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PubMed1981

Is wheat mitochondrial 5S ribosomal RNA prokaryotic in nature?

Küntzel et al. (1981) (Nucleic Acids Res. 9, 1451-1461) recently concluded that the sequence of wheat mitochondrial 5S rRNA is significantly more related to prokaryotic than to eukaryotic 5S rRNA sequences, and displays an especially high affinity to that of the thermophilic Gram-negative bacterium, Thermus aquaticus. However, the sequence on which this conclusion was based, although attributed to us, differs in several places from the one determined by us. We show here that the correct sequence (Spencer, D.F., Bonen, L. and Gray, M.W. (1981) Biochemistry, in press) does not support the conclusions of Küntzel et al. about potential secondary structure in wheat mitochondrial 5S rRNA and its phylogenetic significance. We further show that when the wheat mitochondrial 5S rRNA sequence is matched against published alignments for E. coli, T. aquaticus, and wheat cytosol 5S rRNAs, the mitochondrial sequence shows no greater homology to the T. aquaticus sequence than to the E. coli sequence, and only slightly more homology to these two sequences than to wheat cytosol 5S rRNA. This analysis confirms our original view (Biochemistry, in press) that wheat mitochondrial 5S rRNA is neither obviously prokaryotic nor eukaryotic in nature, but shows characteristics of both classes of 5S rRNA, as well as some unique features.

Base Sequence↗

PubMed2005

Gene expression studies in isolated mitochondria: Solanum tuberosum rps10 is recognized by cognate potato but not by the transcription, splicing and editing machinery of wheat mitochondria.

The complex gene expression mechanisms that occur in plant mitochondria, such as RNA editing and splicing, are not yet well understood. RNA editing in higher plant mitochondria is a highly specific process which modifies mRNA sequences by C-to-U conversions. It has been suggested that in some cases this process is required for splicing. Here, we use an experimental model based on the introduction of DNA into isolated mitochondria by electroporation to study organellar gene expression events. Our aim was to compare processing and editing of potato small ribosomal protein 10 gene (rps10) transcripts in heterologous (wheat mitochondria) and homologous (potato mitochondria) contexts. rps10 is a suitable model because it contains a group II intron, is absent in wheat mitochondria but is actively expressed in potato mitochondria, where transcripts are spliced and undergo five C-to-U editing events. For this purpose, conditions for electroporating isolated potato mitochondria were established. rps10 was placed under the control of either potato or wheat cox2 promoters. We found that rps10 was only transcribed under the control of a cognate promoter. In wheat mitochondria, rps10 transcripts were neither spliced nor edited while they are correctly processed in potato mitochondria. Interestingly, a wheat editing site grafted into rps10 was not recognized by wheat mitochondria but was correctly edited in potato mitochondria. Taken together, these results suggest that editing might occur only when the transcripts are engaged in processing and that they would not be available to editing factors outside of a putative RNA maturation machinery complex.

Electron Transport Complex IV↗

PubMed1999

Effects of phytase supplementation, individually and in combination, with glycanase, on the nutritive value of wheat and barley.

Four experiments were conducted to examine the effects of a microbial phytase (Natuphos), individually and in combination with glycanase preparations with predominantly xylanase (Natugrain Blend) and glucanase (Natugrain) activities, on the nutritive value of wheat and barley. In Experiment 1, the addition of xylanase and phytase increased the AME of a low-AME wheat by 9.7 and 5.3%, respectively. The differences, however, were not significant (P > 0.05). The combination of the two enzymes increased (P < 0.05) the AME of wheat by 19.0% from 2,646 to 3,149 kcal/kg dry matter. A similar trend was seen in terms of ileal amino acid digestibility values of the wheat-casein diet. In Experiment 2, the AME of normal wheat was increased (P < 0.05) by 6.3 and 4.5%, respectively, with the addition of xylanase and phytase. The combination of the two enzymes, however, did not further improve (P > 0.05) the AME values. In Experiment 3, performance of broilers fed a wheat-based diet was not influenced by the addition of individual enzymes, but increasing inclusion levels of the xylanase plus phytase combination linearly improved weight gain (r = 0.58; P < 0.01) and feed efficiency (r = 0.71; P < 0.001). In Experiment 4, the AME of barley was not influenced by the addition of glucanase or phytase. The enzyme combination marginally (P < 0.07) improved the AME at lower concentrations, but had no benefit at the highest concentration.

