Genes controlling hydroxylations of phytosiderophores are located on different chromosomes in barley (Hordeum vulgare L.)

Phytosiderophores, mugineic acids, have been demonstrated to be involved in Fe acquisition in gramineous plants. In this study, chromosomal arm locations of genes encoding for biosynthesis of various phytosiderophores were identified in a cultivar of barley (Hordeum vulgare L. cv. Betzes). Using wheat (Triticum aestivum L. cv. Chinese Spring)-barley (cv. Betzes) ditelosomic addition lines for 4HS and 4HL, a gene for hydroxylation of 2′-deoxymugineic acid to mugineic acid was localized to the long arm of barley chromosome 4H. To locate the gene for hydroxylation of mugineic acid to 3-epihydroxymugineic acid, hybrids between the 4H addition line and other wheat-barley addition lines were studied. Only a hybrid between 4H and 7H addition lines produced 3-epihydroxymugineic acid. The gene was further localized to the long arm of chromosome 7H by feeding mugineic acid to ditelosomic addition lines for 7HS and 7HL. A new phytosiderophore was discovered in both 7H and 7HL addition lines, which was identified to be 3-epihydroxy-2′-deoxymugineic acid by detailed nuclear magnetic resonance studies. These results revealed that in barley there are two pathways from 2′-deoxymugineic acid to 3-epihydroxymugineic acid: 2′-deoxymugineic acid → mugineic acid → 3-epihydroxymugineic acid and 2′-deoxymugineic acid → 3-epihydroxy-2′-deoxymugineic acid → 3-epihydroxymugineic acid. Barley genes encoding for the hydroxylations of phytosiderophores are located in different chromosomes and each gene hydroxylates different C-positions: the long arm of chromosome 4H carries the gene for hydroxylating the C-2′ position and the long arm of chromosome 7H carries the gene for hydroxylating the C-3 position of the azetidine ring. Received: 10 August 1998 / Accepted: 30 September 1998     

Chromosomal assignment and deletion mapping of barley EST markers

From about 10000 PCR-based EST markers of barley we chose 1421 EST markers that were demonstrated to be amplified differently by PCR between wheat (Triticum aestivum cv. Chinese Spring) and barley (Hordeum vulgare cv. Betzes). We assigned them to the seven barley chromosomes (1H to 7H) by PCR analysis using a set of wheat-barley chromosome addition lines. We successfully assigned 701 (49.3%) EST markers to the barley chromosomes: 75 to 1H, 127 to 2H, 119 to 3H, 94 to 4H, 108 to 5H, 81 to 6H and 97 to 7H. By using a set of Betzes barley telosomic addition lines of Chinese Spring, we could successfully determine the chromosome-arm (S or L) location of at least 90% of the EST markers assigned to each barley chromosome. We conducted a trial mapping using 90 EST markers assigned to 7HS (49) or 7HL (41) and 19 wheat lines carrying 7H structural changes. More EST markers were found in the distal region than in the proximal region.     

Why are young rice plants highly susceptible to iron deficiency?

The reason why young rice plant is highly susceptible to Fe-deficiency was clarified as follows: Among Gramineae plants rice secreted a very low amount of deoxy-MA as a phytosiderophore even under Fe-deficiency, and the secretion by rice ceased within 10 days under Fe-deficiency although barley secreted MAs during a period of more than one month. When iron depletion continued, the rice root tips become chimeric and epidermal cells became necrotic. The mitochondrial membrane systems in the cortex cells were also severely damaged. Iron starvation occurred even in the mitochondria, and energy charge in the root decreased. This reduced energy charge has firstly diminished the secretion activity of deoxy-MA from the roots, secondly reduced the activity of the transporter which absorb deoxy-MA-Fe^III chelate and finally reduced the synthesis of deoxy-MA from metionine. Consequently, the depletion of Fe^II in the shoot was induced and severe chlorosis rapidly developed in the young rice plant under Fe-deficiency.Abbreviations DCCD dicyclohexylcarbodiimide - CCCP carbonylcyanide-m-chlorophenylhydrazone - MA mugineic acid - MAs mugineic acid-family phytosiderophores, it contains deoxy-MA, MA, epihydroxy-MA, hydroxy-MA, avenic acid and distichonic acid     

