Molecular dissection of variation in carbohydrate metabolism related to water-soluble carbohydrate accumulation in stems of wheat

Water-soluble carbohydrates (WSCs; composed of mainly fructans, sucrose [Suc], glucose [Glc], and fructose) deposited in wheat (Triticum aestivum) stems are important carbon sources for grain filling. Variation in stem WSC concentrations among wheat genotypes is one of the genetic factors influencing grain weight and yield under water-limited environments. Here, we describe the molecular dissection of carbohydrate metabolism in stems, at the WSC accumulation phase, of recombinant inbred Seri/Babax lines of wheat differing in stem WSC concentrations. Affymetrix GeneChip analysis of carbohydrate metabolic enzymes revealed that the mRNA levels of two fructan synthetic enzyme families (Suc:Suc 1-fructosyltransferase and Suc:fructan 6-fructosyltransferase) in the stem were positively correlated with stem WSC and fructan concentrations, whereas the mRNA levels of enzyme families involved in Suc hydrolysis (Suc synthase and soluble acid invertase) were inversely correlated with WSC concentrations. Differential regulation of the mRNA levels of these Suc hydrolytic enzymes in Seri/Babax lines resulted in genotypic differences in these enzyme activities. Down-regulation of Suc synthase and soluble acid invertase in high WSC lines was accompanied by significant decreases in the mRNA levels of enzyme families related to sugar catabolic pathways (fructokinase and mitochondrion pyruvate dehydrogenase complex) and enzyme families involved in diverting UDP-Glc to cell wall synthesis (UDP-Glc 6-dehydrogenase, UDP-glucuronate decarboxylase, and cellulose synthase), resulting in a reduction in cell wall polysaccharide contents (mainly hemicellulose) in the stem of high WSC lines. These data suggest that differential carbon partitioning in the wheat stem is one mechanism that contributes to genotypic variation in WSC accumulation.     

Linked gene networks involved in nitrogen and carbon metabolism and levels of water-soluble carbohydrate accumulation in wheat stems

High levels of water-soluble carbohydrates (WSC) provide an important source of stored assimilate for grain filling in wheat. To better understand the interaction between carbohydrate metabolism and other metabolic processes associated with the WSC trait, a genome-wide expression analysis was performed using eight field-grown lines from the high and low phenotypic tails of a wheat population segregating for WSC and the Affymetrix wheat genome array. The 259 differentially expressed probe sets could be assigned to 26 functional category bins, as defined using MapMan software. There were major differences in the categories to which the differentially expressed probe sets were assigned; for example, probe sets upregulated in high relative to low WSC lines were assigned to category bins such as amino acid metabolism, protein degradation and transport and to be involved in starch synthesis-related processes (carbohydrate metabolism bin), whereas downregulated probe sets were assigned to cell wall-related bins, amino acid synthesis and stress and were involved in sucrose breakdown. Using the set of differentially expressed genes as input, chemical–protein network analyses demonstrated a linkage between starch and N metabolism via pyridoxal phosphate. Twelve C and N metabolism-related genes were selected for analysis of their expression response to varying N and water treatments in the field in the four high and four low WSC progeny lines; the two nitrogen/amino acid metabolism genes demonstrated a consistent negative association between their level of expression and level of WSC. Our results suggest that the assimilation of nitrogen into amino acids is an important factor that influences the levels of WSC in the stems of field-grown wheat.     

Senescence and net photosynthesis of the flag leaf and grain growth of wheat in response to fungicide

Green leaf area and net photosynthesis of the flag leaves of Avalon and Maris Huntsman winter wheat crops were studied in relation to grain growth following the application of the fungicide propiconazole at flag leaf emergence. Disease levels were low during grain-fill, and were not significantly affected by the fungicide. Propiconazole significantly maintained green leaf area and photosynthesis per unit area during the period of rapid flag leaf senescence, but it had no effect on stem weight, grain growth or yield.     

