Purification of the Trehalase GMTRE1 from Soybean Nodules and Cloning of Its cDNA. GMTRE1 Is Expressed at a Low Level in Multiple Tissues1

Trehalose (α-d-glucopyranosyl-1,1-α-d-glucopyranoside), a disaccharide widespread among microbes and lower invertebrates, is generally believed to be nonexistent in higher plants. However, the recent discovery of Arabidopsis genes whose products are involved in trehalose synthesis has renewed interest in the possibility of a function of trehalose in higher plants. We previously showed that trehalase, the enzyme that degrades trehalose, is present in nodules of soybean (Glycine max [L.] Merr.), and we characterized the enzyme as an apoplastic glycoprotein. Here we describe the purification of this trehalase to homogeneity and the cloning of a full-length cDNA encoding this enzyme, named GMTRE1 (G. max trehalase 1). The amino acid sequence derived from the open reading frame of GMTRE1 shows strong homology to known trehalases from bacteria, fungi, and animals. GMTRE1 is a single-copy gene and is expressed at a low but constant level in many tissues.

Differential Expression of Alternative Oxidase Genes in Soybean Cotyledons during Postgerminative Development1

The expression of the alternative oxidase (AOX) was investigated during cotyledon development in soybean (Glycine max [L.] Merr.) seedlings. The total amount of AOX protein increased throughout development, not just in earlier stages as previously thought, and was correlated with the increase in capacity of the alternative pathway. Each AOX isoform (AOX1, AOX2, and AOX3) showed a different developmental trend in mRNA abundance, such that the increase in AOX protein and capacity appears to involve a shift in gene expression from AOX2 to AOX3. As the cotyledons aged, the size of the mitochondrial ubiquinone pool decreased. We discuss how this and other factors may affect the alternative pathway activity that results from the developmental regulation of AOX expression.

Release of Flavonoids by the Soybean Cultivars McCall and Peking and Their Perception as Signals by the Nitrogen-Fixing Symbiont Sinorhizobium fredii1

Sinorhizobium fredii strain USDA191 forms N-fixing nodules on the soybean (Glycine max L. Merr.) cultivars (cvs) McCall and Peking, but S. fredii strain USDA257 nodulates only cv Peking. We wondered whether specificity in this system is conditioned by the release of unique flavonoid signals from one of the cultivars or by differential perception of signals by the strains. We isolated flavonoids and used nodC and nolX, which are nod-box-dependent and -independent nod genes, respectively, to determine how signals activate genes in the microsymbionts. Seeds of cv McCall and cv Peking contain the isoflavones daidzein, genistein, and glycitein, as well as their glucosyl and malonylglucosyl glycosides. Roots exude picomolar concentrations of daidzein, genistein, glycitein, and coumestrol. Amounts are generally higher in cv Peking than in cv McCall, and the presence of rhizobia markedly influences the level of specific signals. Nanomolar concentrations of daidzein, genistein, and coumestrol induce expression of nodC and nolX in strain USDA257, but the relative nolX-inducing activities of these signals differ in strain USDA191. Glycitein and the conjugates are inactive. Strain USDA257 deglycosylates daidzin and genistin into daidzein and genistein, respectively, thereby converting inactive precursors into active inducers. Although neither soybean cultivar contains unique nod-gene-inducing flavonoids, strain- and cultivar-specific interactions are characterized by distinct patterns of signal release and response.

Specific Soybean Lipoxygenases Localize to Discrete Subcellular Compartments and Their mRNAs Are Differentially Regulated by Source-Sink Status1

Members of the lipoxygenase multigene family, found widely in eukaryotes, have been proposed to function in nitrogen partitioning and storage in plants. Lipoxygenase gene responses to source-sink manipulations in mature soybean (Glycine max [L.] Merr.) leaves were examined using gene-specific riboprobes to the five vegetative lipoxygenases (vlxA–vlxE). Steady-state levels of all vlx mRNAs responded strongly to sink limitation, but specific transcripts exhibited differential patterns of response as well. During reproductive sink limitation, vlxA and vlxB messages accumulated to high levels, whereas vlxC and vlxD transcript levels were modest. Immunolocalization using peptide-specific antibodies demonstrated that under control conditions, VLXB was present in the cytosol of the paraveinal mesophyll and with pod removal accumulated additionally in the bundle-sheath and adjacent cells. With sink limitation VLXD accumulated to apparent high levels in the vacuoles of the same cells. Segregation of gene products at the cellular and subcellular levels may thus permit complex patterns of differential regulation within the same cell type. Specific lipoxygenase isoforms may have a role in short-term nitrogen storage (VLXC/D), whereas others may simultaneously function in assimilate partitioning as active enzymes (VLXA/B).

