An analysis of relative elemental growth rate, epidermal cell size and xyloglucan endotransglycosylase activity through the growing zone of ageing maize leaves

The effect of development on leaf elongation rate (LER) andthe distribution of relative elemental growth rate (REGR), epidermalcell length, and xyloglucan endotransglycosylase (XET) activitythrough the growing zone of the third leaf of maize was investigated.As the leaf aged and leaf elongation slowed, the length of thegrowing zone (initially 35 mm) and the maximal REGR (initially0.09 mm mm^–1 h^–1) declined. The decline in REGRwas not uniform through the growth profile. Leaf ageing sawa maintenance of REGR towards the base of the leaf. Epidermalcell size was not constant at a given position in the growingzone, but was seen to increase as the leaf aged. There was apeak of XET activity close to the base of the growing zone.The peak of XET activity preceded the zone of maximum REGR.XET activity declined as leaves aged and their elongation rateslowed. When leaf elongation was complete a distinct peak ofXET activity remained close to the base of the leaf. Key words: Leaf elongation rate (LER), relative elemental growth rate (REGR), xyloglucan endotransglycosylase (XET)     

Expression of XET-related genes and its relation to elongation in leaves of barley (Hordeum vulgare L.)

Five cDNA clones were isolated from barley (Hordeum vulgare L.) that encoded mRNAs related to xyloglucan endotransglycosylase (XET). One of the clones encoded a protein with XET activity in vitro. Sequence comparisons revealed five families of XET-related sequences, one of which (containing two of the barley genes) was novel. Hybridization studies using clone-specific probes indicated that the corresponding genes were represented once, or possibly twice, in the barley genome. Treatment of dwarf mutants with gibberellic acid (GA₃), or homozygosity at the ‘slender’ (sln1) locus, resulted in a 2.5-fold (approximately) stimulation of blade elongation rate. Three of the five clones detected mRNAs that were maximally expressed towards the base of the blade, and present in greater quantities in GA₃-treated or slender seedlings. The remaining two clones detected mRNAs that were maximally expressed in the middle of the blade. Relative elemental growth rate (REGR) profiles of leaves growing with or without GA₃ treatment revealed similar maximal REGR values despite a 2.5-fold difference in leaf elongation rate. Segments of GA₃-treated leaves attained their maximal REGR values more rapidly, this being associated with enhanced expression of the three ‘basal’ XET-related mRNAs. Highest XET activities were detected in the base of the elongation zone, and in GA₃-treated seedlings a second activity peak was observed near the distal end of the elongation zone. We conclude that there are likely to be several XET isoenzymes with different expression patterns, and identify those XET-related proteins potentially involved in leaf elongation.     

Mechanism for increased leaf growth in elevated CO2

The effect of exposure to elevated CO₂ on the processes of leafcell production and leaf cell expansion was studied using primaryleaves of Phaseolus vulgaris L. Cell division and expansionwere separated temporally by exposing seedlings to dim red lightfor 10 d (when leaf cell division was completed) followed byexposure to bright white light for 14 d (when leaf growth wasentirely dependent on cell expansion). When plants were exposedto elevated CO₂ during the phase of cell expansion, epidermalcell size and leaf area development were stimulated. Three piecesof evidence suggest that this occurred as a result of increasedcell wall loosening and extensibility, (i) cell wall extensibility(WEx, measured as tensiometric extension using an Instron) wassignificantly increased, (ii) cell wall yield turgor (V, MPa)was reduced and (iii) xyloglucan endotransglycosylase (XET)enzyme activity was significantly increased. When plants wereexposed to elevated CO₂ during the phase of cell division, thenumber of epidermal cells was increased whilst final cell sizewas significantly reduced and this was associated with reducedfinal leaf area, WEx and XET activity. When plants were exposedto elevated CO₂ during both phases of cell division and expansion,leaf area development was not affected. For this treatment,however, the number of epidermal cells was increased, but cellexpansion was inhibited, despite exposure to elevated CO₂ duringthe expansion phase. Assessments were also made of the spatialpatterns of WEx across the expanding leaf lamina and the datasuggest that exposure to elevated CO₂ during the phase of leafexpansion may lead to enhanced extensibility particularly atbasal leaf margins which may result in altered leaf shape. The data show that both cell production and expansion were stimulatedby elevated CO₂, but that leaf growth was only enhanced by exposureto elevated CO₂ in the cell expansion phase of leaf development.Increased leaf cell expansion is, therefore, an important mechanismfor enhanced leaf growth in elevated CO₂, whilst the importanceof increased leaf cell production in elevated CO₂ remains tobe elucidated. Key words: Phaseolus vulgaris L., dwarf beans, elevated CO², biophysics of cell expansion, xyloglucan endotransglycosylase, XET, water relations     

