Effects of Mechanical Impedance on Root Growth in Barley (Hordeum vulgare L. ): I. EFFECTS ON THE ELONGATION AND BRANCHING OF SEMINAL ROOT AXES

Apparatus is described in which the root system of plants canbe exposed to accurately known external stresses in beds ofballotini for periods up to 3 weeks. A constant flow of aeratednutrient solution was supplied. Modifications of the apparatusallowed the emerging coleoptile also to be subjected to mechanicalstress. Earlier observations that roots are unable to decrease in diameterto enter small pores were confirmed. The effect of externalpressure on the rate of root elongation was independent of thediameter of pores provided this was smaller than the penetratingroot. Pressures of 20 and 50 kPa reduced the elongation of theseminal axes of barley by 50 and 80 per cent respectively. Significantreductions in root extension were also caused by 50 kPa or lessin more limited experiments with a number of other crop plants.The extension of the coleoptile and the first leaf respondedsimilarly to that of the roots. When the diameter of pores inthe rooting medium was intermediate between those of axes andlaterals, the former being impeded, an enhanced developmentof laterals led to a dense superficial root system. This considerablemodification of root form was without effect on nutrient uptakeprovided that an adequate supply of nutrients was availablein the restricted rooting zone. Attention is directed to some of the implications of these responsesfor plant growth under field conditions. However, considerationof the physiological mechanisms responsible for the effectsof low external pressures and the changes in cell structureto which they lead are deferred to subsequent papers.     

Effects of Mechanical Impedance on Root Growth in Barley, Hordeum vulgare L.: II. EFFECTS ON CELL DEVELOPMENT IN SEMINAL ROOTS

Anatomical changes induced in the roots of barley plants (Hordeumvulgare cv. Proctor) by an applied pressure of 2 ? 10⁴ Pa (0.2bar) have been examined; previous studies had shown that thelength of roots grown under these conditions was about 50% ofthat of unimpeded roots. Cryomicrotomy/light microscopy andscanning electron microscopy were used. Mechanical impedanceincreased the diameter of roots, this being due largely to anincreased thickness of the cortex; the number of cells in transversesection was increased and the diameter of the outer cells wasgreater though that of the inner ones was reduced.     

Lysine Catabolism in Barley (Hordeum vulgare L.)

Lysine catabolism in seedlings of barley (Hordeum vulgare L. var. Emir) was studied by direct injection of the following tracers into the endosperm of the seedlings: aspartic acid-3-(14)C, 2-aminoadipic acid-1-(14)C, saccharopine-(14)C, 2,6-diaminopimelic acid-1-(7)-(14)C, and lysine-1-(14)C. Labeled saccharopine was formed only after the administration of either labeled 2,6-diaminopimelic acid or labeled lysine to the seedlings. The metabolic fate of the other tracers administered also supported a catabolic lysine pathway via saccharopine, and apparently proceeding by a reversal of some of the biosynthetic steps of the 2-aminoadipic acid pathway known from lysine biosynthesis in most fungi. Pipecolic acid seems not to be on the main pathway of l-lysine catabolism in barley seedlings.     

Phosphoinositides in Barley (Hordeum vulgare L.) Aleurone Tissue

[3H]Inositol labeling of barley (Hordeum vulgare L. cv Himalaya) aleurone layers and analysis of phospholipids by deacylation revealed the presence of phosphatidylinositol (PtdIns), PtdIns3P, and PtdIns4P but not PtdInsP2 species. In contrast to an earlier report (P.P.N. Murthy, G. Pliska-Matyshak, L.M. Keranen, P. Lam, H.H. Mueller, N. Bhuvarahamurthy [1992] Plant Physiol 98: 1498-1501) systematic chemical degradation of PtdIns revealed no evidence of a second isomer of PtdIns. Evidence of the widespread occurrence of 3-phosphorylated PtdIns within the plant kingdom is presented.     

Growth Regulators and the Effect of Barley Yellow Dwarf Virus on Barley (Hordeum vulgare L.)

