Study of the effects produced by gamma-irradiation of common wheat F1 seeds using gliadins as genetic markers

A study of the effects produced by gamma-irradiation of dry F₁ seeds of common wheat at 200 Gy is described in the work. The experimental material was hybrids produced from near-isogenic lines based on Bezostaya 1. Irradiation led to a significant decrease in the productivity traits of F₁ plants but did not affect the survival of plants under the given growth conditions. It has been found that one of the effects of the F₁ seed irradiation was a relative, compared to the control, increase in the frequency of male gametes with the 1BL/1RS translocation that participated in the production of F₂ grains. The irradiation induced mutations at gliadin loci with a frequency of 7.4% (against 0.5% in the control).     

Polymorphism and inheritance of gliadin polypeptides in T. monococcum L.

Summary More than 80 different gliadin electrophoretic patterns (spectra) have been found in 109 accessions of the diploid wheat Triticum monococcum. Each pattern consists of 15–20 gliadin bands. Some patterns are clearly related and might arise from one another through single mutations in the gliadin-coding loci. From the analysis of 15 grains of each, only 61 accessions were found to be uniform; others consisted of two or more grain variants differing in their gliadin spectrum. An analysis of F₂ grains from three crosses between different accessions showed that groups (blocks) of components are jointly and codominantly inherited. Two independent major Gli loci were established. The close resemblance of the composition of some blocks of T. monococcum to some of those in polyploid wheats indicates that one locus in each T. monococcum genotype is located on chromosome 1A (Gli-A1) and the other on 6A (Gli-A2). However, the blocks of T. monococcum include more bands than corresponding (equivalent) blocks of polyploid wheats. Two out of 275 F₂ grains of the cross k-14244 x k-20409 were found to have gliadin spectra which can be explained as a result of intralocus recombination. Also, a second gliadin-coding locus on chromosome 1A was found in the cross k-46140 x k-46753. This locus recombines with the main Gli-A1 locus with a frequency of about 22% and was clearly analogous to the additional Gli locus found earlier on chromosome 1A of certain polyploid wheats.     

Analysis of common wheat varieties and near-isogenic lines by PCR with allele-specific primers for Gli-1 and Glu-3 loci

The allelic characteristics of the Gli-A1, Gli-B1, Gli-D1 and Glu-A3 loci of 14 bread wheat varieties and 6 near-isogenic lines derived from Bezostaya 1 have been detected by PCR analysis. The conformity of molecular-genetic data and electrophoresis of storage proteins has been determined: the allelic variants of gliadins Gli-A1o and Gi-A1m correspond to the PCR-allele GliA1.2, the gliadin variants Gli-A1f, Gli-A1b, Gli-A1c correspond to the PCR-allele GliA 1.1, the allelic variants Gli-B1b, Gli-B1d--to the PCR-allele GliB1.1 and the variants Gli-B1e, Gli-B1g, Gli-B1c-to the PCR-allele GliB1.2. A new PCR-allele at the GliB) locus in the line Gli-B1-12 (with the gliadin block Gli-B1o from Levent) was identified.     

Genetic control of gliadin components in wheat <Emphasis Type="Italic">Triticum spelta</Emphasis> L.

The componental composition of electrophoretic spectra of gliadin in Triticum spelta L. was studied. By analogy with common wheat T. aestivum L., it was established that genes controlling gliadin components in spelt are also located in short arms of chromosomes of homeological groups 1 and 6. Analysis of gliadin spectra in F₂ grains from the crosses k-20539 × Ershovskaya 32 and k-20558 × Ershovskaya 32 revealed linkage of some components and their grouping into blocks (alleles) of coinherited gliadin components. Alleles of gliadin-coding loci identical to alleles of common wheat and new alleles earlier unknown for wheat populations have been identified.     

Gliadin alleles in Canadian western red spring wheat cultivars: use of two different procedures of acid polyacrylamide gel electrophoresis for gliadin separation.

Gliadin allele compositions of 21 Canadian spring common wheat cultivars, most of which belong to the Canada western red spring (CWRS) class, were studied and great similarity in their genotypes was confirmed. It was found that alleles frequent in the set of Canadian wheats (such as Gli-B1d, Gli-D1j, Gli-A2m, and Gli-D2h) are very rare or absent in common wheat cultivars from other regions and countries studied earlier, indicating that germplasm of CWRS cultivars is rather unique. It may be suggested that alleles frequent in Canadian cultivars relate to important technological characteristics of these wheats and may possibly serve as marker genes during selection for quality traits. Similarity of gliadin electrophoregrams obtained by two different acid polyacryl-amide gel electrophoretic procedures for the same genotype was established, and the component composition of allelic variants of blocks of gliadin components found in the set of Canadian cultivars and in standard cultivars Chinese Spring and Bezostaya 1 are described.     