6-Phytase↗

PubMed2003

Effects of whole wheat feeding on the development of coccidial infection in broiler chickens.

A complete ground and pelleted feed was compared to free choice feeding of whole wheat and a pelleted protein concentrate during three experimental infections with coccidia in broiler chickens. At 22 d of age birds were inoculated with different doses of sporulated oocysts of a cecal species (Eimeria tenella) in experiment 1 or intestinal species E. maxima or E. acervulina in experiments 2 and 3, respectively. The effects of diets were assessed on weight gain, hematocrit (during cecal coccidiosis), serum coloration (during intestinal coccidiosis), oocyst excretion, and lesion score until 7 d post-inoculation. In experiment 1 before inoculation, the birds fed whole grain had more beneficial microflora with lower counts of coliform bacteria. As shown by oocyst output and lesion score, whole wheat feeding increased parasite development during infection with the cecal parasite E. tenella. This led to significantly lower weight gain with whole wheat than with ground wheat from 5 to 6 d post-inoculation and to lower hematocrit at the highest infective dose. Parasite development in experiments 2 and 3 was similar among diets, during intestinal infection with E. maxima and E. acervulina, respectively, with no significant differences in lesion score. During the acute phase (4 to 7 and 3 to 5 d post-inoculation), when a difference appeared between diets, whole wheat fed-birds were always more affected than ground diet-fed birds in terms of serum coloration and weight gain. These results might be explained by modifications of digestive physiology and intestinal microflora by whole wheat.

Animal Feed↗

PubMed2003

The ABC transporter MgAtr4 is a virulence factor of Mycosphaerella graminicola that affects colonization of substomatal cavities in wheat leaves.

The role in virulence of the ATP-binding cassette (ABC) transporters MgAtr1, MgAtr2, MgAtr3, MgAtr4, and MgAtr5 from Mycosphaerella graminicola was analyzed by gene disruption or replacement on seedlings of the susceptible wheat cultivar Obelisk. Disruption strains of MgAtr1 and MgAtr2 and replacement strains of MgAtr3 and MgAtr5 displayed the same phenotype as control strains, while virulence of the MgAtr4 disruption strains was significantly reduced. This reduction in virulence was independent of the wheat cultivar used. Histopathological analysis of the infection process revealed that MgAtr4 disruption strains colonize substomatal cavities less efficiently and display reduced intercellular growth in the apoplast of wheat leaves. In vitro growth experiments in different media showed no fitness penalty associated with the disruption of MgAtr4. Expression analysis demonstrated that transcripts of the constitutively expressed gene CYP51 encoding the fungal-specific cytochrome P450 sterol 14alpha-demethylase from M. graminicola were not detectable in interaction RNA from wheat infected with MgAtr4 disruption strains, thus confirming the reduced intercellular growth of these strains. The results indicate that MgAtr4 is a virulence factor of M. graminicola during pathogenesis on wheat and may function in protection against fungitoxic compounds present around the substomatal cavities of wheat leaves. MgAtr4 is the first virulence factor cloned from this important plant pathogen.

ATP-Binding Cassette Transporters↗

PubMed1999

Protection against cancer by wheat bran: role of dietary fibre and phytochemicals.

Human intervention and animal studies have shown that supplementing the diet with wheat bran can protect against the development of a range of cancers, especially those of the colon and breast. Wheat bran is a rich source of dietary fibres (plant cell walls) that have structures and compositions which indicate that they may protect against cancer. Nevertheless, dietary fibre makes up less than half of wheat bran. Other nutrients and phytochemicals are present in wheat bran, some of which may also protect against cancer. These include phytic acid and various phenolic components such as phenolic acids, lignans and flavonoids. A major goal of future research on wheat bran should be to determine the relative roles in cancer prevention of the different components in wheat bran.

Animals↗

PubMed2006

Simulation of fertilizer requirement for irrigated wheat in eastern India using the QUEFTS model.