Estimation of cell number in the third leaf of barley (Hordeumvulgare L. C I 666) infected with barley yellow dwarf virus(BYDV) showed a marked decrease in the mitotic activity of theinfected plants Assay of endogenous gibberellins revealed adecrease in the level of a substance corresponding to gibberellicacid (GA₃) in BYDV-infected plants No significant differencein the level of endogenous auxins was observed Application ofgibberellic acid to infected plants reversed the dwarfing effectbut the response was not significantly different from the responseof healthy plants and was found to be due to increased cellelongation. It is suggested that the dwarfing of BYDV-infectedplants is a result of reduced mitotic activity This may be relatedto the reduced level of endogenous gibberellins.     

Apoplast Acidification in Growing Barley (Hordeum vulgare L.) Leaves

Apoplast acidification associated with growth is well documented in roots, coleoptiles, and internodes but not in leaves. In the present study, advantage was taken of the high cuticle permeability in the elongation zone of barley leaves to measure apoplast pH and growth in response to application of test reagents. The role of the plasma membrane H⁺-ATPase (PM-H⁺-ATPase) and K⁺ in this process was of particular interest. pH microelectrodes and an in vitro gel system with bromocresol purple as pH indicator were used to monitor apoplast pH. Growth was measured in parallel or in separate experiments using a linear variable differential transformer. Test reagents that blocked (vanadate) or stimulated (fusicoccin) PM-H⁺-ATPase or that reduced (Cs⁺, tetraethylammonium) K⁺ uptake were applied. Apoplast pH was lower in growing than in nongrowing leaf tissue and increased in the elongation zone with increasing apoplast K⁺. Vanadate increased apoplast pH and reduced growth, whereas fusicoccin caused the opposite effects. It is concluded that barley leaves exhibit acid-growth-type mechanisms in that apoplast pH is lower in elongating leaf tissue. Both growth and apoplast pH depend on the activity of the PM-H⁺-ATPase and K⁺ transport processes. However, not all of the growth displayed by leaves is dependent on a lower apoplast pH in the elongation zone; up to 50 % of growth is retained when apoplast pH in the elongation zone increases to a value observed in mature tissue.     

大麦转基因座位结构的研究

利用质粒营救法获得基因枪法转化的4种转绿色荧光蛋白基因(green fluorescent protein,GFP)大麦的转基因座位序列,序列分析显示4种材料的转基因座位中均有完整栽体的串联重复现象,表明转基因整合是同源重组的结果.同时转基因座位中也存在不完整载体片段、基因组片段的混杂排列,说明转基因整合时也发生异常重组.微粒轰击的转基因整合是由异常重组和同源重组共同完成的.     

Effects of Red and Blue LED Light on the Growth and Photosynthesis of Barley (Hordeum vulgare L.) Seedlings

According to the action spectrum of photosynthesis, photosynthetic efficiency is highest for red light. However, long-term growth with only red light leads to unfavorable changes in plant morphology, decrease in photosynthetic capacity and plant productivity. Detailed mechanisms behind these changes are still poorly understood. We studied the effects of narrow-band red (RL) and blue (BL) LED lighting on the morphology and photosynthesis of barley (Hordeum vulgare L.) seedlings at 9 days old, when energy for plant growth comes mostly from the endosperm, and light has a mainly morphogenic effect on plant growth. Plants grown with white fluorescent lamps (WL) were used as a control. At this developmental stage, light spectrum had small but significant effects on most morphometric parameters, which may become more prominent as the plant grows. These effects were more pronounced in RL-grown plants and were similar to the ‘shade-avoidance response’, which is unusual as in nature it occurs when the fraction of red light in the spectrum is low. RL-grown plants also had impaired photosynthetic photochemical efficiency (as assessed by PAM-fluorometry and leaf absorption). BL-grown plants had a stronger similarity to control plants in their morphology and photosynthetic characteristics than RL-grown plants; however, they had higher NPQ and different NPQ induction kinetics than WL- and RL-grown plants. Our results suggest that photoregulation of plant morphology and photosynthesis evolutionarily adapted to natural light is miscoordinated in narrow-band LED light. We discuss possible reasons for this miscoordination and for the formation of observed phenotypes on the level of photoreceptors.

