unguiculata, a steady-state analysis of transcripts involved in seed development (flowers, pods and dry seeds) and germination (soaked seeds) was performed and systemic co-expression of VuAox1 and VuAox2b was observed during germination. The analysis of Aox transcripts in leaves from seedlings under different stress conditions (cold, PEG, salicylate and H2O2 revealed stress-induced co-expression of both VuAox genes. Transcripts of VuAox2a and 2b were detected in all control seedlings, which was not the case for VuAox1 mRNA. Estimation of the primary transcript lengths of V. unguiculata and soybean Aox genes showed an intron length reduction for VuAox1 and 2b, suggesting that the two genes have converged in transcribed sequence length. Indeed, a bioinformatics analysis of VuAox1 and 2b promoters revealed a conserved region related to a cis-element that is responsive to oxidative stress. Taken together, the data provide evidence for co-expression of Aox1 and Aox2b in response to stress and also during the early phase of seed germination. The dual nature of VuAox2b expression (constitutive and induced) suggests that the constitutive Aox2b gene of V. unguiculata has acquired inducible regulatory elements.
1-Deoxy-d-xylulose 5-phosphate synthase (DXS, EC: 18.104.22.168), the first enzyme in the 2C-methyl-d-erythritol 4-phosphate (MEP) pathway, is known to be responsible for the rate-limiting step of isoprenoid biosynthesis in Escherichia coli and Arabidopsis thaliana. In this study, the dxs gene from Croton stellatopilosus, designated csdxs, was cloned from leaf tissue using the rapid amplification of cDNA ends (RACE) technique. Leaves of C. stellatopilosus contain plaunotol, an acyclic diterpene alcohol. The csdxs cDNA containing the open reading frame of 2163 base pairs appeared to encode a polypeptide of 720 amino acids. Analysis of the deduced amino acid sequence revealed that the NH(2)-terminus of CSDXS carried a chloroplast transit peptide, a thiamine diphosphate binding site, and a transketolase motif, which are the important characteristics of DXS enzymes in higher plants. Multiple alignments of CSDXS with other plant DXSs have indicated that CSDXS has identity ranging between 68% and 89%. Expression levels of csdxs and genes encoding key enzymes in the plaunotol biosynthetic pathway, namely 2C-methyl-d-erythritol 4-phosphate synthase (meps) and geranylgeranyl diphosphate synthase (ggpps), were analysed by measuring transcript levels in leaves of different developmental stages. The results showed that dxs, meps, and ggpps are all active in young leaves prior to full expansion when plaunotol is synthesised from the DXP precursor in chloroplasts. The dense presence of chloroplasts and oil globules in the palisade cells of these leaves support the view that these genes are involved in plaunotol biosynthesis in chloroplast-containing tissues.
Low temperature is one of the most common environmental stresses affecting plant growth and agricultural production. Serine/threonine protein phosphatases 2C (PP2Cs) have been suggested to play an important role in stress signaling. To identify potential new member of the PP2C proteins in maize and investigate its functions for stress responses, the ZmPP2C2 gene, encoding a new PP2C protein from maize roots, was cloned by RT-PCR and RACE-PCR. Its constitutive expression in roots, stems and leaves of maize seedlings was detected by RNA gel blot, and its regulation in response to cold stress was also examined. To further evaluate its function in the cold stress response, we over-expressed the ZmPP2C2 gene in tobacco under the control of the Cauliflower Mosaic Virus (CaMV) 35S promoter, and assessed a series of physiological changes in wild type and transgenic plants under low temperatures. Compared with wild type tobacco under cold stress, plants that over-expressed ZmPP2C2 displayed higher germination speed and rate, higher antioxidant enzyme (SOD, POD, CAT) activities, with lower cold-induced electrolyte leakage and malondialdehyde (MDA) contents. These results show that over-expression of ZmPP2C2 in tobacco enhanced tolerance to cold stress, suggesting that this new gene, ZmPP2C2, may act as a positive regulator of cold resistance in plants.
