Characterization of Prolamins from Meadow Grasses: Homology with those of Wheat, Barley and Rye

Prolamin fractions were extracted from seeds of five speciesof temperate meadow grasses. SDS-PAGE showed the presence ofpolypeptidcs with M_rs of 20–30000 in Phieum pratense andDactylis glomerata, and between 30000 and 65 000 in Lolium perenneand in two species of Festuca (F. rubra and F. arundinacea).The fractions had broadly similar amino acid compositions withhigh glutamate+glutamine (29 to 35 mol %) and phenylalaninc(8 to 10 mol %), but there was some variation in the contentsof prolinc (10 to 23 mol %) and several other amino acids includinglysine (0.3 to 1.9 mol %). Automated Edman degradation of twogel filtration fractions from L perenne and one fraction eachfrom the other four species showed single major N.terminal aminoacid sequences. These were homologous with each other, and withthe -typc prolamins of wheat, barley and rye. Key words: Prolamins, meadow grasses, SDS-PAGE     

Characterization and genetic control of the prolamins of Haynaldia villosa: Relationship to cultivated species of the Triticeae (rye,wheat, and barley)

Haynaldia villosa is a wild grass of the tribe Triticeae, other members of which include the cultivated cereals barley, rye, and wheat. We have made an electrophoretic and chemical characterization of the major seed storage proteins (prolamins) of H. villosa and determined the chromosomal locations of the structural genes for some components using the available wheat/H. villosa chromosome addition lines. As in wheat, barley, and rye, groups of high molecular weight (polymeric), sulfur-poor (monomeric), and sulfur-rich (monomeric -type and polymeric) prolamins can be recognized. Most of the components are encoded by genes on chromosome 1 Ha, which is homologous with the chromosomes controlling many of the prolamins in wheat and rye and all of those in barley. In addition, H. villosa also contains -type sulfur-rich prolamins, previously detected only in wheat and its close relatives. These may be encoded by genes on chromosome 6Ha, which is homologous with the group 6 chromosomes that control the -type gliadins of wheat. Despite the proposed close relationship between Haynaldia and ryes, no evidence was found for the presence of proteins closely related to the M _r 75,000 -secalins which are characteristic of wild and cultivated species of Secale.     

Cereal prolamin evolution and homology revealed by sequence analysis

Prolamin mixtures were isolated from oats, rice, normal and high-lysine sorghum, two varieties of pearl millet, two strains of teosinte, and gamma grass and subjected to NH₂-terminal amino acid sequence determinations. In each case (except for rice, whose prolamins apparently have blocked or unavailable NH₂-terminal residues), primarily a single sequence was observed despite significant heterogeneity, suggesting that prolamin homology in each cereal arose through duplication and mutation of a single ancestral gene. Comparisons were then made to prolamin sequences previously determined for wheat, corn, barley, and rye. Within genera, different varieties or subspecies exhibited few differences, but more distantly related genera, subtribes, and tribes showed increasingly large differences. Within the subfamily Festucoideae, no homology was apparent between prolamins of oats and those of the subtribe Triticinae (including wheat, rye, and barley, for which prolamin homology was previously demonstrated). Within the subfamily Panicoideae, corn was shown to be closely related to teosinte but more distantly to Tripsacum. Sorghum was shown to have diverged less from corn than had millet. These comparisons demonstrate that prolamin sequence analyses can successfully predict and clarify evolutionary relationships of cereals.     

Electron-probe Microanalysis Studies of Silicon in the Epicarp Hairs of the Caryopses of Hordeum sativum Jess., Avena sativa L., Secale cereale L. and Triticum aestivum L.