     

Barley (Hordeum vulgare L.) inositol monophosphatase: gene structure and enzyme characteristics

The cellular myo-inositol (Ins) pool is important to many metabolic and signaling pathways in plants. Ins monophosphatase (IMPase; EC 3.1.3.25) activity is essential for the de novo synthesis of myo-Inositol (Ins), and for recycling of Ins in Ins(1,4,5)P3. However, proteins encoded by at least one family of IMP genes also have L-galactose-1-P phosphatase activity important to ascorbic acid synthesis, indicating a bifunctionality that links these two branches of carbon metabolism. As part of research into the regulation of Ins synthesis and supply during seed development, the barley IMP-1 gene and gene products were studied. The 1.4 kb barley IMP-1 promoter contains one low temperature response element (RE), two heat shock REs, one gibberellin and two auxin REs, and five sugar REs. Barley IMP-1 is expressed in all tissues assayed, and expression levels were not greatly altered by abiotic stress treatments. Reduced use of Ins for Ins P6 synthesis in developing seed of barley low phytic acid (lpa) mutants results in Ins accumulation, and IMP-1 expression is reduced in proportion to the increase in Ins level. The barley recombinant enzyme had a lower Km, indicating higher affinity, for D/L-Ins(3)P1 (Km = 9.7 microM) as compared with reported Km (Ins P1) values for other eukaryotic IMPases (43-330 microM) or with a reported Km (L-Gal-1P) of 150 microM for a kiwifruit (Actinidia deliciosa) enzyme. These and other data indicate that the barley IMP-1 gene is regulated at least in part in response to Ins metabolic needs, and that the enzyme it encodes displays catalytic properties well suited for a role in Ins synthesis, in addition to other roles as an L-gal-1-P phosphatase important to ascorbate synthesis, or as an IMPase important to Ins(1,4,5)P3 signal recycling.     

Proteins in Intercellular Washing Fluids from Leaves of Barley (Hordeum vulgare L.)

Primary leaves of barley were detached, infiltrated with variousbuffers, and centrifuged to yield ‘intercellular washingfluid’ (IWF). Effective pH control of the IWF was obtainedonly with Tris, among all buffers tried. In these liquids, upto 30 proteins were detected by gradient gel electrophoresis.Intracellular protein from injured cells at the cut ends ofleaves was present in IWF but did not contribute significantlyto the total protein recovered in this liquid. The yield ofprotein in the IWF depended on the buffer used for infiltrationand on the concentration of the buffer. Higher concentrationsof buffer yielded more protein. In other experiments leaves were infiltrated with Tris, centrifuged,and then infiltrated a second time with this buffer containingvarious concentrations of the zwitterionic detergent CHAPS,a sulphobetaine derivative of cholate. Gel electrophoresis ofthe IWF obtained after the second centrifugation revealed protein‘bands’ not detected when the detergent had beenomitted from the infiltration buffer. The electrophoretic patternsof protein ‘bands’ in the gels differed dependingon the CHAPS concentration used for infiltration. The effect of CHAPS on plasmalemma integrity was studied byobserving infiltrated tissue with the electron microscope andby treating isolated protoplasts with the detergent. After infiltrationwith CHAPS at 0.6 mM or 2.0 mM no plasmalemma breaks were detectedin leaves, and isolated protoplasts survived exposure to CHAPSat these concentrations for 2 h without bursting. Evidently,CHAPS at these low concentrations did not destroy the integrityof the plasmalemma; the additional protein recovered in theIWF under these conditions probably originated in the cell wall.Infiltration of leaves with 6.0 mM CHAPS resulted in breaksof the plasmalemma, in tissue collapse and leaf tip necrosis.Isolated protoplasts burst within minutes after being exposedto CHAPS at this concentration. Key words: Cell wall permeability, Intercellular space, Detergent, CHAPS, Protoplasts     

Inhibitory Effects of Jasmonic Acid and its Analogues on Barley (Hordeum vulgare L.) Anther Extrusion

Plant hormones play many important roles in plant growth and development. We previously tried to control barley (Hordeum vulgare L.) flowering in ear cultures treated with plant hormones and related compounds, and found that anther extrusion from florets was reduced by treatment with 100 ppm methyl jasmonate (MJA). In the present study, we show that 10 ppm MJA also inhibits anther extrusion in barley ear cultures. Spraying field-grown barley ears with MJA clarified the efficacy of this chemical under field conditions, but higher concentrations (100 to 1000 ppm MJA) were required. In addition, we investigated the activity of jasmonic acid (JA) analogues and cucurbic acid analogues in barley ear cultures. Some esters of JA showed similar effects to that of MJA, but effects of 9,10-dihydrojasmonic acid (DJA) and cucurbic acid derivatives were lower than that of MJA. In light of these data, we discuss the structural requirements for increased inhibition of barley anther extrusion.     