Chromosomal Location of Genes Encoding Barley (1-->3, 1-->4)-beta-Glucan 4-Glucanohydrolases

Preparations of DNA from wheat (Triticum aestivum, cv Chinese Spring), barley (Hordeum vulgare, cv Betzes) and six euplasmic wheat-barley addition lines were digested to completion with restriction endonucleases and the products probed by Southern blot analysis using a cDNA-encoding barley (1→3, 1→4)-β-glucanase isoenzyme II. It is shown that one of the barley (1→3, 1→4)-β-glucanase genes is located on chromosome 1.     

Efficient production of wheat-barley hybrids and preferential elimination of barley chromosomes

Summary Intergeneric hybridization between four common wheat cultivars, Triticum aestivum L. cultivars Chinese Spring, Norin 12, Norin 61, and Shinchunaga, and cultivated barley, Hordeum vulgare L. cultivars Betzes, Nyugoruden, Harunanijou, and Kinai 5 were carried out in a greenhouse under 15 – 20 °C and long-day (15 h) photoperiod conditions. Two days prior to pollination, a 100 mg/1 2,4-D solution was injected into wheat stems. Among wheat cultivars, Norin 12, Norin 61, and Shinchunaga showed higher crossabilities than that of Chinese Spring, suggesting the presence of crossability gene(s) other than the kr system of Chinese Spring. Variation was also found among the barley cultivars as male parents. Betzes barley showed the highest crossability with wheat. Thus, the cross Norin 12×Betzes showed the highest crossability (8.25%), followed by Norin 61 ×Betzes (6.04%), Shinchunaga×Betzes (5.00%), and Shinchunaga×Kinai 5 (5.00%). The embryos were rescued by culture at 15–20 days after pollination. Seventyfour plants were obtained from 82 embryos. The morphology of the hybrid plants resembled that of wheat parents. Among 60 seedlings observed, 28 had 28 chromosomes, 8 had 21, 23 had aneuploid numbers of chromosomes (22–27), and 1 had 29 chromosomes. About half of the aneuploid hybrids showed mosaicism for chromosome number. By analyzing five isozyme markers of barley chromosomes, the chromosome constitutions of the aneuploid hybrids were determined. Barley chromosomes 1 and 5 were found to be preferentially eliminated in the hybrids, while chromosomes 2 and 4 were eliminated infrequently. The conditions and genetic factors for high crossability and the tendency of barley chromosome elimination are discussed.     

应用基因组原位杂交及RFLP标记鉴定小麦中的大麦染色体

用生物素（Biotin-6-dUTP)标记的大麦Betzes基因组DNA作探针,以普通小麦中国春总DNA作封阻进行基因组原位杂交（Genomeinsituhybridization,简称GISH）,从13株小麦-大麦杂交后代中鉴定出2个含有3条大麦Betzes2H染色体的材料（2n＝43）;2个2H单体异代换系（2n＝42）;7个2H二体异代换系（2n＝42）。用已定位在小麦第2部分同源群短臂上的探针psr131进行RFLP分析,结果表明大麦Betzes、代换系A5有1条区别于小麦中国春的特异带,A     

A Quantitative Scale of Spike Initial and Pistil Development in Barley and Wheat

Spring barley cv. Koru and spring wheat cv. Highbury were grownin constant controlled conditions of 16 h photoperiod, and temperaturesof 10 and 15 °C respectively. A range of spike growth anddevelopmental attributes were closely monitored from seedlingemergence to pollination in the most advanced floret. Simple, quantitative scales of development from seedling emergence(0) to pollination (10) are proposed, based on the morphogenesisof the spike initial, then the floret and finally the pistil.These scales allow developmental (ontogenetic) progress to bequantified without involving any attribute of growth or sizeof the plant or its organs. Hordeum sativum Jess., Triticum aestivum L., barley, wheat, quantification of development, ontogenesis, morphogenesis, growth, spike initial, pistil, gynoecium     