Crop modeling can provide us with information about fertilizer dose to achieve the target yield, crop conditions, etc. Due to conventional and imbalanced fertilizer application, nutrient use efficiency in wheat is low. Estimation of fertilizer requirements based on quantitative approaches can assist in improving yields and nutrient use efficiency. Field experiments were conducted at 20 sites in eastern India (Nadia district of West Bengal) to assess the soil supply, requirement, and internal efficiency of N, P, K, and Zn in wheat. The data were used to calibrate the QUEFTS (Quantitative Evaluation of the Fertility of Tropical Soils) model for site-specific, balanced fertilizer recommendations. The parameters of maximum accumulation (a) and maximum dilution (d) in wheat were calculated for N (35, 100), P (129, 738), K (17, 56), and Zn (21502, 140244). Grain yield of wheat showed statistically significant correlation with N (R2 = 0.937**), P (R2 = 0.901**), and K uptake (R2 = 0.801**). The NPK ratio to produce 1 tonne grain yield of wheat was calculated to be 4.9:1.0:8.9. The relationships between chemical properties and nutrient-supplying capacity of soils were also established. The model was validated using the data from four other experiments. Observed yields with different amounts of N, P, K, and Zn were in good agreement with the predicted values, suggesting that the validated QUEFTS model can be used for site-specific nutrient management of wheat.

Agriculture↗

PubMed1993

Wheat (Triticum aestivum L.) [gamma]-Gliadin Accumulates in Dense Protein Bodies within the Endoplasmic Reticulum of Yeast.

Following their sequestration into the endoplasmic reticulum (ER), wheat storage proteins may either be retained and packaged into protein bodies within this organelle or transported via the Golgi to vacuoles. We attempted to study the processes of transport and packaging of wheat storage proteins using the heterologous expression system of yeast. A wild-type wheat [gamma]-gliadin, expressed in the yeast cells, accumulated mostly within the ER and was deposited in protein bodies with similar density to natural protein bodies from wheat endosperm. This suggested that wheat storage proteins contain sufficient information to initiate the formation of protein bodies in the ER of a heterologous system. Only a small amount of the [gamma]-gliadin was transported to the yeast vacuoles. When a deletion mutant of the [gamma]-gliadin, lacking the entire N-terminal repetitive region, was expressed in the yeast cells, the mutant was unable to initiate the formation of protein bodies within the ER and was completely transported to the yeast vacuole. This strongly indicated that the information for packaging into dense protein bodies within the ER resides in the N-terminal repetitive region of the [gamma]-gliadin. The advantage of using yeast to identify the signals and mechanisms controlling the transport of wheat storage proteins and their deposition in protein bodies is discussed.

Journal Article↗

PubMed2006

RNA interference-based gene silencing as an efficient tool for functional genomics in hexaploid bread wheat.

Insertional mutagenesis and gene silencing are efficient tools for the determination of gene function. In contrast to gain- or loss-of-function approaches, RNA interference (RNAi)-induced gene silencing can possibly silence multigene families and homoeologous genes in polyploids. This is of great importance for functional studies in hexaploid wheat (Triticum aestivum), where most of the genes are present in at least three homoeologous copies and conventional insertional mutagenesis is not effective. We have introduced into bread wheat double-stranded RNA-expressing constructs containing fragments of genes encoding Phytoene Desaturase (PDS) or the signal transducer of ethylene, Ethylene Insensitive 2 (EIN2). Transformed plants showed phenotypic changes that were stably inherited over at least two generations. These changes were very similar to mutant phenotypes of the two genes in diploid model plants. Quantitative real-time polymerase chain reaction revealed a good correlation between decreasing mRNA levels and increasingly severe phenotypes. RNAi silencing had the same quantitative effect on all three homoeologous genes. The most severe phenotypes were observed in homozygous plants that showed the strongest mRNA reduction and, interestingly, produced around 2-fold the amount of small RNAs compared to heterozygous plants. This suggests that the effect of RNAi in hexaploid wheat is gene-dosage dependent. Wheat seedlings with low mRNA levels for EIN2 were ethylene insensitive. Thus, EIN2 is a positive regulator of the ethylene-signaling pathway in wheat, very similar to its homologs in Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa). Our data show that RNAi results in stably inherited phenotypes and therefore represents an efficient tool for functional genomic studies in polyploid wheat.