The Effect of (2-chloroethyl)-trimethylammonium Chloride on the Growth of Barley

Barley (Hordeum vulgare L. C.I.666) was shown to be susceptibleto the growth retardant (2-chloroethyl)-trimethylammonium chloride(CCC). The estimation of cell number in the dwarfed third leafblade indicated that a decrease in mitotic activity had occurredin treated plants. There was also a decrease in cell size intreated plants. The dwarfing action of CCC was reversed by exogenousgibberellic acid (GA₃) but this was shown to be the result ofincreased cell elongation only. GA₃ did not promote cell divisionin healthy or CCC-treated plants. Assay of endogenous gibberellinsshowed a significant reduction in the level of a substance correspondingto GA₃ in CCC-treated plants. It is suggested that CCC-induceddwarfing of barley is largely the result of a reduction in meristematicactivity. This may be related to an effect on gibberellin biosynthesisbut is not reversed by the application of exogenous GA ₃.     

Gibberellin-regulated XET is differentially induced by auxin in rice leaf sheath bases during gravitropic bending

The asymmetric distribution of auxin plays a fundamental role in plant gravitropism, yet little is understood about how its lateral distribution stimulates growth. In the present work, the asymmetric distribution not only of auxin, but also that of gibberellins (GAs), was observed in rice leaf sheath bases following gravistimulation. Gravistimulation induced the transient accumulation of greater amounts of both IAA and GA in the lower halves of the leaf sheath bases of rice seedlings. OsGA3ox1, a gene of active GA synthesis, was differentially induced by gravistimulation. Furthermore, 2,3,5-tri-iodobenzoic acid (TIBA), an inhibitor of auxin transport, substantially decreased the asymmetric distribution of IAA and the gradient of OsGA3ox1 expression. Externally applied GA(3) restored the gravitropic curvature of rice leaf sheaths inhibited by either TIBA or by ancymidol, a GA synthesis inhibitor. The expression of XET (encoding xyloglucan endotransglycosylase) was differentially induced in the lower halves of gravistimulated leaf sheath bases and was also up-regulated by exogenous IAA and GA(3). Both ancymidol and TIBA decreased the gradient of XET expression. These data suggest that the asymmetric distribution of auxin effected by gravistimulation induced a gradient of GAs via asymmetric expression of OsGA3ox1 in rice leaf sheath bases, and hence caused the asymmetric expression of XET. Cell wall loosening in the curvature site of the leaf sheath triggered by the expression of XET would contribute to gravitropic growth.     

Distribution of Endogenous Gibberellins in Vegetative Shoots of Rice

Levels of endogenous gibberellins in rice seedlings (Oryza sativaL., cv. Nipponbare) were compared between young and old leavesat the 4- and 5-leaf stages. The levels of GA₁, GA₁₉ and GA₅₃were higher in the youngest leaf than in older leaves at the5-leaf stage, but they did not differ significantly betweenthe leaf sheath and the leaf blade. At the 4-leaf stage, thelevel of GA₁, was highest in the third leaf sheaths which containedyoung elongating tissues. These results indicate that gibberellinsare synthesized in young vegetative tissues to promote theirelongation growth. The levels of GA₁ in the youngest leaf sheathsof two cultivars of dwarf rice, Tan-ginbozu and Waito-C, wereapproximately 10% of that in the normal rice at the 5-leaf stage.This result could explain the retardation of shoot elongationin these dwarf cultivars. (Received February 15, 1995; Accepted June 1, 1995)     

Xyloglucan endotransglycosylase activity in pea internodes. Effects of applied gibberellic acid.