小麦开花后旗叶中蔗糖合成与籽粒中蔗糖降解

在小麦开花后,旗叶中蔗糖磷酸合成酶（SPS）活性在开花后14d内一直维持较高水平,蔗糖合成酶（SS）的活性在开花后14-28d较高,蔗糖的含量与SPS活性呈显著正相关,籽粒中蔗糖合成酶（SS）在开花后28d内一直维持较高的活性;与此相对应,籽粒蔗糖的含量在开花后28d内呈明显的下降趋势。而旗叶和籽粒中SS活性均与籽粒淀粉的积累速率呈极显著正相关。     

大穗型小麦叶片性状、养分含量及氮素分配特征

采用田间试验研究了8个大穗型小麦新品系和西农979不同生育期比叶面积(SLA)、叶干物质含量(LDMC)与养分含量的差异性,并对其不同器官氮素积累和分配情况进行了探讨。结果表明,大穗型小麦的SLA和LDMC平均值小于西农979;小麦叶片养分含量由大到小依次为全氮(N)、全钾(K)和全磷(P);成熟期大穗型小麦品系各器官平均氮素积累能力都高于对照,氮素积累量及分配比例的大小顺序为籽粒>茎+鞘>穗草(穗轴+颖壳)>旗叶>倒二叶>余叶;大穗型小麦平均转移率和贡献率较西农979低;抽穗期旗叶SLA和LDMC呈负相关关系,SLA和N含量呈不显著的正相关,和P、K呈极显著的负相关。小麦叶片结构性状与养分含量影响体内有机物质的积累与转运,通过水肥调控措施来提高大穗型小麦养分利用效率的研究需进一步加强。     

Biochemical and genetic analysis of carbohydrate accumulation in Allium cepa L

Onion and shallot (Allium cepa L.) exhibit wide variation in bulb fructan content, and the Frc locus on chromosome 8 conditions much of this variation. To understand the biochemical basis of Frc, we conducted biochemical and genetic analyses of Allium fistulosum (FF)-shallot (A. cepa Aggregatum group) alien monosomic addition lines (AALs; FF+1A-FF+8A) and onion mapping populations. Sucrose and fructan levels in leaves of FF+2A were significantly lower than in FF throughout the year, and the springtime activity of acid invertase was also lower. FF+8A showed significantly higher winter sucrose accumulation and sucrose phosphate synthase (SPS) activity. Inbred high fructan (Frc_) lines from the 'W202Ax Texas Grano 438' onion population exhibited significantly higher sucrose levels prior to bulbing than low fructan (frcfrc) lines. Sucrose synthase (SuSy) activity in these lines was correlated with leaf hexose content but not with Frc phenotype. Markers for additional candidate genes for sucrose metabolism were obtained by cloning a major SPS expressed in onion leaf and exhaustively mining onion expressed sequence tag resources. SPS and SuSy loci were assigned to chromosome 8 and 6, respectively, using AALs and linkage mapping. Further loci were assigned, using AALs, to chromosomes 1 (sucrose phosphate phosphatase), 2 (SuSy and three invertases) and 8 (neutral invertase). The concordance between chromosome 8 localization of SPS and elevated leaf sucrose levels conditioned by high fructan alleles at the Frc locus in bulb onion or alien monosomic additions of chromosome 8 in A. fistulosum suggest that the Frc locus may condition variation in SPS activity.     

Sucrose metabolism in spring and winter wheat in response to high irradiance, cold stress and cold acclimation

Effects of low‐temperature stress, cold acclimation and growth at high irradiance in a spring (Triticum aestivum L. cv. Katepwa) and a winter wheat (Triticum aestivum L. cv. Monopol) were examined in leaves and crowns with respect to the sucrose utilisation and carbon allocation. Light‐saturated and carbon dioxide (CO₂)‐saturated rates of CO₂ assimilation were decreased by 50% in cold‐stressed spring and winter wheat cultivars. Cold‐ or high light‐acclimated Katepwa spring wheat maintained light‐saturated rates of CO₂ assimilation comparable to those of control spring wheat. In contrast, cold‐ or high light‐acclimated winter wheat maintained higher light and CO₂‐saturated rates of CO₂ assimilation than non‐acclimated controls. In leaves, during either cold stress, cold acclimation or acclimation to high irradiance, the sucrose/starch ratio increased by 5‐ to 10‐fold and neutral invertase activity increased by 2‐ to 2.5‐fold in both the spring and the winter wheat. In contrast, Monopol winter wheat, but not Katepwa spring wheat, exhibited a 3‐fold increase in leaf sucrose phosphate synthase (SPS) activity, a 4‐fold increase in sucrose:sucrose fructosyl transferase activity and a 6.6‐fold increase in acid invertase upon cold acclimation. Although leaves of cold‐stressed and high light‐grown spring and winter wheat showed 2.3‐ to 7‐fold higher sucrose levels than controls, these plants exhibited a limited capacity to adjust either sucrose phosphate synthase or sucrose synthase activity (SS[s]). In addition, the acclimation to high light resulted in a 23–31% lower starch abundance and no changes at the level of fructan accumulation in leaves of either winter or spring wheat when compared with controls. However, high light‐acclimated winter wheat exhibited a 1.8‐fold higher neutral invertase activity and high light‐acclimated spring wheat exhibited an induction of SS(d) activity when compared with controls. Crowns of Monopol showed higher fructan accumulation than Katepwa upon cold and high light acclimation. We suggest that the differential adjustment of CO₂‐saturated rates of CO₂ assimilation upon cold acclimation in Monopol winter wheat, as compared with Katepwa spring wheat, is associated with the increased capacity of Monopol for sucrose utilisation through the biosynthesis of fructans in the leaves and subsequent export to the crowns. In contrast, the differential adjustment of CO₂‐saturated rates of CO₂ assimilation upon high light acclimation of Monopol appears to be associated with both increased fructan and starch accumulation in the crowns.     