Silicon deposits in the epicarp hairs of the caryopses of mature,field-grown specimens of Hordeum sativum, Avena sativa, Secalecereale and Triticum aestivum were investigated using electron-probemicroanalysis. In all four cereals, silicon was most concentratedat the extreme tips of the hairs. In barley, it was the onlyelement detected; in the other three cereals potassium and calciumwere located below the tip. In wheat, chlorine was also detected. The hair bases of the different cereals displayed variationin the elements detected. Silicon and polassium were presentin all four and calcium present in all except rye. The hairbases of wheat also contained chlorine; phosphorus and zincwere located in barley. The latter alone showed variation ofelements between hair bases. Scanning electron microscopy revealed heavy striations of thehair tips of barley and oats. In rye and wheat, the tips weresmooth but there were slight surface markings below the tip. The results are discussed in relation to the possible functionsand significance of the epicarp hairs and their silicification. Avena sativa L., Hordeum sativum Jess., Secale cereale L., Triticum aestivum L., oat, barley, rye, wheat, epicarp hairs, silicon, electron-probe microanalysis, scanning electron microscopy     

Immunochemical studies on barley seed storage proteins

The antigenic relationships between the prolamins of barley, rye and wheat have been studied by examining the specificity of an antibody to C hordein in a quantitative study using a laser nephelometer. The antibody reacts weakly with B hordein and strongly with 75-kdalton and 40-kdalton -secalins from rye and ₃ some -gliadins from wheat. Absorption experiments and immunodiffusion tests indicate that there are shared antigenic determinants for most of the prolamins. All the species with reacting prolamins belong to the sub-family Festucoideae of the Gramineae. The prolamins of maize, pearl millet and sorghum, species of the sub-family Panicoideae, do not react. The results confirm the known lack of homology between the prolamins of the two sub-families and also indicate the presence of relationships not yet established between C hordein, the 75-kdalton and 40-kdalton -secalins and also ₃ gliadin.Abbreviations HMW high molecular weight - PAGE polyacnylamide-gel electrophoresis - PBS phosphate-buffered saline - RLS relative light scattering - SDS sodlum dodecyl sulphate     

Pathogenicity of Fusarium graminearum Schwabe isolates from Poland towards seedlings of cereal species

Ten isolates ofFusarium graminearum Schwabe originating from diseased cereal plants and kernels were tested for pathogenicity to various cultivars of wheat, rye, triticale and oats. The isolates varied greatly in their pathogenicity to the seedlings of the species, and were most pathogenic to rye and triticale, less pathogenic to barley and wheat and least pathogenic to oats.     

Infrared Spectroscopic Study of the Secondary Structure of Wheat,Rye, and Barley Prolamins

The secondary structure of prolamins, the storage proteins of wheat (Triticum aestivum L.), rye (Secale cereale L.), and barley (Hordeum vulgare L.), was studied by infrared (IR) spectroscopy. The secondary structure of wheat gliadin was characterized by a high content of unordered conformations of polypeptide chains (64–67%) and a low content of short -helical regions (4–5%). Elongated conformations, formed by poly-L-proline II helices (61 and 53%) and -turns (15 and 19%), predominated in rye secalins and barley hordeins. Prolamin proteins of cereal crops also contained -sheet structure regions comprising 9–12% of the polypeptide chains.     

Molecular evolution of the seed storage proteins of barley, rye and wheat

The major storage proteins (prolamins) of barley, rye and wheat are characterized by the presence of two or more unrelated structural domains, one of which contains repeated sequences. Because of this repetitive structure and their restricted distribution (only in grasses), it has been suggested that the prolamins are of recent origin. Contrary to this hypothesis, we show that parts of the non-repetitive domain of one group of prolamins are homologous with sequences present in a large group of seed proteins from monocotyledonous and dicotyledonous plants; including Bowman-Birk protease inhibitors, cereal inhibitors of alpha-amylase and trypsin, and 2 S globulin storage proteins of castor bean and oil seed rape. This implies an ancient origin for these non-repetitive domains. The origins of the repetitive domains are not known but may lie within the grasses.     