The PR-2d promoter/uidA (GUS) gene construct was introduced into the cucumber (Cucumis sativus L.) genome and several transgenic lines were produced. Activation of the PR-2d promoter was investigated in these plants in response to inoculation with fungal pathogens and after salicylic acid (SA) or cold treatments. Treatment with exogenous SA increased GUS activity 2 to 11 fold over that of the control. Endogenous SA and its conjugate salicylic acid glucoside (SAG) rose in parallel after inoculation with the fungal pathogen Pseudoperonospora cubensis, with SAG becoming the predominant form. The free SA levels increased 15 fold above the basal level at 5 dpi and preceded the induction of the PR-2d promoter by five days, which occurred at 10 dpi with a 12 fold increase over the control. Inoculation with another fungal pathogen, Erysiphe polyphage, increased GUS activity 4 to 44 fold over that of the control. During normal development of flowers in the cucumber, the PR-2d/uidA gene expressed in the floral organs was similar to that of the primary host. In addition, we present the first evidence that the PR-2d promoter was induced (624 fold) under cold stress. We demonstrate that in the heterologous state the gene construct was expressed according to the signalling pattern of the native species and was stably transmitted to progeny over four generations.
It is postulated that leaf thermonasty (leaf curling) in rhododendrons under sub-freezing temperatures is caused by water redistribution due to extracellular freezing. We hypothesize that aquaporins (AQPs), the transmembrane water-channels, may be involved in regulating water redistribution and thus leaf curling. Our experimental system includes two Rhododendron species with contrasting leaf curling behavior whereby it was observed in R. catawbiense but not in R. ponticum. We compared leaf movements and the expression of two AQPs, i.e. R. catawbiense/ponticum plasma-membrane intrinsic protein 2 (Rc/RpPIP2;1 and Rc/RpPIP2;2), in the two species under freezing-rewarming and dehydration-rehydration cycles. To determine the relationship between extracellular freezing and leaf-curling, we monitored leaf-curling in R. catawbiense with or without controlled ice-nucleation. Our data indicate that extracellular freezing may be required for leaf curling. Moreover, in both species, PIP2s were up-regulated at temperatures that fell in ice-nucleation temperature range. Such up-regulation could be associated with the bulk-water efflux caused by extracellular freezing. When leaves were frozen beyond the ice-nucleation temperature range, PIP2s were continuously down-regulated in R. catawbiense along with the progressive leaf curling, as also observed for RcPIP2;2 in dehydrated leaves; as leaves uncurled during re-warming/rehydration, RcPIP2 expression was restored. On the other hand, R. ponticum, a non-curling species, exhibited substantial up-regulation of RpPIP2s during freezing/dehydration. Taken together, our data suggest that RcPIP2 down-regulation was associated with leaf curling. Moreover, the contrasting PIP2 expression patterns combined with leaf behavior of R. catawbiense and R. ponticum under these two cycles may reflect different strategies employed by these two species to tolerate/resist cellular dehydration.
The application of nitrogen fertilisers leads to different ecological problems such as nitrate leaching and the release of nitrogenous gases. N2O is a gas involved in global warming, therefore, agricultural soils can be regarded as a source of global warming. Soil N2O production comes from both the nitrification and denitrification processes. From an ecological viewpoint, using nitrification inhibitors with ammonium based fertilisers may be a potential management strategy to lower the fluxes of N2O, thus decreasing its undesirable effect. In this study, the nitrification inhibitors (NIs) dicyandiamide (DCD) and 3,4-dimethyl pyrazole phosphate (DMPP) have been evaluated as management tools to mitigate N2O emissions from mineral fertilisation and slurry application in grassland systems (experiments 1 and 2), and to assess the phytotoxic effect of these inhibitors per se on clover (experiment 3). Both nitrification inhibitors acted in maintaining soil nitrogen (N) in ammonium form, decreasing cumulative N2O emissions. DCD, but not DMPP, produced phytotoxic effects and yield reduction in white clover. A nutrient imbalance, which led to a senescence process visually observed as chlorosis and necrosis at the border of the leaves, was noted.