Mechanism of Septum Opening in Anthers of Two-rowed Barley (Hordeum vulgare L.)

To test the hypothesis that the rapid swelling of pollen grainsdriven by potassium movement opens the septum in anthers ofpoaceous plants, we studied (1) the behaviour of pollen grainsduring unfolding of the locule and (2) the distribution of potassiumin the locule in two-rowed barley. In the first experiment,the unfolding of decapitated anthers was observed. The pollengrains paved the inner wall of the locule during the unfoldingprocess, suggesting that the pollen grains bend the locule walloutward when they adhere to the wall. In the second experiment,the distribution of potassium in transverse sections of loculesin dehisced and indehisced anthers was observed. In indehiscedanthers, potassium was detected outside the pollen grains. Incontrast, in dehisced anthers, potassium was detected insidepollen grains. This suggested potassium ions moved from theinter-pollen space (locular fluid) into the pollen grains inthe locule at the time of pollen-grain swelling. Copyright 2000Annals of Botany Company Hordeum vulgare L., locule unfolding, pollen grain swelling, potassium ion, two-rowed barley     

Flow Cytometric Analysis and Chromosome Sorting of Barley (Hordeum vulgare L.)

Flow cytometric analysis was systematically performed to optimize the concentration and duration of hydroxyurea (DNA synthesis inhibitor) and trifluralin (metaphase blocking reagent) treatments for synchronizing the cell cycle and accumulating metaphase chromosomes in barley root tips. A high metaphase index (76.5% in the root tip meristematic area) was routinely achieved. Seedlings of about 1.0-cm length were treated with 1.25 mM hydroxyurea for 14 h to synchronize the root tip meristem cells at the S/G2 phase. After rinsing with hydroxyurea, the seedlings were incubated in a hydroxyurea-free solution for 2 h and were treated with 1 microM trifluralin for 4 h to accumulate mitotic cells in the metaphase. The consistent high metaphase index depended on the uniform germination of seeds prior to treatment. High-quality and high-quantity isolated metaphase chromosomes were suitable for flow cytometric analysis and sorting. Flow karyotypes of barley chromosomes were established via univariate and bivariate analysis. A variation of flow karyotypes was detected among barley lines. Two single chromosome types were identified and sorted. Bivariate analysis showed no variation among barley individual chromosomes in AT and GC content.     

Root contribution to nitrate reduction in barley seedlings (Hordeum vulgare L.)

Summary The leaf and root nitrate reductase activities were measured in 7 day-old barley seedlings by anoxic nitrite accumulation in darkness, during 48h after the transfer from a N-starved medium to a 1.5 mM K¹⁵NO₃ medium. Thisin situ nitrate reduction was compared with the¹⁵N incorporation in the reduced N fraction of the whole seedlings.The nitrate reduction integrated fromin situ measurements was lower than the reduced¹⁵N accumulation. The rootin situ nitrate reductase activity seemed to account for only the third of the real root nitrate reduction, which may have been responsible for the overall underestimation. This discrepancy was partly explained by the ability of the root to reduce nitrite in an anoxic environment.These results suggest that, after correction of thein situ estimation of the nitrate reduction. the roots contribute to about 50% of the total assimilation.     

Rubisco Content and Specific Activity in Barley (Hordeum vulgare L.): I. GENETIC VARIABILITY

Seven barley varieties, differing widely in their geographicorigin, were analysed at the seedling stage under controlledconditions for the following traits: (A) Soluble leaf proteincontent, (B) Rubisco content, (C) Total carboxylase activity,as well as for combinations of these basic characteristics:relative Rubisco concentration, relative carboxylase activity,and specific carboxylase activity of the enzyme. A very large genotypic variability was detected in all traits,especially for the total activity of Rubisco, with strong genotypiccorrelations between these traits, whereas environmental correlationswere almost non-existent. There were also relatively good correlationswith the gross assimilation rate (Pg) among the same range ofvarieties, measured in the adult stage in a previous investigation.For two out of five crosses analysed in the F₁ generation, significantdifferences were found between reciprocal hybrids. These results are discussed in terms of heredity patterns ofthe characteristics concerned, namely the double genetic determinismof Rubisco: a large subunit controlled by maternally inheritedchloroplast DNA and a small subunit controlled by nuclear DNA. Key words: Rubisco, barley, genetic variability, maternal inheritance     

Proton Flux and Elongation in Primary Roots of Barley (Hordeum vulgare L.)