Carotenoids↗

PubMed1995

Insect cell expression of recombinant imidazoleglycerolphosphate dehydratase of Arabidopsis and wheat and inhibition by triazole herbicides.

Imidazoleglycerolphosphate dehydratase (IGPD; EC 4.2.1.19), which is involved in the histidine biosynthetic pathway of Arabidopsis thaliana and wheat (Triticum aestivum), has been expressed in insect cells using the baculovirus expression vector system. N-terminal amino acid sequencing indicated that recombinant IGPDs (rIGPDs) were produced as mature forms via nonspecific proteolytic cleavages in the putative transit peptide region. The wheat rIGPD contained one Mn atom per subunit, and the Mn was involved in the assembly of the subunits to form active IGPDs. Protein-blotting analysis, using antibodies raised against the wheat rIGPD, indicated that IGPD was located in the chloroplasts of wheat. The rIGPDs of Arabidopsis and wheat, which were 86% identical in their primary structures deduced from the cDNAs, exhibited similar properties in terms of the molecular mass, pH optimum, and the Km for the substrate, imidazoleglycerolphosphate. However, the nonselective herbicides 3-amino-1,2,4-triazole and a newly synthesized triazole [(1R*, 3R*)-[3-hydroxy-3-(2H-[1,2,4]triazole-3-yl)-cyclohexyl]- phosphonic acid], inhibited Arabidopsis and wheat IGPDs in a mixed-type and a competitive manner, respectively.

Amino Acid Sequence↗

PubMed1998

Characterization of cadmium binding, uptake, and translocation in intact seedlings of bread and durum wheat cultivars

High Cd content in durum wheat (Triticum turgidum L. var durum) grain grown in the United States and Canada presents potential health and economic problems for consumers and growers. In an effort to understand the biological processes that result in excess Cd accumulation, root Cd uptake and xylem translocation to shoots in seedlings of bread wheat (Triticum aestivum L.) and durum wheat cultivars were studied. Whole-plant Cd accumulation was somewhat greater in the bread wheat cultivar, but this was probably because of increased apoplastic Cd binding. Concentration-dependent 109Cd2+-influx kinetics in both cultivars were characterized by smooth, nonsaturating curves that could be dissected into linear and saturable components. The saturable component likely represented carrier-mediated Cd influx across root-cell plasma membranes (Michaelis constant, 20-40 nm; maximum initial velocity, 26-29 nmol g-1 fresh weight h-1), whereas linear Cd uptake represented cell wall binding of 109Cd. Cd translocation to shoots was greater in the bread wheat cultivar than in the durum cultivar because a larger proportion of root-absorbed Cd moved to shoots. Our results indicate that excess Cd accumulation in durum wheat grain is not correlated with seedling-root influx rates or root-to-shoot translocation, but may be related to phloem-mediated Cd transport to the grain.

Journal Article↗

PubMed1999

The localization and expression of the class II starch synthases of wheat.

The starch granules of hexaploid wheat (Triticum aestivum) contain a group of three proteins known as SGP-1 (starch granule protein-1) proteins, which have apparent molecular masses of 100, 108, and 115 kD. The nature and role of these proteins has not been defined previously. We demonstrate that these polypeptides are starch synthases that are present in both the starch granule and the soluble fraction at the early stages of wheat endosperm development, but that are exclusively granule bound at mid and late endosperm development. A partial cDNA clone encoding a fragment of the 100-kD protein was obtained by screening a wheat endosperm cDNA expression library using monoclonal antibodies. Three classes of cDNA were subsequently isolated from a wheat endosperm cDNA library by nucleic acid hybridization and were shown to encode the 100-, 108-, and 115-kD proteins. The cDNA sequences are highly homologous to class II starch synthases and have the highest homology with the maize SSIIa (starch synthase IIa) gene. mRNA for the SGP-1 proteins was detected in the leaf, pre-anthesis florets, and endosperm of wheat and is highly expressed in the leaf and in the grain during the early to mid stages of development. We discuss the roles of the SGP-1 proteins in starch biosynthesis in wheat.