Identification of Inositol 1,3,4-Trisphosphate 5-Kinase and Inositol 1,3,4,5-Tetrakisphosphate 6-Kinase in Immature Soybean Seeds

In extracts of immature soybean (Glycine max [L.] Merr.) seeds inositol tetrakisphosphate was formed from [3H]inositol 1,3,4-trisphosphate but not from [3H]inositol 1,4,5-trisphosphate. Inositol 1,3,4-trisphosphate kinase was purified to a specific activity of 3.55 min−1 mg−1 by polyethylenimine clarification and anion-exchange chromatography. The partially purified enzyme converted [3H]inositol 1,3,4-trisphosphate to inositol 1,3,4,5-tetrakisphosphate as the major product and inositol 1,3,4,6- and/or 1,2,3,4-tetrakisphosphate as the minor product. Subsequent experiments revealed a separate inositol 1,3,4,5-tetrakisphosphate 6-kinase activity, which could link these enzymes to inositol hexakisphosphate synthesis via the previously reported inositol 1,3,4,5,6-pentakisphosphate 2-kinase. The apparent Km values for inositol 1,3,4-trisphosphate kinase were 200 ± 0 nm for inositol 1,3,4-trisphosphate and 171 ± 4 μm for ATP, and the reaction was not reversible. The kinetics were such that no activity could be detected using unlabeled inositol 1,3,4-trisphosphate and [γ-32P]ATP, which suggested that other kinases may have been observed when less purified fractions were incubated with radiolabeled ATP. Inositol 1,3,4-trisphosphate kinase was nonspecifically inhibited more than 80% by various inositol polyphosphates at a concentration of 100 μm.

Resistance gene analogs are conserved and clustered in soybean.

Sequences of cloned resistance genes from a wide range of plant taxa reveal significant similarities in sequence homology and structural motifs. This is observed among genes conferring resistance to viral, bacterial, and fungal pathogens. In this study, oligonucleotide primers designed for conserved sequences from coding regions of disease resistance genes N (tobacco), RPS2 (Arabidopsis) and L6 (flax) were used to amplify related sequences from soybean [Glycine max (L.) Merr.]. Sequencing of amplification products indicated that at least nine classes of resistance gene analogs (RGAs) were detected. Genetic mapping of members of these classes located them to eight different linkage groups. Several RGA loci mapped near known resistance genes. A bacterial artificial chromosome library of soybean DNA was screened using primers and probes specific for eight RGA classes and clones were identified containing sequences unique to seven classes. Individual bacterial artificial chromosomes contained 2-10 members of single RGA classes. Clustering and sequence similarity of members of RGA classes suggests a common process in their evolution. Our data indicate that it may be possible to use sequence homologies from conserved motifs of cloned resistance genes to identify candidate resistance loci from widely diverse plant taxa.

Do decomposing organic matter residues reduce phosphorus sorption in highly weathered soils?

Many studies have shown a reduction in P sorption in highly weathered soils when organic matter (OM) is applied, suggesting competition between OM decomposition products and P for sorption sites. However, such studies seldom consider the P released from the added OM. To delineate the effects of OM addition on P availability through sorption competition and P addition, water leachate from incubated soybean (SB) [Glycine mar (L.) Merr.] and Rhodes grass (RG) (Chloris gayana Knuth cv. Callide) was used in competitive P sorption studies both undiluted and after acidification (i.e., the fulvic acid [FA] component). Addition of two rates (0.2 and 2 mL) of SB leachate to an Oxisol significantly increased P sorption at the higher rate, while a similar trend was observed following RG leachate addition at the same rates. Extending the range of highly weathered soils examined (two Oxisols, an Ultisol, and an acidic Vertisol) resulted in no observed decrease in P sorption following addition of OM leachate. Surprisingly, SB leachate transiently increased P sorption in the two Oxisol soils. Addition of the FA component of the leachates resulted in a transient (< 6 d) decrease in P sorption in three of the four soils examined and constituted the only evidence in this study that decomposing OM residues reduced P sorption. This research provides further evidence contradicting the long held assumption that inhibition of P sorption by dissolved organic compounds, derived from decomposing OM, is responsible for increased P phytoavailability when P fertilizer and OM are applied together.