Xyloglucan endotransglycosylase (XET) activity extractable from internodes of tall and dwarf varieties of pea (Pisum sativum L.) was assayed radiochemically using tamarind seed xyloglucan as donor substrate and an oligosaccharidyl-[3H]alditol as acceptor substrate. Internodes I and II showed little elongation during the period 15 to 21 d after sowing; XET activity remained relatively constant and was unaffected by exogenous gibberellic acid (GA3). A single application of GA3 to the dwarf genotype resulted in a small enhancement of elongation in internode III between d 17 and 21 and caused a small increase in XET activity in internode III. Repeated applications of GA3 caused internode V to elongate between d 20 and 26, to the same extent as in the tall variety, and concomitantly led to greatly elevated XET activity (expressed per unit fresh weight, per unit of extractable protein, and per internode). Thus, XET activity correlated with GA3-enhanced length in pea internodes; the possibility that this represents a causal relationship is discussed.     

Inhibition of Gibberellic Acid-Induced Elongation-Growth of Pea Epicotyls by Xyloglucan Oligosaccharides

The biological activity of a cell wall-derived xyloglucan nonasaccharide(XG9) was investigated using a bioassay with entire pea epicotyls(Pisum sativum cv. Progress). The xyloglucan fragment was foundto inhibit gibberellic acid-induced elongation of etiolatedpea epicotyls with maximum inhibition at concentrations rangingfrom 10^–11 to 10^–9M. Growth of etiolated epicotylsin the absence of exogenously applied GA₃ was also inhibitedby XG9 in the same concentration range. A cell wall-derivedheptasaccharide (XG7) lacking the fucosyl-galactosyl-side chainshowed no inhibitory effect in the pea epicotyl bioassay withand without exogenous GA₃. Furthermore, the biological activityof a synthetic pentasaccharide (XG5), containing the fucosylgalactosyl-sidechain which is necessary for the biological activity was investigatedin the same bioassay. Compared to XG9 the pentasaccharide hada similar inhibitory activity on GA₃-promoted elongation aswell as on the endogenous growth in the absence of exogenouslyapplied GA₃, but did not exhibit a distinct concentration optimum. Key words: Elongation-growth, gibberellic acid (GA₃), oligosaccharides, pea, XG9     

Regulation of Leaf Shape in the Solanifolia Mutant of Tomato (Lycopersicon esculentum) by Plant Growth Substances

The solanifolia mutant (sf/sf) of tomato (Lycopersicon esculentum)produces leaves consisting of leaflets with entire margins,unlike the lobed leaflets of normal plants. Normal plants treatedwith gibberellic acid (GA₃) produced leaves with entire marginswhereas mutant plants exposed to 2-chloroethyl-trimethyl ammoniumchloride (CCC)—an inhibitor of gibberellin biosynthesis—producedlobing of leaflets. The leaf area of the mutant was significantlygreater than that of the normal, but was not significantly differentfrom GA₃-treated normal leaves. Similarly, in CCC-treated mutantleaves the leaf area was not different from that of normal untreatedleaves. These observations suggest that the sf/sf mutation affectsthe leaf shape through its effect on endogenous gibberellinsand/or inhibitory substances. Leaf shape, Lycopersicon esculentum, plant growth substances, tomato     

Morphogenetic Effects on Vegetative Plants of Poa pratensis L. of 6-Azauracil, (2-Chloroethyl)- phosphonic acid, and (2-Chloroethyl) trimethyl-ammonium chloride and their Interaction with Gibberellic Acid

Fresh and dry weights and leaf size of Poa pratensis were reducedwhen treated with 6-azauracil (AzU), (2-chloroethyl)phosphonicacid (CEPA), or (2-chloroethyl)trimethylammonium chloride (CCC).AzU and CEPA inhibited epidermal cell division without inhibitingcell elongation, while CCC inhibited mainly cell elongationand cell division to a small extent. The ratio of blade lengthto sheath length and the blade length/width ratio were reduced,but leaf emergence and tillering were increased by AzU and CEPA.CCC affected only the latter three features. Like GA₃, CEPAinduced stem formation, but internodes were shorter. GA₃ was ineffective in preventing leaf-growth inhibition byAzU, which inhibited Ga₃-induced cell elongation. The inhibitoryeffect of CEPA on leaf growth was apparently reversed by GA₃,but this was due solely to increased cell elongation, the reductionin cell number being unaffected. Ga₃ reversed the effect ofCCC on leaf length, as well as on cell size and number. Simultaneousapplication of the inhibitors produced a complex interactionin reducing leaf length and number and size of epidermis cells.It is postulated that AzU, CEPA, and CCC have different modesof action because they have specific effects on plant growthand different effects on GA₃-induced cell elongation.     