Leaf photosynthetic rate and mesophyll cell anatomy changes during ontogenesis in backcrossed <Emphasis Type="Italic">indica</Emphasis> × <Emphasis Type="Italic">japonica</Emphasis> rice inbred lines

The high-yielding indica rice variety, ‘Takanari’, has the high rate of leaf photosynthesis compared with the commercial japonica varieties. Among backcrossed inbred lines from a cross between ‘Takanari’ and a japonica variety, ‘Koshihikari’, two lines, BTK-a and BTK-b, showed approximately 20% higher photosynthetic rate than that of ‘Takanari’ for a flag leaf at full heading. This is a highest recorded rate of rice leaf photosynthesis. Here, the timing and cause of the increased leaf photosynthesis in the BTK lines were investigated by examining the photosynthesis and related parameters, as well as mesophyll cell anatomy during ontogenesis. Their photosynthetic rate was greater than that of ‘Takanari’ in the 13th leaf, as well as the flag leaf, but there were no differences in the 7th and 10th leaves. There were no consistent differences in the stomatal conductance, or the leaf nitrogen and Rubisco contents in the 13th and flag leaves. The total surface area of mesophyll cells per leaf area (TA_mes) in the 13th and flag leaves increased significantly in the BTK lines due to the increased number and developed lobes of mesophyll cells compared with in ‘Takanari’. The mesophyll conductance (g _m) became greater in the BTK lines compared with ‘Takanari’ in the flag leaves but not in the 10th leaves. A close correlation was observed between TA_mes and g _m. We concluded that the increased mesophyll conductance through the development of mesophyll cells during the reproductive period is a probable cause of the greater photosynthetic rate in the BTK lines.     

Comparison of fructan dynamics in two wheat cultivars with different capacities of accumulation and remobilization under drought stress

Remobilization of stored carbohydrates in the stem of wheat plants is an important contributor to grain filling under drought stress (DS) conditions. A massive screening on Iranian wheat cultivars was performed based on stem dry weight changes under well-watered and DS conditions. Two cultivars, Shole and Crossed Falat Hamun (CFH), with different fructan accumulation and remobilization behavior were selected for further studies. Water-soluble carbohydrates (WSCs) and fructan metabolizing enzymes were studied both in the stem penultimate and in sucrose (Suc) treated, excised leaves. Under drought, CFH produced higher grain yields than Shole (412 vs 220 g m(-2)). Also, grain yield loss under drought was more limited in CFH than in Shole (17 vs 54%). Under drought, CFH accumulated more graminan-type fructo-oligosaccharides than Shole. After anthesis, fructan 6-exohydrolase (6-FEH; EC 3.2.1.154) activities increased more prominently than fructan 1-exohydrolase (EC 3.2.1.153) activities during carbon remobilization. Interestingly, CFH showed higher 6-FEH activities in the penultimate than Shole. The field experiment results suggest that the combined higher remobilization efficiency and high 6-FEH activities in stems of wheat could contribute to grain yield under terminal drought. Similar to the penultimate, fructan metabolism differed strongly in Suc-treated detached leaves of selected cultivars. This suggests that variation in the stem fructan among wheat cultivars grown in the field could be traced by leaf blade induction experiments.     