A New Class of Wheat Gliadin Genes and Proteins

The utility of mining DNA sequence data to understand the structure and expression of cereal prolamin genes is demonstrated by the identification of a new class of wheat prolamins. This previously unrecognized wheat prolamin class, given the name δ-gliadins, is the most direct ortholog of barley γ3-hordeins. Phylogenetic analysis shows that the orthologous δ-gliadins and γ3-hordeins form a distinct prolamin branch that existed separate from the γ-gliadins and γ-hordeins in an ancestral Triticeae prior to the branching of wheat and barley. The expressed δ-gliadins are encoded by a single gene in each of the hexaploid wheat genomes. This single δ-gliadin/γ3-hordein ortholog may be a general feature of the Triticeae tribe since examination of ESTs from three barley cultivars also confirms a single γ3-hordein gene. Analysis of ESTs and cDNAs shows that the genes are expressed in at least five hexaploid wheat cultivars in addition to diploids Triticum monococcum and Aegilops tauschii. The latter two sequences also allow assignment of the δ-gliadin genes to the A and D genomes, respectively, with the third sequence type assumed to be from the B genome. Two wheat cultivars for which there are sufficient ESTs show different patterns of expression, i.e., with cv Chinese Spring expressing the genes from the A and B genomes, while cv Recital has ESTs from the A and D genomes. Genomic sequences of Chinese Spring show that the D genome gene is inactivated by tandem premature stop codons. A fourth δ-gliadin sequence occurs in the D genome of both Chinese Spring and Ae. tauschii, but no ESTs match this sequence and limited genomic sequences indicates a pseudogene containing frame shifts and premature stop codons. Sequencing of BACs covering a 3 Mb region from Ae. tauschii locates the δ-gliadin gene to the complex Gli-1 plus Glu-3 region on chromosome 1.     

Two Related Biosynthetic Pathways of Mugineic Acids in Gramineous Plants

The biosynthesis of mugineic acids was studied by feeding 2H- or 13C-labeled compounds to water-cultured roots in several gramineous plants. The fate of labeled compounds was monitored by using 2H- and 13C-nuclear magnetic resonance. On investigating the proton changes during biosynthesis by feeding D,L-[3,3,4,4-d4]-methionine (98.6% 2H), 2H-labeled 2[prime]-deoxymugineic, mugineic, and 3-epihydroxymugineic acids were isolated from root washings of wheat (Triticum aestivum L. cv Minori), barley (Hordeum vulgare L. cv Minorimugi), and beer barley (Hordeum vulgare L. cv AM Nijo Tochigi), respectively. The 2H-nuclear magnetic resonance study indicated that 12 deuteriums were incorporated into the labeled 2[prime]-deoxymugineic acid, suggesting that three molecules of L-[3,3,4,4-d4]methionine were combined. In comparison, one of the deuteriums at C-2[prime] position in the mugineic acid, and one each of the deuteriums at C-2[prime] and C-3 positions in the 3-epihydroxymugineic acid, were lost. However, all other deuteriums were incorporated in a manner similar to that of the labeled 2[prime]-deoxymugineic acid. When [1,4[prime],4"-13C3]2[prime]-deoxymugineic acid (20% 13C) was fed to oat roots (Avena sativa L. cv Amuri II), avenic acid A, which was 13C enriched at the corresponding positions, was obtained. These results revealed that L-methionine was the precursor for all these mugineic acids and that cleavage of the azetidine ring or hydroxylation of the 2[prime]-deoxymugineic acid produced two related biosynthetic pathways in different gramineous plant species: L-methionine -> 2[prime]-deoxymugineic acid -> avenic acid A in oat; and L-methionine -> 2[prime]-deoxymugineic acid -> mugineic acid -> 3-epihydroxymugineic acid in barley and beer barley.     