Effects of Plant Growth Regulators and Nutrient Supply on Tiller Bud Outgrowth in Barley (Hordeum distichum L.)

Seedlings of spring barley were raised in 100 and 20% nutrientsolution and treated with a foliar application of Terpal, Cerone,TIBA, GA₃ or BAP. The growth of individual tiller buds and tillers,the main shoot and the root system was recorded over the following15 d. Terpal and Cerone stimulated tiller bud elongation within5 d at both nutrient levels and after 15 d the number of emergedtillers was increased at the higher nutrient level. Terpal characteristicallypromoted the growth of secondary tiller buds whereas Ceronepromoted the emergence of the coleoptile tiller; both thesePGRs also retarded the development of the main shoot. TIBA increasedthe number of elongating tiller buds but this did not resultin greater tillering. GA₃ reduced the number of elongating tillerbuds and restricted their growth, especially in the high nutrientregime; this was accompanied by an increase in main shoot elongation.The growth and development of tiller buds was reduced by BAPand the number of emerged tillers was reduced at 15 d in bothnutrient levels; main shoot dry weight and root elongation werealso reduced. The results are considered in relation to theoverall influence of hormonal factors and mineral supply ontiller bud outgrowth. Hordeum distichum, spring barley, tiller bud outgrowth, plant growth regulators, Terpal, Cerone, GA₃, BAP, nutrient supply, apical dominance, TIBA     

The effects of temperature and the Rht3 dwarfing gene on growth, cell extension, and gibberellin content and responsiveness in the wheat leaf

The effects of low temperature and the Rht3 dwarfing gene onthe dynamics of cell extension in leaf 2 of wheat were examinedin relation to gibberellin (GA) content and GA-responsivenessof the extension zone. Leaf 2 of wild-type (rht3) wheat closelyresembled that of the Rht3 dwarf mutant when seedlings weregrown at 10C. The maximum relative elemental growth rate (REGR)within the extension zone in both genotypes was lower at 10Cthan at 20C, but the position with respect to the leaf basewas unaffected by temperature. The size of the extension zoneand epidermal cell lengths were similar in both genotypes at10C. Growth at 20C, instead of 10C, increased the lengthof the extension zone beyond the point of maximum REGR in thewild type, but not in the Rht3 mutant. Increasing temperatureresulted in longer epidermal cells in the wild type. Treatingwild-type plants at 10C with gibberellic acid (GA₃) also increasedthe length of the extension zone, but the Rht3 mutant was GA-non-responsive.However, the concentrations of endogenous GA₁ and GA₃ remainedsimilar across the extension zone of wild-type plants grownat both temperatures, despite large differences in leaf growthrates. The period of accelerating REGR as cells enter the extensionzone, and the maximum REGR attained, are apparently not affectedby GA. It is proposed that GA functions as a stimulus for continuedcell extension by preventing cell maturation in the region beyondmaximum REGR and that low temperature increases the sensitivitythreshold for GA action. Key words: Cell extension, gibberellin, Rht3 dwarfing gene, temperature, wheat leaf     

Changes in the Sugar Content of the Leaf Sheath of Dwarf Maize Seedlings Associated with Gibberellin-Induced Growth