Possibilities of insertion of glutenin subunits and gliadin components of glu B1, glu B3 and gli B1 loci fromTriticum turgidum intoTriticum aestivum

An electrophoretic investigation of the glutenin subunits and gliadin components in F₄ progenies between the hexaploid wheat (Triticum aestivum L. cv. Bezostaya 1) and the wild tetraploid wheat (Triticum turgidum var.dicoccoides Körn) is carried out. The possibility of inserting subunits and components of Glu B1, Glu B3 and Gli B1 loci fromT. dicoccoides in F₄ progenies ofT. aestivum is investigated. For this purpose parental forms with different allelic variants in these loci are chosen. It is established cytologically that the chromosomal number of progenies is 42. Genotype classes and the frequency of progenies are analyzed for which we judge by the phenotype manifestation of the allelic variants of the loci investigated. Different frequency of the transfer of subunits and components of Glu B1, Glu B3 and Gli B1 loci fromT. dicoccoides in the hexaploid wheat genome is established and it is connected with the disposition of the loci on chromosomes arms.     

Recombination mapping of some chromosome 1A-, 1B-, 1D- and 6B-controlled gliadins and low-molecular-weight glutenin subunits in common wheat

 Inheritance of low-molecular-weight glutenin subunits (LMW GS) and gliadins was studied in the segregating progeny from several crosses between common wheat genotypes. The occurrence of a few recombinants in the F₂ grains of the cross Skorospelka Uluchshennaya×Kharkovskaya 6 could be accounted for by assuming that the short arm of chromosome 1D contains two tightly linked loci each coding for at least one gliadin plus one C-type LMW GS. These loci were found to recombine at a frequency of about 2%, and to be linked to the Glu-D3 locus coding for B-type LMW GS. Some proteins showing biochemical characteristics of D-type or C-type LMW GS were found to be encoded by the Gli-B1 and Gli-B2 loci, respectively. Strongly stained B-type LMW GS in cvs Skorospelka Uluchshennaya and Richelle were assigned to the Glu-B3 locus, but recombination between this locus and Gli-B1 was not found. Analogously, in the cross Bezostaya 1×Anda, no recombination was found between Gli-A1 and Glu-A3, suggesting the maximum genetic distance between these loci to be 0.97% (P=0.05). A B-type LMW GS in cv Kharkovskaya 6 was assigned to the Glu-B2 locus, with about 25% recombination from the Gli-B1 locus. The present results suggested that alleles at Gli loci may relate to dough quality and serve as genetic markers of certain LMW GS affecting breadmaking quality. Received: 9 July 1996/Accepted: 15 November 1996     

A novel globular protein electrospun fiber mat with the addition of polysilsesquioxane

The aim of this work has been to elaborate well defined gliadin nanofibers with incorporation of inorganic molecules, such as polyhedral oligomeric silsesquioxane (POSS). Nanofibers were obtained by electrospinning processing, controlling the relevant parameters such as tip-to-collector distance, voltage and feed rate. The fiber mats were characterized by SEM, confocal images, DSC, viscosity, FTIR and conductivimetry analysis. FTIR spectra showed characteristic absorption bands related to the presence of POSS-NH₂ within the matrices. SEM micrographs showed that gliadin fibers decreased their dimensions as the amount of POSS-NH₂ increased in the spinning solution. The electrical conductivity of gliadin solutions diminished as the concentration of POSS-NH₂ was increased. Besides, confocal micrographs revealed that POSS-NH₂ might be dispersed as nanocrystals into gliadin and gluten fibers. The dimension of gluten nanofibers was also affected by the POSS-NH₂ concentration, but conversely, this dependence was not proportional to the POSS-NH₂ amount. Somehow, the interaction between gliadin and POSS-NH₂ in aqueous TFE affected the solution viscosity and, as a consequence, higher jet instabilities and thinner fiber dimensions were obtained.     