Enhancement of ferric-mugineic acid uptake by iron deficient barley roots in the presence of excess free mugineic acid in the medium

To investigate the mechanism of mugineic acid-Fe^III uptake by barley roots, plasma membrane fractions were isolated from Fe-deficient barley roots using an aqueous two-phase partition method. Utilizing the plasma membrane vesicles, we developed an assay system for studying mugineic acid-⁵⁵Fe^III binding to the plasma membrane. However, no efficient active transport of mugineic acid-⁵⁵Fe^III into the plasma membrane vesicle was detected, because of large amount of non-specific adsorption of ⁵⁵Fe^III onto the vesicle. And the adsorption could be decreased by adding excess amount of free mugineic acid to the assay system. From the results it is speculated that an excess of free mugineic acids is necessary in the medium for effective uptake of mugineic acid-Fe^III by Fe-deficient barley roots. Support for this speculation came from a multi-compartment transport box experiment with excised roots of Fe-deficient barley.Abbreviations CCCP carbonylcyanide-m-chlorophenylhydrazone - MA mugineic acid     

Uptake of Lysine by Wild-Type and S-(2-Aminoethyl)-L-cysteine-Resistant Suspension-Cultured Cells of Triticum aestivum

The kinetics of uptake of L-lysine in wheat (Triticum aestivumcv. Chinese Spring) were analyzed in wild-type cells and inAEC-1 variant cells that are resistant to S-(2-aminoethyl)-L-cysteine(AEC). Uptake of lysine by AEC-1 cells was considerably slowerthan that by the wild-type cells. In the presence of carbonylcyanidem-chlorophenylhydrazone, the rates of uptake by both types ofcell were reduced to a similar linear component. Fitting theuptake data to one linear (diffusional) component and one Michaelis-Menten(active) system showed that, as compared to wild-type cells,AEC-1 cells have a reduced V_max and an increased K_m with respectto the active component, byt they have a similar diffusionalcomponent. Inhibition experiments with various amino acids indicatedthat the active component represents a carrier specific forbasic amino acids, which was competitively inhibited by AEC.The AEC-1 cells also showed reduced uptake of several neutraland acidic amino acids, but the rate of uptake of 3-O-methylglucosewas somewhat higher than that by wild-type cells. (Received May 16, 1989; Accepted September 4, 1989)     

Short communication. Biosynthesis of phytosiderophores in several Triticeae species with different genomes

To examine variation in phytosiderophore biosynthesis in Triticeae, phytosiderophores were investigated in wild and cultivated species of wheat and barley with different genomes. All wheats tested including hexaploid (AABBDD), tetraploid (AABB),and diploid (AA or DD) lines produced only one phytosiderophore, 2-deoxymugineic acid. The phytosiderophores biosynthesized in wild barleys varied among species. Using substitution-type triticale lines and wheat-barley addition lines. it was revealed that, in triticale, genes for the biosynthesis of both mugineic and hydroxymugineic acids were located in the long arm of chromosome 5R and that, in barley, the gene for production of mugineic acid was located in the long arm of chromosome 4H.     

Molecular and chromosomal organization of DNA sequences coding for the ribosomal RNAs in cereals

The chromosomal locations of ribosomal DNA in wheat, rye and barley have been determined by in situ hybridization using high specific activity ¹²⁵I-rRNA. The 18S-5.8S-26S rRNA gene repeat units in hexaploid wheat (cv. Chinese Spring) are on chromosomes 1B, 6B and 5D. In rye (cv. Imperial) the repeat units occur at a single site on chromosome 1R(E), while in barley (cv. Clipper) they are on both the chromosomes (6 and 7) which show secondary constrictions. In wheat and rye the major 5S RNA gene sites are close to the cytological secondary constrictions where the 18S-5.8S-26S repeating units are found, but in barley the site is on a chromosome not carrying the other rDNA sequences. — Restriction enzyme and R-loop analyses showed the 18S-5.8S-26S repeating units to be approximately 9.5 kb long in wheat, 9.0 kb in rye and barley to have two repeat lengths of 9.5 kb and 10 kb. Electron microscopic and restriction enzyme data suggest that the two barley forms may not be interpersed. Digestion with EcoR1 gave similar patterns in the three species, with a single site in the 26S gene. Bam H1 digestion detected heterogeneity in the spacer regions of the two different repeats in barley, while in rye and wheat heterogeneity was shown within the 26S coding sequence by an absence of an effective Bam H1 site in some repeat units. EcoR1 and Bam H1 restriction sites have been mapped in each species. — The repeat unit of the 5S RNA genes was approximately 0.5 kb in wheat and rye and heterogeneity was evident. The analysis of the 5S RNA genes emphasizes the homoeology between chromosomes 1B of wheat and 1R of rye since both have these genes in the same position relative to the secondary constriction. In barley we did not find a dominant monomer repeat unit for the 5S genes.     