Changes in the time in sugar and starch content of the firstleaf sheath were compared among normal, d₅, and GA₁-treated(1µg/plant) d₅ seedlings of Zea mays L., from which endospermshad been removed. The patterns of changes in the levels of thesecarbohydrates were more or less similar to one another amongthe three groups of seedlings. Generally, the levels decreasedtoward the time of maturation of the leaf sheath except forthe level of sucrose which showed a sharp increase followedby a decrease. There was an inverse relationship over time betweengrowth and levels of glucose and fructose. Although treatmentwith GA₁ resulted in a decrease in sucrose content, it had practicallyno significant effect on the levels of the other sugars. Inthe leaf sheath of d₅, growth is probably not limited by thelevels of these sugars, and GA-induced elongation does not seemto have any significant correlation with the changes in thelevels of sugars. ¹Present address: Department of Botany, Suri Vidyasagar College,Suri, Dt. Birbhum, West Bengal, India. (Received March 18, 1988; Accepted November 21, 1988)     

A lack of correlation between the biological activity and rate of metabolism of ent-(3H)-17-kaurenoic acid by seedlings of dwarf rice cv. Tan-ginbozu

Significant leaf sheath elongation occurred within 24 hr afterapplication of 10 µg (0.67, µCi) of ent-(³H)-17-kaurenoicacid (KA) to individual seedlings of dwarf rice cv. Tan-ginbozu,but this growth was unaccompanied by production of significantlevels of radioactivity in more polar, acidic, ethyl acetate-solublemetabolites of (³H)-KA. However modest levels of radioactivityappeared in the highly water-soluble fraction by hour 24, subsequentto the most rapid phase of KA-induced growth. Growth continuedand by hour 48 was accompanied by the appearance of small amountsof radioactivity in polar, acidic products. It would appearthat KA per se, and not its metabolic products, may be responsiblefor the leaf sheath elongation noted at hour 24. On the speculation that it might be a metabolite of KA, gibberellinA₁₄ (GA₁₄) was applied simultaneously with (³H)-KA to individualrice seedlings. Several changes in the metabolism of ³H-KA inthe presence of GA₁₄ were noted, and GA₁₄ antagonized the KA-inducedsheath elongation. ¹Present address: Botany Department, Rhodes University, Grahamstown,6140, South Africa. ²Present address: Crop Science Department, University of Saskatchewan,Saskatoon, Sask. S7N OWO, Canada. (Received May 12, 1975; )     

Differential Effects of Carbon Dioxide on the Growth Rates of Coleoptiles and the Enclosed Leaves of Hordeum vulgare L. seedlings

Carbon dioxide appears to modulate the growth rates of the barley(Hordeum vulgare L.) coleoptile and the enclosed leaves duringearly germination. Its effect is to stimulate the coleoptileto elongate at a rate equal to or greater than that of the enclosedleaves. Such stimulation was observed directly in barley coleoptilesfloated on CO₂-sarurated water or indirectly using a DCMU solution.This mechanism ensures that the tender leaves remain enclosedwithin the structurally strong coleoptile sheath until the shootpushes through the soil surface. It is suggested that CO₂ respiredby the enclosed leaves effects this modulation: the CO₂ couldpass into the coleoptile chamber and thence into the coleoptilevia stomata present on coleoptile inner walls.     

The Effect of Calmodulin Antagonists on Gibberellic Acid-induced Enzyme Secretion in Barley Aleurone Layers

Calmodulin activity was detected and assayed in barley aleuronecells. The effect of calmodulin antagonists on GA₃-induced enzymesynthesis and secretion in barley aleurone layers was also investigated.These calmodulin antagonists (chlorpromazine, haloperidol) inhibitedonly GA₂-induced -amylase secretion. This inhibitory effectwas intensified after 6 h of GA₃-incubation. This leads us tosuggest that some calmodulin-controlled mechanism is involvedin GA₂-induced -amylase secretion. Hordeum vulgare L., barley aleurone cells, gibberellic acid, -amylase secretion, calmodulin, calmodulin antagonist     

Functional characterisation of Nicotiana tabacum xyloglucan endotransglycosylase (NtXET-1): generation of transgenic tobacco plants and changes in cell wall xyloglucan