Comparative analysis of the genetic diversity dynamics at gliadin loci in the winter common wheat Triticum aestivum L. cultivars developed in Serbia and Italy over 40 years of scientific breeding

The dynamics of genetic transformations at gliadin-coding loci in the winter common wheat cultivars produced in Serbia and Italy over 40 years of scientific breeding was studied. It was demonstrated that a number of alleles unique for the wheat cultivars of each country were substituted with the alleles of a limited number of donor cultivars, in particular, cultivar Bezostaya 1 and its derivatives. On the background of preserved heterogeneity values during various time periods of breeding in each country, the genetic diversity in the total region decreased, as demonstrated by similarity in the sets of alleles of gliadin loci and their frequencies in the modem cultivars of these two Southern European countries. This decrease in the genetic diversity is an erosion of genetic resources within the region, which results in a loss of unique coadapted gene complexes.     

基于醇溶蛋白的20份小麦种质遗传完整性分析

采用醇溶蛋白电泳技术对同一品种不同繁殖年份的20份小麦种质进行遗传完整性分析。结果表明：供试种质中有10份具有一种醇溶蛋白谱带带型的同质性种质;另外10份具有2～4种醇溶蛋白谱带带型的异质性种质,其中6份为地方品种。表明地方品种具有较高的遗传多样性。在10份异质性种质中,两个繁殖年份种质之间的醇溶蛋白带型频率变化差异不显著的有5份,其第一繁殖年份的种质发芽率均高于75％,而另外5份存在显著差异的种质,第一年份的发芽率都低于66％。进一步分析表明,这10份异质性种质在两个繁殖年份之间,其发芽率差值与带型频率差值之间呈极显著正相关,相关系数为0．8665。上速结果表明,小麦更新时较高的发芽率是维持异质性种质遗传完整性的关键因素。     

Comparative analysis of the genetic diversity dynamics at gliadin loci in the winter common wheat Triticum aestivum L. cultivars developed in Serbia and Italy over 40 years of scientific breeding

The dynamics of genetic transformations at gliadin-coding loci in the winter common wheat cultivars produced in Serbia and Italy over 40 years of scientific breeding was studied. It was demonstrated that a number of alleles unique for the wheat cultivars of each country were substituted with the alleles of a limited number of donor cultivars, in particular, cultivar Bezostaya 1 and its derivatives. On the background of preserved heterogeneity values during various time periods of breeding in each country, the genetic diversity in the total region decreased, as demonstrated by similarity in the sets of alleles of gliadin loci and their frequencies in the modern cultivars of these two Southern European countries. This decrease in the genetic diversity is an erosion of genetic resources within the region, which results in a loss of unique coadapted gene complexes.     

Does Solid-state <Superscript>15</Superscript>N NMR Spectroscopy Detect all Soil Organic Nitrogen?

Virtually all of the N detected by ¹⁵N cross polarization (CP) NMR spectra of four HF-treated soil clay fractions is amide N. However, the intensity of this ¹⁵N CP NMR signal (per unit N) is 27–57% lower than detected for a wheat protein, gliadin. There are two possible explanations – either the amide N in the soil clay fractions produces proportionately less NMR signal than does the amide N in gliadin, or part of the N in the soil clay fractions produces little or no NMR signal. The cross polarization dynamics of the gliadin amide resonance and amide resonances detected for the soil clay fractions are very similar and thus should produce similar amounts of signal, ruling out the first possibility. Therefore up to half or even more of the organic N in these soil clay fractions must be in a form that is insensitive to NMR detection. For a model compound (caffeine), non-protonated heterocyclic N produced less than 20% of the signal of an equivalent amount of amide N in gliadin. Results from several ¹³C NMR techniques provide further evidence that much of the undetected N in the soil clay fractions may be heterocyclic.     

Gliadin Fragments and a Specific Gliadin 33-mer Peptide Close KATP Channels and Induce Insulin Secretion in INS-1E Cells and Rat Islets of Langerhans

In non-obese diabetic (NOD) mice, diabetes incidence is reduced by a gluten-free diet. Gluten peptides, such as the compound gliadin, can cross the intestinal barrier and may directly affect pancreatic beta cells. We investigated the effects of enzymatically-digested gliadin in NOD mice, INS-1E cells and rat islets. Six injections of gliadin digest in 6-week-old NOD mice did not affect diabetes development, but increased weight gain (20% increase by day 100). In INS-1E cells, incubation with gliadin digest induced a dose-dependent increase in insulin secretion, up to 2.5-fold after 24 hours. A similar effect was observed in isolated rat islets (1.6-fold increase). In INS-1E cells, diazoxide reduced the stimulatory effect of gliadin digest. Additionally, gliadin digest was shown to decrease current through K_ATP-channels. A specific gliadin 33-mer had a similar effect, both on current and insulin secretion. Finally, INS-1E incubation with gliadin digest potentiated palmitate-induced insulin secretion by 13% compared to controls. Our data suggest that gliadin fragments may contribute to the beta-cell hyperactivity observed prior to the development of type 1 diabetes.     