Cytoplasmic DNA variation and relationships in cereal genomes

Summary Chloroplast (cp) and mitochondrial (mt) DNAs were isolated from four cereal genomes (cultivated wheat, rye, barley and oats) and compared by restriction nuclease analysis. Cleavage of cp and mt DNAs by Sal I, Kpn I, Xho I and EcoR I enzymes indicated that each cereal group contains specific cytoplasmic DNAs. A phylogenetic tree of cereal evolution has been obtained on the basis of cp DNA homologies. It is suggested that wheat and rye diverged after their common ancestor had diverged from the ancestor of barley. This was preceded by the divergence of the common ancestor of wheat, rye and barley and the ancestor of oats.The molecular weight of the different cp DNAs was determined from the Sal I and Kpn I patterns. cp DNAs from wheat, rye, barley and oats appeared to be characterized by a very similar molecular weight of about 80–82.10⁶ d.In the case of the mt DNAs, the great number of restriction fragments obtained with the restriction enzymes used prevented precise comparisons and determination of molecular weights.     

Chromosomal location of genes controlling seed proteins in species related to wheat

Summary The seed proteins of Chinese Spring wheat stocks which possess single chromosomes from other plant species related to wheat have been separated by gel electrophoresis in the presence of sodium dodecyl sulphate. Marker protein bands have been detected for both arms of barley chromosome 5, chromosome E (= 1R) and B (= 2R) of rye, chromosomes A,B (= 1C^u) and C (= 5C^u) of Aegilops umbellulata and chromosomes I and III of Agropyron elongatum. These studies, and previous findings, indicate that chromosome 5 of barley, chromosome 1R of rye, chromosome I of Ag. elongatum and possibly chromosome 1C^u of Ae. umbellulata are similar to chromosomes 1A, 1B and 1D in hexaploid wheat in that they carry genes controlling prolamins on their short arms and genes controlling high-molecular-weight (apparent molecular weight greater than 86,000) seed protein species on their long arms. These findings support the idea that all these chromosomes are derived from a common ancestral chromosome and that they have maintained their integrity since their derivation from that ancestral chromosome.     

Lacewings (Chrysopidae,Neuroptera) in cereal agrocenoses of the forest-steppe of Western Siberia

Twelve species of lacewings: Chrysopa altaica, Ch. commata, Ch. perplexa, Ch. phyllochroma, Ch. dasyptera, Ch. carnea, Ch. formosa, Ch. intima, Ch. perla, Ch. prasina, Ch. septempunctata, and Nineta inpunctata were found in cereal agroecosystems of the forest-steppe zone of Western Siberia. The dominant species were Ch. carnea and Ch. phyllochroma. The biological characteristics, seasonal dynamics of the abundance of lacewings in the agrocenoses of winter rye, spring wheat, and oats are given. The abundance of larval and adult lacewings in the spring wheat agrocenosis was not affected by the level of chemicalization (upon condition of the rational use of insecticides), tillage variant, and predecessor crop. In the years when the density of lacewings was low, no differences in their population were found between the agrocenoses of wheat (after fallow), winter rye, oats, vetch-oats, canola, barley, and barley with melilot. In the years characterized by relatively high abundance of lacewings, they occurred more frequently in the crop rotations with wheat after fallow and with oats. These plants were settled by cereal aphids to the greatest extent. In all the years studied, the density of lacewings on alfalfa was 2–2.8 times as great as that on wheat after fallow.     

The Enzymic Decarboxylation of {gamma}-methyleneglutamic Acid by Plant Extracts

-Methyleneglutamic acid, an acidic amino-acid isolated fromgroundnut plants, was decarboxylated by enzymes present in extractsof Capsicum fruits, barley roots, and tulip leaves, and alsoby intact cells of Clostridium welchii S.R. I2. The amino-acidwas attacked in a similar manner to, but in all cases at a slowerrate than, l-glutamic acid. The nature of the enzyme responsiblefor the decarboxylation of -methyleneglutamic acid was furtherinvestigated using preparations from barley roots (which donot contain the amino-acid) and from tulip leaves (in whichthe amino-acid is normally present, together with larger amountsof its amide form, -methyleneglutamine). The effects of pH,inhibitors, and partial heat denaturation upon the enzyme systemspresent in the barley and tulip extracts indicated that a singleenzyme was responsible for the decarboxylation of both l-glutamicacid and -methyleneglutamic acid. Although the Cl. welchii rapidlydeamidated and then decarboxylated l-glutamine, -methyleneglutaminewas not attacked by the organism.     