To study the function of xyloglucan endotransglycosylase (XET) in vivo we isolated, a tomato (Lycopersicon esculentum Mill.) XET cDNA (GenBank AA824986) from the homologous tobacco (Nicotiana tabacum L.) clone named NtXET-1 (Accession no. D86730). The expression pattern revealed highest levels of NtXET-1 mRNA in organs highly enriched in vascular tissue. The levels of NtXET-1 mRNA decreased in midribs with increasing age of leaves. Increasing leaf age was correlated with an increase in the average molecular weight (MW) of xyloglucan (XG) and a decrease in the relative growth rates of leaves. Transgenic tobacco plants with reduced levels of XET activity were created to further study the biochemical consequences of reduced levels of NtXET-1 expression. In two independent lines, total XET activity could be reduced by 56% and 37%, respectively, in midribs of tobacco plants transformed with an antisense construct. The decreased activity led to an increase in the average MW of XG by at least 20%. These two lines of evidence argue for NtXET-1 being involved in the incorporation of small XG molecules into the cell wall by transglycosylation. Reducing the incorporation of small XG molecules will result in a shift towards a higher average MW. The observed reduction in NtXET-1 expression and increase in the MW of XG in older leaves might be associated with strengthening of cell walls by reduced turnover and hydrolysis of XG. Received: 24 January 2000 / Accepted: 21 July 2000     

Seasonal Changes of GA1, GA19 and Abscisic Acid in Three Rice Cultivars

The levels of endogenous gibberellin A₁ and A₁₉ (GA₁ and GA₁₉)and abscisic acid (ABA) in three rice (Oryza sativa L.) cultivars,Nihonbare (normal), Tan-ginbozu (semid-warf) and Tong-il (semi-dwarf),were measured at various stages of internode elongation andear development. GA₁₉ was the main GA in Nihonbare and Tong-ilthroughout the life cycle but was not detected in Tan-ginbozu.The levels of GA₁ in the ears of all three cultivars were lowand reached their maxima after anthesis. Similarly, the earsof all three cultivars contained high ABA levels which peakedafter anthesis. Shoots contained low quantities of ABA throughoutthe life cycle. The roles of GAs and ABA are discussed withrespect to physiological phenomena, such as internode elongation,ear development and dwarfism. (Received May 9, 1981; Accepted July 18, 1981)     

The role of gibberellins in the growth of floral organs of Pharbitis nil

Evidence that the synthesis of GA₃ is involved in the growthof floral orga'ns of Pharbitis nil is presented. GAs in floralorgans at different developmental stages were surveyed usingTLC followed by the bioassay with two dwarf rice seedlings,‘Tanginbozu’ and ‘Waito-C’. The amountof GAs in the petal and stamen increased rapidly after the petalemerged from calyx, reached a maximum 12 hr before anthesis,then declined markedly thereafter. The GA content in the calyxremained unchanged before and after anthesis, and that in thepistil increased after anthesis. Pharbitis flowers containedat least two active GAs, one of which was probably GA₃, theother appeared to be GA₁₉. GA₃ was detected in relatively largeamounts in both the petal and stamen during their rapid elongation.In the calyx, which showed little increase in fresh weight duringrapid flower growth, GA₉ was the dominant GA. Exogenously suppliedGA₃ promoted elongation of sections in excised young filaments.Sucrose was necessary for definite growth promotion by GA₃.GA₁₉ had little effect on filament elongation, and IAA was ratherinhibitive. (Received July 29, 1972; )     

An Analysis of the Effects of Gibberellic Acid on Tomato Leaf Growth

The effects of four consecutive daily sprays of gibberellicacid (GA₃) on the growth of leaves of young tomato plants cv.Potentate were studied. Total leaf weight and area were increasedby GA₃. The percentage changes were larger in the younger leavesthan in the older but the absolute increases of the middlleleaves accounted for most of the total response. Chlorophyllcontent, both total and per unit weight, was reduced by GA inthe older leaves and increased in the younger; on an area basisit was reduced in all but the youngest leaves. Palisade cell length and palisade cell number per unit sectionlength were reduced by GA₃ in the oldest leaves and increasedin the youngest. There were larger intercellular spaces in bothmesophyll layers and a larger transectional area of the mid-ribsof the oldest and two youngest leaves in GA₃ plants. The ‘surfaceareas’ of epidermal cells were also increased by GA₃ treatment.Leaf fresh weight per unit area was only a true index of laminathickness in the two oldest leaves.