Resolution of Trp near UV CD spectra of calmodulin-domain peptide complexes into the 1La and 1Lb component spectra

Near uv CD spectra of Trp residues in proteins frequently show a complex line shape deriving from the overlap of ¹L_a and ¹L_b electronic transitions. This study presents an original empirical method of resolving these components, based on the near uv CD spectra of well-defined complexes of calmodulin domains with target peptides containing a single Trp residue and derived from the skeletal muscle myosin light chain kinase target sequence. Spectra of 4 complexes were used to obtain the ¹L_a and ¹L_b component spectra that were then used to analyze further complexes. The broad and featureless ¹L_a spectrum is centered at 279 nm, the ¹L_b spectrum shows vibrational fine structure with maxima at 274.9, 281.5, and 289.8 nm. The CD spectrum of most complexes could successfully be fitted with one ¹L_a and one ¹L_b spectrum, the ¹L_b spectrum being negative for all complexes but the ¹L_a spectrum showing either positive or negative sign. Spectra of some complexes, however, failed to be adequately represented by only one ¹L_a and one ¹L_b spectrum. Instead, they could be fitted with one ¹L_b spectrum and two ¹L_a spectra with different sign and position. The method is successful in identifying and quantitating the relative intensities of a two-component system, consistent with a single conformation for tryptophan in a protein, and provides a simple indication of cases where a more complicated explanation is required. © 1998 John Wiley & Sons, Inc. Biopoly 45: 493–501, 1998     

Protein markers of plant traits in breeding spring wheat for grain yield and quality

A comparative study of the composition of gliadin proteins in grains of different-quality wheat cultivars Rollo and Drott and four forms of the hybrids F₉ and F₁₀ obtained by crossing these cultivars has been performed. The grain yield of ears, quality of flour, dough, and gluten were compared. Groups of genetically linked gliadin components, controlled by chromosomes 1 and 6 of homologous groups, have been identified as protein markers of selectively valuable plant traits.Translated from Prikladnaya Biokhimiya i Mikrobiologiya, Vol. 41, No. 1, 2005, pp. 121–126.Original Russian Text Copyright © 2005 by Berezovskaya, Trufanov, Mitrofanova, Kazmiruk.     

Gliadin fragments promote migration of dendritic cells

In genetically predisposed individuals, ingestion of wheat gliadin provokes a T‐cell‐mediated enteropathy, celiac disease. Gliadin fragments were previously reported to induce phenotypic maturation and Th1 cytokine production by human dendritic cells (DCs) and to boost their capacity to stimulate allogeneic T cells. Here, we monitor the effects of gliadin on migratory capacities of DCs. Using transwell assays, we show that gliadin peptic digest stimulates migration of human DCs and their chemotactic responsiveness to the lymph node‐homing chemokines CCL19 and CCL21. The gliadin‐induced migration is accompanied by extensive alterations of the cytoskeletal organization, with dissolution of adhesion structures, podosomes, as well as up‐regulation of the CC chemokine receptor (CCR) 7 on cell surface and induction of cyclooxygenase (COX)‐2 enzyme that mediates prostaglandin E2 (PGE₂) production. Blocking experiments confirmed that gliadin‐induced migration is independent of the TLR4 signalling. Moreover, we showed that the α‐gliadin‐derived 31–43 peptide is an active migration‐inducing component of the digest. The migration promoted by gliadin fragments or the 31–43 peptide required activation of p38 mitogen‐activated protein kinase (MAPK). As revealed using p38 MAPK inhibitor SB203580, this was responsible for DC cytoskeletal transition, CCR7 up‐regulation and PGE₂ production in particular. Taken together, this study provides a new insight into pathogenic features of gliadin fragments by demonstrating their ability to promote DC migration, which is a prerequisite for efficient priming of naive T cells, contributing to celiac disease pathology.     