Specificity of an antibody to a subunit of high-molecular-weight storage protein from wheat seed and its reaction with other cereal storage proteins (prolamins)

An antiserum to subunit 2 from the high-molecular-weight (HMW) subunits of the glutenin fraction of Triticum aestivum cv. Highbury was shown to react with related subunits from other cultivars of wheat. The reaction was measured quantitatively by laser nephelometry in polyethylene glycol phosphate-buffered saline after dissolving the HMW fraction in 0.1 M acetic acid; urea used to dissolve the HMW prolamins inhibited the reaction, in some cases at the low concentration of 0.06 M. A study of the comparative reactions of other cereal prolamins was made. D hordein, the homologous HMW protein of barley, showed less reaction, which was more inhibited by urea than the wheat subunits. Some -gliadins from the wheat cultivars Chinese Spring and Cheyenne reacted more strongly than the injected fraction and there was less inhibition by urea. A-, - and ₃ of wheat also reacted with the antiserum while a secalin of rye of M_r 40000 gave a weak reaction.Abbreviations HMW high molecular weight - PAGE polyacrylamide-gel electrophoresis - PBS phosphate-buffered saline - PE pyridylethylated - SDS sodium dodecyl sulphate     

SOME OBSERVATIONS ON THE CEREAL-ROOT EELWORM POPULATION OF FIELD PLOTS OF CEREALS WITH DIFFERENT SOWING TIMES AND FERTILIZER TREATMENTS

In the autumn of 1953 an experiment was begun to follow changes in the cereal-root eelworm population of small plots on a field in Shropshire. The plots were cropped with either oats, wheat, barley or rye, sown in the autumn and spring, and some plots had fertilizer. Each plot received the same treatment for 3 years; in the fourth year an indicator crop of spring oats was grown on all the plots.
Under rye and autumn-sown wheat the eelworm population fell to a level which permitted a good oat crop in 1957. Autumn-sown wheat, barley and rye generally produced lower eelworm populations than their spring-sown counterparts, but autumn-sown oats proved to be the most efficient host. The order of host efficiency was oats (best), barley, wheat, rye. The eelworm populations were generally higher on plots receiving fertilizer treatment. In this experiment all oat plots, and spring-sown barley plus fertilizer, produced populations which severely damaged the 1957 oat crop.     

Comparisons of Peptide hydrolase activities in cereals

Carboxypeptidase activity (hydrolysis of N-carbobenzoxy-l-phenylalanyl-l-alanine) is high in a number of temperate zone cereals, originating in Asia Minor (wheat, barley, oats, wild oats, rye, triticale) compared to other cereals originating in central America or Asia (maize, sorghum, rice). However, endopeptidase activity (hydrolysis of azocasein or hemoglobin) is relatively much higher in the latter group. Comparison of trichloroacetic acid (TCA)-soluble products derived from the hydrolysis of hemoglobin showed that carboxyterminal amino acids (histidine, arginine, and tyrosine), are released when extracts from wheat and barley endosperms are used. With extracts from corn endosperms, much more TCA-soluble ultraviolet- absorbing material is released, but very little is released as free amino acids within the first 2 hours and the expected C-terminal amino acids of hemoglobin are not detected in significant amounts. These results suggest that the method of hydrolysis of the storage proteins may be significantly different in these two classes of cereals.     

Toward the assessment of food toxicity for celiac patients: characterization of monoclonal antibodies to a main immunogenic gluten peptide

Background and Aims

Celiac disease is a permanent intolerance to gluten prolamins from wheat, barley, rye and, in some patients, oats. Partially digested gluten peptides produced in the digestive tract cause inflammation of the small intestine. High throughput, immune-based assays using monoclonal antibodies specific for these immunotoxic peptides would facilitate their detection in food and enable monitoring of their enzymatic detoxification. Two monoclonal antibodies, G12 and A1, were developed against a highly immunotoxic 33-mer peptide. The potential of each antibody for quantifying food toxicity for celiac patients was studied.