Diurnal Growth of Tall Fescue Leaf Blades : II. Dry Matter Partitioning and Carbohydrate Metabolism in the Elongation Zone and Adjacent Expanded Tissue

The spatial distributions of net deposition rates of water soluble carbohydrate-free dry matter (WSC-free DM) and WSC were evaluated within and above the elongation zone of tall fescue (Festuca arundinacea Schreb.) leaf blades during light and darkness. Imported DM used for WSC-free DM synthesis during darkness (67% of the total in experiment I and 59% in experiment II) was greater than during light (47% in both experiments), suggesting that the 65% higher leaf elongation rate during darkness was accompanied by higher rates of synthesis of cellular structural components. Deposition rates of WSC in the basal and central part of the elongation zone (0-20 mm from the ligule) were similar during light and darkness, but above 20 millimeters WSC deposition occurred during light and WSC loss occurred during darkness. WSC deposition and loss throughout the elongation zone and the recently expanded tissue were mostly due to net synthesis and degradation of fructan. Fructan was predominantly low molecular weight and contributed about 50% of the total osmotic partial pressure of WSC. In the most actively growing region, where fructan synthesis was most rapid, no diurnal change occurred in molecular weight distribution of fructan. WSC solute concentrations were diluted in the most actively growing tissue during darkness because net monosaccharide and fructan deposition were unaltered and sucrose deposition was decreased, but growth-associated water deposition was increased by 77%. Net rates of fructan synthesis and degradation were not related to tissue sucrose concentration, but appeared to respond to the balance between assimilate import and assimilate use in synthesis of cellular structural components (i.e. WSC-free DM) and deposition of monosaccharides. Fructan synthesized in tissue during most active elongation was degraded when the respective tissue reached the distal limit of the elongation zone where assimilate import in darkness was insufficient to maintain synthetic processes associated with further differentiation of cells.     

Expression analysis of the genes involved in accumulation and remobilization of assimilates in wheat stem under terminal drought stress

Stem reserve mobilization and expression of major genes involved in fructan metabolism during grain filling in wheat (Triticum aestivum L.) cultivars, Zagros and Marvdasht were studied under terminal drought through withholding water at the anthesis. Mobilized dry matter, maximum specific weight and mobilization efficiency were observed to be higher in the internodes of tolerant cultivar (Zagros), both under well-watered and stress conditions, which resulted in enhanced translocation of stem reserves to the grains. Water soluble carbohydrates (WSC) and its constituent compounds were observed to be higher in the internodes of Zagros than those of sensitive cultivar (Marvdasht). Maximum relative expression of 1-SST, 6-SFT, INV, 1-FEHw1, 1-FEHw2, 1-FEHw3 and 6-FEH was significantly higher in the peduncle and penultimate of Zagros compared to Marvdasht cultivar under both drought and well-watered conditions. Expression of 1-FEHw3 and 6-FEH were increased during carbon remobilization in Zagros cultivar, suggesting that both genes are necessary for an efficient degradation and translocation of stem fructans. The mRNA levels of two fructan synthetic enzymes (1-SST and 6-SFT) in the stem were positively correlated with stem WSC concentrations, while the mRNA levels of enzymes involved in fructan hydrolysis (INV, 1-FEHw3 and 6-FEH) were inversely correlated with WSC concentration. According to the achieved results, it can be concluded that certain characteristics of Zagros cultivar, enhanced capability of fructan storage, higher mobilization efficiency and high gene expression level of 1-SST, 6-SFT, 1-FEHw3 as well as 6-FEH genes might help the drought tolerant cultivar to cope the stress conditions.     