Developing pineapple fruit has a small transcriptome dominated by metallothionein

In a first step toward understanding the molecular basis of pineapple fruit development, a sequencing project was initiated to survey a range of expressed sequences from green unripe and yellow ripe fruit tissue. A highly abundant metallothionein transcript was identified during library construction, and was estimated to account for up to 50% of all EST library clones. Library clones with metallothionein subtracted were sequenced, and 408 unripe green and 1140 ripe yellow edited EST clone sequences were retrieved. Clone redundancy was high, with the combined 1548 clone sequences clustering into just 634 contigs comprising 191 consensus sequences and 443 singletons. Half of the EST clone sequences clustered within 13.5% and 9.3% of contigs from green unripe and yellow ripe libraries, respectively, indicating that a small subset of genes dominate the majority of the transcriptome. Furthermore, sequence cluster analysis, northern analysis, and functional classification revealed major differences between genes expressed in the unripe green and ripe yellow fruit tissues. Abundant genes identified from the green fruit include a fruit bromelain and a bromelain inhibitor. Abundant genes identified in the yellow fruit library include a MADS box gene, and several genes normally associated with protein synthesis, including homologues of ribosomal L10 and the translation factors SUI1 and eIF5A. Both the green unripe and yellow ripe libraries contained high proportions of clones associated with oxidative stress responses and the detoxification of free radicals.

Legume nodulation: successful symbiosis through short- and long-distance signalling

Nodulation in legumes provides a major conduit of available nitrogen into the biosphere. The development of nitrogen-fixing nodules results from a symbiotic interaction between soil bacteria, commonly called rhizobia, and legume plants. Molecular genetic analysis in both model and agriculturally important legume species has resulted in the identification of a variety of genes that are essential for the establishment, maintenance and regulation of this symbiosis. Autoregulation of nodulation (AON) is a major internal process by which nodule numbers are controlled through prior nodulation events. Characterisation of AON-deficient mutants has revealed a novel systemic signal transduction pathway controlled by a receptor-like kinase. This review reports our present level of understanding on the short- and long-distance signalling networks controlling early nodulation events and AON.

Structure repair of a compacted vertisol with wet-dry cycles and crops

We hypothesized that the four rotation crops: wheat (Triticum aestivum L.), sorghum [Sorghum bicolor (L.) Merr.], lablab [Lablab purpureus (L.) Sweet] and mung bean [ Vigna radiata (L.) R. Wilczek] differ in their ability to repair soil structure. The study was conducted on a Typic Haplustert, Queensland, Australia, locally termed a Black Earth and considered a prime cropping soil. Large (0.5-m depth by 0.3-m diam.) soil cores, collected from compacted wheel furrows in an irrigated cotton (Gossypium hirsutum L.) field, were subjected to three, six, or nine wet-dry cycles that simulated local flood irrigation practices. After each cycle, soil profiles were sampled for clod bulk density, image analysis of soil structure, and evapotranspiration. Generally, all crops improved soil structure over the initial field condition but lablab and mung bean gave improvements to greater depths and more rapidly than wheat and sorghum. Mung bean and lablab caused up to a threefold increase in clod porosity in the 0.1- to 0.4-m soil layer after only three wet-dry cycles, whereas sorghum required nine wet-dry cycles to increase clod porosity in only the 0.2- to 0.3-m layer, and wheat gave no improvement even after nine wet-dry cycles. Image analysis of soil structure showed that lablab and mung bean rapidly (by three wet-dry cycles) produced smaller peds with more interconnected pore space than wheat and sorghum. By nine wet-dry cycles, sorghum achieved deep cracking of the soil but the material between the cracks remained large and dense. Evapotranspiration was double under lablab and mung bean compared with wheat and sorghum. Our results indicate greater cycles of wetting and drying under lablab and mung bean than wheat and sorghum that have led to rapid repair of soil compaction.