The elongation zone of the primary root of barley (Hordeum vulgare L.) has been reported to be markedly basic in pH, in apparent contradiction of the acid-growth theory. We determined simultaneously the location of the elongation zone and the basic zone in these roots and found them indeed to be the same. However, sections of barley root elongation zones were found to respond to acidic, basic, and neutral solutions as predicted by the acid-growth theory.     

Properties and Regulation of Aspartate Kinase from Barley Seedlings (Hordeum vulgare L.)

Aspartate kinase (EC 2.7.2.4) has been purified 8-fold and characterized from germinating barley (Hordeum vulgare) seedlings. The enzyme is inhibited 50% by 0.7 mm l-lysine and almost completely at 5 mm. l-Methionine does not affect the enzyme on its own, but at low concentrations (0.1-1 mm) increases the inhibition in the presence of lysine, indicating that the two amino acids act as cooperative feedback regulators.     

The Uptake and Transport of Chromium by Barley Seedlings (Hordeum vulgare L.)

Uptake of ⁵¹CrO^2–₄ by intact barley seedlings was linearover 24 h and was stimulated by Ca2⁺ but inhibited by SO^2–₄and other Group VI anions. Uptake increased with increasingchromate concentration, but unless the concentration was high(100 µM) less than 1 per cent of the isotope absorbedwas transported to the shoots. The results of solvent extraction,subcellular fractionation and efflux studies indicated thatmost of the isotope accumulated by the roots was present ina soluble non-particulate form in the vacuoles. The possiblereasons for the restriction in chromate transport are discussedin relation to the metabolism of the element.     

Inositol Trisphosphate Metabolism in Subcellular Fractions of Barley (Hordeum vulgare L.) Mesophyll Cells

Phosphatases in cytosolic fractions, vacuoles, and vacuolar membranes from barley (Hordeum vulgare L.) leaves were found to dephosphorylate inositol 1,4,5-trisphosphate (IP3). 1,4-inositol bisphosphate (1,4-IP2) is the main product of IP3 dephosphorylation by the cytosolic fraction. The activity was strictly Mg2+ dependent. In contrast, IP3 dephosphorylation activity of both the soluble vacuolar and the tonoplast fractions was inhibited up to 50% by Mg2+. When vacuolar membranes were incubated with IP3, 1,4-IP2 was produced only under neutral and slightly alkaline conditions. Under acidic conditions, however, dephosphorylation yielded putative 4,5-inositol bisphosphate. Li+ (20 mM) and Ca2+ (100 [mu]M) strongly inhibited activity in the soluble vacuolar fraction but had only a slight effect on the activities of the cytosolic and tonoplast fractions.     

In vivo Nitrate Reductase Activity in Barley (Hordeum vulgare L.) during Ear Development

Experiments were conducted during the 1974–75 and 1975–76winter season with the barley (Hordeum vulgare L.) cultivarJyoti. From amongst the various plant parts, the flag leaf bladehad higher in vivo nitrate reductase (NR) activity than thelower two leaf blades, glumes, and grains. However, the potentialof a plant part to reduce NO₃^– is a function of its freshweight and the NR per unit fresh weight. On this basis, thesecond and third leaf blades could reduce more NO₃^– thanthe flag leaf blade. N fertilizer application resulted in enhancementof the activity of the leaf blades alone. N fertilizer appliedduring the reproductive phase was taken up and assimilated bythe various plant parts. The studies suggest that, even whenthe fertilizer is applied at optimum levels for obtaining maximumyields, the upper leaf blades have sub-optimal NR activity andthat there is a likelihood of either a preferential flow ofNO₃^– to the leaf blades or transnational barriers to NO₃^–movement to the ear.