The regulation of leaf elongation and xyloglucan endotransglycosylase by gibberellin in 'Himalaya' barley (Hordeum vulgare L.)

The potential role of xyloglucan endotransglycosylase (XET)in GA-stimulated cell elongation was investigated during leafexpansion in barley (Hordeum vulgare L.). XET activity in aqueousextracts of leaves was detected in all segments along the elongatingblade of leaf 1 of seedlings, but was at highest levels in basalsegments. Leaf 1 elongation rates of gibberellin (GA)-responsivedwarf mutants were lower than the wild type, and accompaniedby reduced levels of XET activity. Leaf elongation rates ofthe dwarfs increased following treatment with gibberellic acid(GA₃) associated with higher levels of XET activity. The slendermutant, crossed into a dwarfing background, exhibited high ratesof leaf 1 elongation and high levels of XET activity withoutadded GA₃. The elongation of leaf 3 in a GA-responsive dwarfmutant was also studied. Treatment with GA₃ resulted in bladeand sheath lengths being 5-fold and 7-fold (respectively) thelengths of controls, and again there were increases in bladeand sheath XET activities. To investigate the basis for changesin XET activity levels two XET-related cDNA clones were isolated.RNAs detected by the two clones occurred at the highest levelsin basal segments of rapidly elongating leaves, but they haddifferent distribution patterns along the leaf. Overall, thedata indicate that an XET-like activity is detectable in barleyleaves, that the activity level and related. Key words: Gibberellin (GA), leaf elongation, Hordeum vulgare, 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.     

Drought effects on cellular and spatial parameters of leaf growth in tall fescue

The effect of drought and recovery on cellular and spatial parametersof the growth process in tall fescue leaves was studied in twoexperiments. In both experiments plants grown on vermiculiteand maintained in a controlled environment were submitted toa 7 d drought period generated by withholding water. Droughtwas followed by a 3 d recovery period in experiment II. As leafelongation rate (LER) decreased during developing drought boththe growth zone length (initially 40 mm) and the maximum relativeelemental growth rate (initially 0.09 mm mm^–1 h^–1during the dark period of diurnal cycles) within the growthzone declined. But the growth zone still exhibited a lengthof approximately 15 mm when LER approached 0 under severe drought(–2.0 MPa predawn leaf water potential). The growth potentialof the basal 15-mm-long portion of the leaf was conserved duringthe period when drought effected the complete arrest of leafelongation. A (retrospective) analysis of the position-timerelationships of epidermal cells identified on leaf replicas(experiment II) indicated that the cell flux out of the growthzone responded very sensitively to drought. Before drought theflux was maximum at approximately 3.2 cells (cell file h)^–1during the dark period. Flux decreased to 0 when leaf elongationstopped. Flux also varied diurnally both under well-wateredand droughted conditions. In well-watered conditions it wasabout 30% less during the light than the dark period. Cell elongationwas also sensitive to drought. Under well-watered conditionsepidermal cell elongation stopped when cells attained a lengthof approximately 480 µm. During developing drought cellsstopped elongating at progressively shorter lengths. When LERhad decreased to almost nil, cells stopped elongating at a lengthof approximately 250 µn. When drought was relieved followinga 2 d complete arrest of leaf elongation then cells shorterthan 250 µm were able to resume expansion. Following rewateringcell flux out of the growth zone increased rapidly to and abovethe pre-drought level, but there was only a slow increase overtime in the length at which cell elongation stopped. About 2d elapsed until the leaf growth zone produced cells of similarlength as before drought (i.e. approximately 480 µm). Key words: Epidermal cell length, cell flux, (leaf) growth zone, leaf elongation rate, relative elemental growth rate, position-time relationships (path line, growth trajectory), drought, water deficit     

Kinetics of Maize Leaf Elongation: I. INCREASED YIELD THRESHOLD LIMITS SHORT-TERM, STEADY-STATE ELONGATION RATES AFTER EXPOSURE TO SALINITY