Amino Acid Sequence↗

PubMed2001

Molecular evolution of receptor-like kinase genes in hexaploid wheat. Independent evolution of orthologs after polyploidization and mechanisms of local rearrangements at paralogous loci.

Hexaploid wheat is a young polyploid species and represents a good model to study mechanisms of gene evolution after polyploidization. Recent studies at the scale of the whole genome have suggested rapid genomic changes after polyploidization but so far the rearrangements that have occurred in terms of gene content and organization have not been analyzed at the microlevel in wheat. Here, we have isolated members of a receptor kinase (Lrk) gene family in hexaploid and diploid wheat, Aegilops tauschii, and barley (Hordeum vulgare). Phylogenetic analysis has allowed us to establish evolutionary relationships (orthology versus paralogy) between the different members of this gene family in wheat as well as with Lrk genes from barley. It also demonstrated that the sequences of the homoeologous Lrk genes evolved independently after polyploidization. In addition, we found evidence for gene loss during the evolution of wheat and barley. Analysis of large genomic fragments isolated from nonorthologous Lrk loci showed a high conservation of the gene content and gene organization at these loci on the homoeologous group 1 chromosomes of wheat and barley. Finally, sequence comparison of two paralogous fragments of chromosome 1B showed a large number of local events (sequence duplications, deletions, and insertions), which reveal rearrangements and mechanisms for genome enlargement at the microlevel.

Base Sequence↗

PubMed1966

Composition of guttation fluid from rye, wheat, and barley seedlings.

This work was undertaken to determine the kinds and amount of substances that would account for the previously demonstrated differential growth of Claviceps purpurea on guttation fluids from Rosen rye, Genesee wheat, and Traill barley seedlings. Chromatographic methods were used for determining amino acids and sugars, spot tests and spectrometric methods for inorganic materials, and microbiological methods for vitamins.Total sugar content is about equal in rye and barley fluids, but lower in wheat. Glucose is the principal sugar component of the rye and barley fluids and galactose highest in wheat. Most of the amino acid in all 3 fluids is aspartic acid or asparagine. Barley fluid is far higher than the other 2 in total amino acids, with wheat the lowest. Most inorganic elements are found to be highest in barley and lowest in wheat, with the exception of iron where rye is highest and barley lowest. Barley fluid is highest in choline, p-aminobenzoic acid, thiamine, and uracil, while rye is highest in inositol and pyridoxine. Wheat is much lower than the other 2 in choline and inositol.

Journal Article↗

PubMed1982

A comparison of the properties of ATPase associated with wheat and cauliflower plasma membranes.

Plasma membrane-associated ATPase obtained from cauliflower (Brassica oleraceae L.) florets isolated and assayed by several different procedures was stimulated 150 to 400% by K(+). In contrast, winter wheat (Triticum aestivum L. cv. Kharkov 22 MC) shoot and root ATPase obtained by the same methods exhibited only 10 to 25% stimulation by K(+). The level of K(+)-stimulation of the wheat enzyme was not significantly increased by purifying the crude microsomal membrane fraction using sucrose density gradients. ATPase associated with density gradient-purified cauliflower membranes was inhibited by Ca(2+), high ATP concentration in the presence of low Mg(2+), and by several metabolic inhibitors. In contrast, the wheat enzyme was largely unaffected by all of these treatments. The plasma membranes of intact wheat and cauliflower cells gave a positive reaction with the plasma membrane-specific, phosphotungstic acid-chromic acid stain (PACP). A high proportion of the cauliflower membrane vesicles in the putative plasma membrane-enriched fraction stained with PACP, whereas only a small proportion of the wheat membrane vesicles reacted positively with PACP. These results indicate that a plasma membrane-enriched fraction has been isolated successfully from cauliflower floret tissue, but that none of the procedures used effectively separate plasma membranes from homogenates of wheat shoots and roots.

Journal Article↗

PubMed1986

Temperature characteristics and adaptive potential of wheat ribosomes.