Comparative mapping of HKT genes in wheat, barley, and rice, key determinants of Na+ transport, and salt tolerance

Salt tolerance of plants depends on HKT transporters (High-affinityK⁺ Transporter), which mediate Na⁺-specific transport or Na⁺-K⁺co-transport. Gene sequences closely related to rice HKT geneswere isolated from hexaploid bread wheat (Triticum aestivum)or barley (Hordeum vulgare) for genomic DNA southern hybridizationanalysis. HKT gene sequences were mapped on chromosomal armsof wheat and barley using wheat chromosome substitution linesand barley–wheat chromosome addition lines. In addition,HKT gene members in the wild diploid wheat ancestors, T. monococcum(A^m genome), T. urartu (A^u genome), and Ae. tauschii (D^t genome)were investigated. Variation in copy number for individual HKTgene members was observed between the barley, wheat, and ricegenomes, and between the different wheat genomes. HKT2;1/2-like,HKT2;3/4-like, HKT1;1/2-like, HKT1;3-like, HKT1;4-like, andHKT1;5-like genes were mapped to the wheat–barley chromosomegroups 7, 7, 2, 6, 2, and 4, respectively. Chromosomal regionscontaining HKT genes were syntenic between wheat and rice exceptfor the chromosome regions containing the HKT1;5-like gene.Potential roles of HKT genes in Na⁺ transport in rice, wheat,and barley are discussed. Determination of the chromosome locationsof HKT genes provides a framework for future physiological andgenetic studies investigating the relationships between HKTgenes and salt tolerance in wheat and barley. Key words: Barley, comparative mapping, HKT, rice, salt tolerance, sodium transport, wheat     

Salt Tolerance in the Triticeae: Solute Accumulation and Distribution in an Amphidiploid derived from Triticum aestivum cv. Chinese Spring and Thinopyrum bessarabicum

An amphidiploid derived by colchicine treatment of a hybridbetween Triticum aestivum cv Chinese Spring and Thinopyrum bessarabicumwas found to be more salt tolerant than the wheat cultivarsChinese Spring, Kharchia and Ciano 79 in terms of survival andgrain yield at 250 mol m^–3 NaCl. Tolerance was relatedto the ability of the amphidiploid to exclude Na and Cl fromthe shoots, and particularly from the young leaves, developinginflorescence and grain. There was no relationship between thesalt tolerance of the different species and varieties testedand changes in the concentrations of other solutes. The amphidiploiddid not inherit the high glycinebetaine concentrations characteristicof the wheatgrass parent. Amphidiploids produced from crossesbetween Thinopyrum species and wheat may be useful as stress-resistantnew crops. Key words: Salt stress, solute accumulation, Thinopyrum, Triticum     

Exogenous 4-hydroxybenzoic acid and salicylic acid modulate the effect of short-term drought and freezing stress on wheat plants

Exogenous salicylic acid has been shown to confer tolerance against biotic and abiotic stresses. In the present work the ability of its analogue, 4-hydroxybenzoic acid to increase abiotic stress tolerance was demonstrated: it improved the drought tolerance of the winter wheat (Triticum aestivum L.) cv. Cheyenne and the freezing tolerance of the spring wheat cv. Chinese Spring. Salicylic acid, however, reduced the freezing tolerance of Cheyenne and the drought tolerance of Chinese Spring, in spite of an increase in the guaiacol peroxidase and ascorbate peroxidase activity. The induction of cross tolerance between drought and freezing stress was observed: drought acclimation increased the freezing tolerance of Cheyenne plants and cold acclimation enhanced the drought tolerance. The induction of drought tolerance in Cheyenne was correlated with an increase in catalase activity.