Effect of assimilate utilization on photosynthetic rate in wheat

Summary Two weeks after anthesis, when the grain is filling rapidly, the rate of photosynthesis by flag leaves of wheat cv. Gabo was between 20 and 30 mg CO₂ dm^-2 leaf surface hour^-1 under the conditions used. About 45% of flag-leaf assimilates were translocated to the ear, and only about 12% to the roots and young shoots.On removing the ear, net photosynthesis by the flag leaves was reduced by about 50% within 3–15 hours, and there was a marked reduction in the outflow of ¹⁴C-labelled assimilates from the flag leaves.Subsequent darkening of all other leaves on plants without ears led to recovery of flag-leaf photosynthesis, as measured by gas analysis and ¹⁴CO₂ fixation, and to increased translocation of assimilates to the roots and young shoots. Minor changes in the rates of dark respiration accompanied these major, reversible changes in photosynthetic rate.After more than a week in continuous, high-intensity light, the rate of photosynthesis by flag leaves of intact plants had fallen considerably, but could be restored again by a period in darkness, or by inhibiting photosynthesis in the ears by spraying them with DCMU. The inhibition of ear photosynthesis increased translocation of labelled assimilates from the flag leaf to the ears, without affecting leaf sugar levels.The application of TIBA to the culm below the ear inhibited auxin movement throught the culm, but had no influence on flag-leaf photosynthesis.These results suggest that, at least in this system, photosynthesis by the flag leaf is regulated directly by the demand for assimilates from the flag leaf and not indirectly through action in the leaf of auxins produced by the sink organs.     

Differential Expression of Genes in the Leaves of Sugarcane in Response to Sugar Accumulation

In C₄ sugarcane (Saccharum spp. hybrids), photosynthetic activity has been shown to be regulated by the demand for carbon from sink tissues. There is evidence, from other plant species, that sink-limitation of photosynthesis is facilitated by sugar-signaling mechanisms in the leaf that affect photosynthesis through regulation of gene expression. In this work, we manipulated leaf sugar levels by cold-girdling leaves (5°C) for 80 h to examine the mechanisms whereby leaf sugar accumulation affects photosynthetic activity and assess whether signaling mechanisms reported for other species operate in sugarcane. During this time, sucrose and hexose concentrations above the girdle increased by 77% and 81%, respectively. Conversely, leaf photosynthetic activity (A) and electron transport rates (ETR) decreased by 66% and 54%, respectively. Quantitative expression profiling by means of an Affymetrix GeneChip Sugarcane Genome Array was used to identify genes responsive to cold-girdling (56 h). A number of genes (74) involved in primary and secondary metabolic pathways were identified as being differentially expressed. Decreased expression of genes related to photosynthesis and increased expression of genes involved in assimilate partitioning, cell wall synthesis, phosphate metabolism and stress were observed. Furthermore four probe sets homologous to trehalose 6-phosphate phosphatase (TPP; EC 5.3.1.1) and trehalose 6-phosphate synthase (TPS; EC 2.4.1.15) were up- and down-regulated, respectively, indicating a possible role for trehalose 6-phosphate (T6P) as a putative sugar-sensor in sugarcane leaves.     

Exogenous nitric oxide effect on fructan accumulation and FBEs expression in chilling-sensitive and chilling-resistant wheat

This study was to investigate the effect of exogenous nitric oxide (NO) on fructan accumulation and fructan biosynthesic enzymes (FBEs) expression in seedlings leaves of two wheat (Triticum aestivum L.) cultivars, winter wheat (Zhoumai18, ZM) and spring wheat (Yanzhan4110, YZ), under 4 °C. The seedlings of two wheat cultivars were subjected to different concentrations of sodium nitroprussiate (SNP) for 0, 24, 48, and 96 h. Relative water content (RWC) was increased by exogenous NO in YZ, but decreased in ZM. Except for glucose, fructose and fructans of degree of polymerization (DP) 3 in YZ, other soluble carbohydrates contents in the two wheat cultivars all increased to different degrees. The activities of FS (including sucrose: sucrose 1-fructosyltransferase (1-SST, EC: 2.4.1.99) and sucrose: fructan 6-fructosyltransferase (6-SFT, EC: 2.4.1.10)) were significantly higher than fructan: fructan 1-fructosyltransferase (1-FFT, EC: 2.4.1.100) in the seedlings of two wheat cultivars. The same phenomenon occurred to FBEs expression. In addition, sucrose content decreased while fructans content increased under low temperature, which was in accordance with the improved 1-FFT activity in ZM. Moreover, fructans content increased to a high level under high concentration of NO in ZM while kept at a constant low level in YZ. The expression levels of FBEs were universally higher in ZM than in YZ, which identified with the high frost resistance of the winter cultivar. It is concluded that exogenous NO treatment on wheat may be a good option to reduce chilling injury by regulating fructan accumulation in leaves. This is the first report owing that exogenous NO alleviated the negative effects of chilling stress by accumulating fructans in wheat.