The uncommon amino acid 3,4-dihydroxyphenylalanine (DOPA) was determined colorimetrically using isonicotinic acid hydrazide (IAH) in the crude extracts obtained from fresh material of Vicia faba minor. DOPA was found to accumulate at concentrations inconsistent with its solubility in water. Other orthodiphenols do not interfere in DOPA determination. The accumulated DOPA was only of the L-form. The DOPA content in the dry material of Vicia faba was significantly lower than that from the corresponding fresh material, suggesting some kind of DOPA oxidation during drying. Subsequent analyses designed to evaluate the actual potential of V. faba minor in the accumulation of DOPA were performed with fresh material. It was observed that the DOPA concentration in the tissues of V. faba minor is reproducible when expressed on a fresh matter weight basis. In young seedlings L-DOPA production is significantly enhanced by light via an increased fresh matter yield. The DOPA content was found to be increased in mature plants grown on media enriched with low concentrations of NaCl (up to 25 mM) that showed a positive effect on their growth. NaCl induced an overproduction of L-DOPA even when growth was restricted by ion excess. On the contrary, the DOPA level dropped in mature plants submitted to water stress. Nodulated plants were found to contain higher content of L-DOPA than the non-nodulated ones grown with NO(3)(-) and NH(4)(+). It is suggested that DOPA hyperproduction by Vicia faba minor might be optimized by growing plants in mild saline conditions and by inoculating them with efficient salt tolerant strains of R. leguminosarum.
The graviresponsiveness of intact and primary maize roots kept horizontally in darkness and humid air is analysed. A precise local application of IAA is possible when using resin beads (diameter: 0.45 +/- 0.05 mm) loaded with IAA. The beads are placed on the upper or lower sides of the caps. They significantly change the root gravireaction. The effect of IAA is discussed in terms of its possible level in the growing and gravibending zones and its transport (acropetal, lateral and basipetal) respectively in the stele, the cap and the cortex of the elongating root.
Microautoradiographic methods were applied to trace (86)Rb and (43)K during the migration within the transpiration stream of Trifolium repens L. and Oxalis acetosella L. During the dark phase of the diurnal cycle, ions moved through the petiole to the insertion of the leaf blade. There the movement stopped. In Oxalis the ions gathered in three distinct spots at the adaxial side of the insertion, each belonging to one of the downwards bending leaflets. In Trifolium the areas of ion accumulation were two little humps at the abaxial periphery of the nodal junction of the inwardly bending leaflets. During the phases of upward movement of the leaflets, Oxalis did not reveal a specific pattern of ion distribution within the pulvinus. However, when Trifolium was loaded during the phases of leaflet opening, (86)Rb and (43)K were mainly found within the parenchymatous bundle sheath of the three laminar pulvini and their junction towards the petiole. The conclusion was that ions are stored within small cells on the flexor side and are removed from there by regulatory processes controlling water and solute flux from the sites of storage to the evaporating surfaces of the leaf blade. Implications on the interpretation of nyctinastic leaf movements are discussed.
Microautoradiographic methods applied to the pulvinar region of Phaseolus vulgaris L. revealed specific patterns of ion distribution according to the phases of upward or downward movements of the leaves: When feeding was taking place during the dark phase of the diurnal cycle, the main source of radioactivity was the dorsal groove of the vascular core on the flexor side. During the phase of leaf opening, (86)Rb and (43)K were mainly found within the parenchymatous bundle sheath. As an additional similarity to the guard cell physiology, toxic metal ions were accumulated in the cells of the bundle sheath when introduced into the plant during the light phase. Additional experiments on the influence of osmotic gradients supported the view that the characteristic distribution of ions during the two phases of the diurnal cycle is the result of regulatory processes controlling water and solute flux from the sites of storage to the evaporating surfaces. The conclusion was that in light the cells of the bundle sheath removed the ions from the flexor sites and that this process is substantially controlled by transpiration rates. Findings of other authors were discussed in view of the hypothesis that K(+) movements in the pulvinar region depend on regulatory systems other than active ion pumps. Among the processes probably involved in the initiation of primary water flux changes are the import of carbohydrates into the cells of the bundle sheath and starch hydrolysis.