Analyses of the community compositions of root rot pathogenic fungi in the soybean rhizosphere soil

Soybean ( Glycine max [L.] Merr.) root rot is an important disease of soybean under continuous cropping, and root rot is widely distributed throughout the world. This disease is extremely harmful, and it is difficult to prevent and control. The study aimed to elucidate the composition of root rot pathogenic fungal communities in the continuous cropping of soybean. In this study, we employed PCRDGGE technology to analyze the communities of root rot pathogenic fungi in soybean rhizosphere soil subjected to continuous cropping during a season with a high incidence of root rot in Heilongjiang province, China, the main soybean producing area in China. The results of 13 DGGE bands were analyzed by phylogenetic revealed that the predominant root rot pathogenic fungi in rhizosphere soil in the test area were Pythium ultimum and Fusarium species. The results of cluster analysis showed that the duration of continuous cropping, the soybean variety and the plant growth stage all had significant effects on the diversity of root rot pathogenic fungi in rhizosphere soil.

Efeito de indutores de florescimento nas cultivares de abacaxizeiro RBR-1, SNG-2 e SNG-3 em Rio Branco-Acre.

Este trabalho teve como objetivo avaliar o efeito de indutores de florescimento em três cultivares de abacaxizeiro em Rio Branco-Acre. O delineamento experimental foi em blocos ao acaso, em esquema fatorial 3 x 2 x 2, com quatro repetições, sendo combinadas três cultivares de abacaxi (RBR-1, SNG-2 e SNG-3), dois indutores de florescimento (carbureto de cálcio-CaC2 e etefon-ácido 2-cloroetilfosfônico) e duas épocas de aplicação (aos 10 e 12 meses do plantio). As plantas induzidas com etefon aos 10 meses apresentaram maior percentagem de florescimento (96,25%) quando com CaC2 (85,42%) quando comparadas com as induzidas aos 10 meses (62,92%).

Functional aspects of floral nectar secretion of Ananas ananassoides, an ornithophilous bromeliad from the Brazilian savanna

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Differences in gastroprotective and mutagenic actions between polar and apolar extracts of Ananas ananassoides

Several plants are used in folk medicine to treat gastrointestinal disorders. Ananas ananassoides (Baker) L. B. Smith (Family Bromeliaceae) is a medicinal plant commonly used in the central region of Brazil against gastric pain. We evaluated two extracts (methanol [MeOH] and dichloromethane [DCM]) obtained from the leaves of A. ananassoides for their ability to protect the gastric mucosa against injuries caused by necrotizing agents (0.3 M HCl/60% ethanol, absolute ethanol, non-steroidal anti-inflammatory drugs, and pylorus ligation) in mice and rats. The best results were obtained after pretreatment with the DCM extract, whereas the MeOH extract did not show any significant anti-ulcerogenic activity but presented mutagenic action. The mechanism of action of the DCM extract suggested the effective participation of endogenous sulfhydryl group in the gastroprotective action. The data, taken together with the absence of acute toxicity and mutagenicity, indicate the apolar extract, instead of the polar, extract of A. ananassoides as a safe and potential new anti-ulcerogenic drug.

Occurrence Of Isoflavonoids In Brazilian Common Bean Germplasm (phaseolus Vulgaris L.).

Common bean (Phaseolus vulgaris) is present in the daily diet of various countries and, as for other legumes, has been investigated for its nutraceutical potential. Thus, 16 genotypes from different gene pools, representing seven types of seed coats and different responses to pathogens and pests, were selected to verify their isoflavone contents. The isoflavonoids daidzein and genistein and the flavonols kaempferol, myricetin, and quercetin were found. Grains of the black type showed the highest concentrations of isoflavonoids and were the only ones to exhibit daidzein. IAC Formoso, with high protein content and source of resistance to anthracnose, showed the greatest concentration of genistein, representing around 11% of the content present in soybean, as well as high levels of kaempferol. Arc 1, Raz 55, and IAC Una genotypes showed high content of coumestrol. The results suggest the use of IAC Formoso to increase the nutraceutical characteristics in common bean.