Methods

Epitope preferences of G12 and A1 antibodies were determined by ELISA with gluten-derived peptide variants of recombinant, synthetic or enzymatic origin.

Results

The recognition sequences of G12 and A1 antibodies were hexameric and heptameric epitopes, respectively. Although G12 affinity for the 33-mer was superior to A1, the sensitivity for gluten detection was higher for A1. This observation correlated to the higher number of A1 epitopes found in prolamins than G12 epitopes. Activation of T cell from gluten digested by glutenases decreased equivalently to the detection of intact peptides by A1 antibody. Peptide recognition of A1 included gliadin peptides involved in the both the adaptive and innate immunological response in celiac disease.

Conclusions

The sensitivity and epitope preferences of the A1 antibody resulted to be useful to detect gluten relevant peptides to infer the potential toxicity of food for celiac patients as well as to monitor peptide modifications by transglutaminase 2 or glutenases.     

The invasion of root tips of cereals by the cereal cyst nematode Heterodera avenue

Newly germinated seedlings of susceptible cultivars of oats, wheat, barley and rye were inoculated with second-stage juveniles of Heterodera avenae in pots of sand. Subsequent examination showed oat root tips to be more commonly invaded, and by a greater range of nematode numbers than the other cereals. A comparison of oats and barley showed that lower nematode numbers in barley were not due to a higher emigration from barley; invasion, establishment and emigration by nematodes all being greater in oats. Second-stage juveniles were more likely to migrate prior to establishment in barley than in oats.     

Identification of Homologous Globulins from Embryos of Wheat, Barley, Rye and Oats

Total globulins from embryos and endosperms of barley, wheat,rye, and oats were separated by SDS-PAGE under reducing andnon-reducing conditions. The preparations from embryos of allfour cereals contained major groups of bands with M_r's of 50-60000,which were not affected by reduction. These have been characterizedpreviously from oats and shown to correspond to subunits ofthe 7S storage globulin. Immunochemical relationships betweenthese bands (and others with M_r's between 40000 and 70000) weredemonstrated by immunodiffusion and ‘Western Blotting’using antiserum raised against the major subunits of the oat7S globulins. The 7S globulins were also prepared from hand-dissectedembryos of the four cereals using sucrose density ultracentrifugation.Their amino acid compositions were broadly similar, but differedfrom those of the 7S vicilins of legumes. It is concluded thatstructurally-related 7S globulins are present in the embryosof the four species of cereals. Key words: Homologous globulins, embryo, wheat, barley, rye, oats     

Biology,host preferences and fitness of <Emphasis Type="Italic">Oulema melanopus</Emphasis> (Coleoptera: Chrysomelidae), a recent invasive pest in Western Canada

The developmental parameters and fitness of a recent invasive insect pest of cereals in western Canada, the cereal leaf beetle, Oulema melanopus (L.) (Coleoptera: Chrysomelidae), were compared on live plants and excised leaves of commercial cultivars of potential cereal hosts including wheat (winter and spring), oats, barley, corn, rye and triticale. Host preference and utilization within the fundamental host range of O. melanopus differed. The biological parameters differed significantly when the larvae were reared on excised tissues versus on live host plants. In both studies, wheat (winter and spring), oat (cv. Morgan) and barley were the most preferred hosts in terms of development, survivorship, adult weights and fecundity. Prolonged development with low fitness gains was noted on corn. Although the development on hosts such as rye and triticale was prolonged and adult fitness was low, the survivorship was high on these hosts. Hence, these crops can act as secondary hosts for the beetle in its new eco-region. Larvae fed foliage of Waldern, a local oat cultivar, had increased developmental time and lower survivorship compared with other cereal hosts. Despite differences in fitness gains among hosts, similar numbers of eggs were laid on all hosts. Hence, the oviposition choice of O. melanopus may not be driven by fitness gains alone and it may indicate adaptive strategy to hosts in newer environments.