Facile measurement of <Superscript>1</Superscript>H–<Superscript>15</Superscript>N residual dipolar couplings in larger perdeuterated proteins

We present a simple method, ARTSY, for extracting ¹J_NH couplings and ¹H–¹⁵N RDCs from an interleaved set of two-dimensional ¹H–¹⁵N TROSY-HSQC spectra, based on the principle of quantitative J correlation. The primary advantage of the ARTSY method over other methods is the ability to measure couplings without scaling peak positions or altering the narrow line widths characteristic of TROSY spectra. Accuracy of the method is demonstrated for the model system GB3. Application to the catalytic core domain of HIV integrase, a 36 kDa homodimer with unfavorable spectral characteristics, demonstrates its practical utility. Precision of the RDC measurement is limited by the signal-to-noise ratio, S/N, achievable in the 2D TROSY-HSQC spectrum, and is approximately given by 30/(S/N) Hz.     

Genetic aspects of wheat gliadin proteins

Inheritance of gliadin components unique to three different varieties of common wheat (Triticum aestivum L.) was studied in F₁ and F₂ seeds of intervarietal crosses using protein patterns obtained by polyacrylamide gel electrophoresis in aluminum lactate buffer (pH 3.2). The patterns of F₁ seeds of the crosses Cheyenne × Justin and INIA 66R × Justin evidenced all the bands present in the patterns of the parents; band intensities reflected gene dosage levels dependent on whether the contributing parent was maternal or paternal in accordance with the triploid nature of endosperm tissue. Most of the gliadin components examined segregated in accordance with control by a single dominant gene, but in two instances single bands in the one-dimensional electrophoretic patterns segregated in the F₂ as expected if controlled by two genes. A method of two-dimensional electrophoresis was developed that resolved these apparently single bands into two components each, which could segregate independently. Linkage analysis provided evidence of codominant alleles and closely linked genes coding for gliadin protein components in both coupling and repulsion situations. The gliadin protein components seem to be coded for by clusters of genes located on chromosomes of homoeologous groups 1 and 6 in hexaploid wheats.Reference to a company or product name does not imply approval by the U.S. Department of Agriculture to the exclusion of others which may also be suitable.     

On the Use of Alleles of Gliadin-Coding Loci as Possible Adaptability Markers in the Spring Wheat (Triticum aestivum L.) Cultivars during Seed Germination

Prolamine proteolysis is assumed to be among numerous adaptability factors in cereals. The patterns of gliadin proteolysis have been studied in 16 cultivars of spring wheat via analysis of electrophoretic spectra. Four proteolytic patterns have been identified. It is hypothesized that the cultivars characterized by early and rapid proteolysis (the first and third types) are the most adaptable. The gliadin genetic formulas of chromosomes of the first homeologous group have been determined. The alleles of gliadin loci (Gli-A1f, Gli-B1e, Gli-D1a, and Gli-D1b) have been found that can be used as markers of adaptability in spring wheat cultivars.     

Absorbance difference spectra of the successive redox states of the oxygen-evolving apparatus of photosynthesis

The spectra of the absorbance changes due to the turnover of the so-called S-states of the oxygen-evolving apparatus were determined. The changes were induced by a series of saturating flashes in dark-adapted Photosystem II preparations, isolated from spinach chloroplasts. The electron acceptor was 2,5-dichloro-p-benzoquinone. The fraction of System II centers involved in each S-state transition on each flash was calculated from the oscillation pattern of the 1 ms absorbance transient which accompanies oxygen release. The difference spectrum associated with each S-state transition was then calculated from the observed flash-induced difference spectra. The spectra were found to contain a contribution by electron transfer at the acceptor side, which oscillated during the flash series approximately with a periodicity of two and was apparently modulated to some extent by the redox state of the donor side. At the donor side, the S₀ → S₁, S₁ → S₂ and S₂ → S₃ transitions were all three accompanied by the same absorbance difference spectrum, attributed previously to an oxidation of Mn(III) to Mn(IV) (Dekker, J.P., Van Gorkom, H.J., Brok, M. and Ouwehand, L. (1984) Biochim. Biophys. Acta 764, 301–309). It is concluded that each of these S-state transitions involves the oxidation of an Mn(III) to Mn(IV). The spectrum and amplitude of the millisecond transient were in agreement with its assignment to the reduction of the oxidized secondary donor Z⁺ and the three Mn(IV) ions.