The translational efficiency of wheat ribosomes was studied as a function of an in vivo temperature pretreatment of wheat seedlings (Triticum aestivum L.). Ribosomes were isolated from heat-pretreated (36 degrees C) and reference (4 degrees C, 20 degrees C) wheat seedlings. The efficiency of the ribosomes in translating polyuridylic acid was assayed. Ribosomes from heat-pretreated seedlings exhibit a threefold enhanced incorporation rate of phenylalanine as compared to ribosomes from wheat seedlings adapted to 20 or 4 degrees C. This difference develops within 24 hours after onset of the heat treatment of seedlings following a 3 hour lag phase. The temperature induced changes can be traced back to the cytoplasmic ribosomes, since cycloheximide inhibits translation almost completely. Thermal inactivation of ribosomes occurs at 45 degrees C, irrespective of the temperature pretreatment of the wheat seedlings. Specific differences in the yield of ribosomes, in the polyribosomal profiles, and in the apparent Arrhenius' activation energy of protein synthesis were observed depending on the age and the temperature pretreatments. The results presented here are considered an important molecular correlation to phenotypical temperature adaptation of in vivo protein synthesis in wheat (M Weidner, C Mathée, FK Schmitz 1982 Plant Physiol 69: 1281-1288).

Journal Article↗

PubMed1990

Crossreactivity of IgE antibodies from sera of subjects allergic to both ryegrass pollen and wheat endosperm proteins: evidence for common allergenic determinants.

Positive RAST (greater than 5% radioactive uptakes) to wheat endosperm proteins were found in approximately one-quarter of subjects who had both a positive skin prick and RAST (greater than 10% radioactive uptake) to ryegrass pollen proteins. Immunoblotting of proteins electrophoretically transferred to nitrocellulose membrane after SDS-PAGE of ryegrass pollen and wheat endosperm proteins confirmed the crossreactive properties of the sera identified by RAST testing. Immunoadsorption of serum IgE onto nitrocellulose membrane, to which ryegrass pollen or wheat endosperm proteins had been adsorbed, removed IgE from crossreactive sera reactive to both ryegrass pollen and wheat endosperm proteins. Elution of the adsorbed IgE from the nitrocellulose membrane after immunoadsorption and probing blotted strips of both ryegrass pollen and wheat endosperm proteins supported the results obtained from the immunoadsorption experiments. This data provides evidence that the crossreactivity of IgE antibodies in sera reacting with both ryegrass pollen and wheat endosperm proteins involves common or related determinants and has implications for the clinical management of these allergic subjects.

Blotting, Western↗

PubMed2006

Airborne exposure to wheat allergens: measurement by human immunoglobulin G4 and rabbit immunoglobulin G immunoassays.

BACKGROUND: Exposure to airborne wheat allergens in the bakery trade is associated with a high risk of occupational allergy and asthma. Control and reduction of allergen exposure require relatively simple but reliable monitoring techniques. We developed new rabbit IgG-based enzyme immunoassays (EIA) for wheat allergens, which might be a convenient alternative for the thus far used human IgG4 inhibition assay. METHODS: The reactivity and specificity of rabbit antibodies were assessed by EIA and immunoblotting, and compared with those of IgE from wheat-sensitized bakers, and with the antibodies used in the IgG4 inhibition EIA. An IgG inhibition and a sandwich EIA were developed for analysis of airborne dust samples. RESULTS: Human IgG4 and rabbit IgG inhibition EIAs had comparable sensitivities, with limits of detection (LOD) between 18 and 88 ng/mL, while the sandwich EIA was much more sensitive (LOD<0.2 ng/mL). Human IgG4 and rabbit IgG reacted in immunoblotting with most of the IgE-binding wheat proteins, although with quantitative differences. All three assays showed a strong reaction with wheat proteins, and some cross-reactivity with rye and barley, but were further highly specific for cereal flour proteins. Concentrations measured with the three EIAs in 432 airborne dust samples were highly correlated (r>0.95) and their absolute values showed less than 10-20% differences. CONCLUSION: The rabbit IgG EIAs are valid substitutes for the human IgG4 inhibition EIA, with important practical advantages. The inhibition EIA is recommended for routine wheat allergen measurements. The sandwich EIA may be used to measure low allergen levels, as in short task-related exposure measurements or in subfractions of airborne dust samples.

Air Pollutants, Occupational↗