The short-term responses of leaf elongation to salinity areinvestigated in this study. The kinetics of maize (Zea maysL.) leaf elongation were measured with Linear Variable DifferentialTransformers (LVDTs). After exposure to salinity (0 to 120 molm^–3 NaCl), leaf elongation rates (LER) declined rapidly.Within 4 h, LER had recovered and reached a new steady-statefor all salinity treatments. These rates were reduced by 10,20, and 60% of control rates by 40, 80 and 120 mol m^–3NaCl, respectively. Osmotic adjustment in the growing zone ofleaves was correlated with the recovery of LER after plant exposureto salinity. However, after 4 h of exposure, the osmolalityof the cell sap continued to increase without effect on steady-stateLER. Estimates of the apparent turgor in the growing zone indicatedthat turgor was no longer limiting LER of salt-stressed plantsafter 4 h. An in vivo technique was developed to apply a unidirectionalforce to intact growing leaves of maize to mimic increases inelongation force. Relative elongation rate (RER) were increasedby adding weights to the LVDT core to increase elongation force.Plots of RER as a function of elongation force gave estimatesof two growth coefficients: the yield threshold and the yieldingcoefficient, mL/(m + L), where m is the cell wall extensibilityand L is the hydraulic conductivity. RER as a function of elongationforce was determined immediately, 05, 4, and 21 h after plantswere salinized. Estimates of the growth coefficients indicatedthat the apparent yield threshold decreased immediately aftersalinization. However, when LER reached steady-state, the yieldthreshold of salt-stressed plants had increased above controlvalues and was the only limiting growth coefficient. There wereno significant effects of salinity on the yielding coefficients,cell wall extensibility or hydraulic conductivity. One of theadvantages of this in vivo technique over other methods is thatyield threshold, yielding coefficient, and cell wall extensibilitycan be determined without the confounding effects of woundingor osmotic stress. This technique may prove widely applicableto the study of other growth regulating factors. Key words: Salinity, leaf growth, Zea mays L     

Kinetics of Maize Leaf Elongation: II. RESPONSES OF A NA-EXCLUDING CULTIVAR AND A NA-INCLUDING CULTIVAR TO VARYING NA/CA SALINITIES

Salinity causes physiological and morphological changes in plantsand calcium can mitigate many of these effects. In this study,the effects of salinity (75 mol m^–3 NaCl) with or withoutsupplemental Ca (10 mol m^–3) on the kinetics of maize(Zea mays L.) leaf elongation were examined using Linear VariableDifferential Transformers (LVDTs). Short-term growth responsesof two cultivars (Dekalb hybrid XL75 and Pioneer hybrid 3906)differing in salt tolerance were compared. Salinity caused animmediate reduction in the leaf elongation rate (LER). Within2 h, elongation rates had increased and reached new steady rates.Significant differences between salinity treatments with highand low Ca could be detected within the first 2 h after impositionof salinity for Dekalb hybrid XL75, but not for Pioneer hybrid3906. After 24 h, distinct differences for both cultivars weredetected. Dekalb hybrid XL75, a Na-includer, was more salt-sensitiveand responsive to supplemental Ca (10 mol m^–3) than Pioneerhybrid 3906, a Na-excluder. Turgor was not reduced 24 h aftersalinization because there was complete osmotic adjustment inthe elongation zone of the leaves. Analysis of the growth parameterslimiting LER indicated that the yield threshold (Y) was increasedfor salt-stressed plants. In addition, both the cell wall extensibilityand hydraulic conductance were reduced 24 h after salinization.Supplemental Ca increased LER of salt-stressed plants by increasinghydraulic conductance. The differences in LER of the two cultivarsunder saline conditions was attributed to differences in theincrease of Y caused by salinity. Key words: Calcium, growth, salinity, sodium, Zea mays L.     

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     

Leaf growth responses to ABA are temperature dependent

The robustness of a leaf elongation bioassay was evaluated byconducting trials with detached shoots of wheat at several differenttemperatures. Leaf elongation rate (LER) was monitored for shootsfed either an artificial xylem solution or xylem solution plus10^–3mol m^–3 abscisic acid (ABA). Consistent resultswere obtained when periodic ruler measurements of many shootswere made and compared with simultaneous measurements on a singleshoot made with a linearly variable displacement transducer(LVDT). ABA treatment consistently inhibited leaf growth; however,the magnitude of the inhibition was dependent on the temperatureat which the assay was conducted. Interpretation of resultsfrom such bioas-says in terms of ABA concentration suppliedto the detached shoots is complicated by this observation sincethere is no unique relationship between leaf growth inhibitionand ABA concentration. The results are discussed in terms ofchemical signalling affecting the growth rate of plants in dryingsoil. Key words: ABA, leaf growth, temperature, leaf elongation bioassay     

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     

Quantitative analysis of the combined effects of temperature, evaporative demand and light on leaf elongation rate in well-watered field and laboratory-grown maize plants

The respective effects of meristem temperature, vapour pressuredeficit (VPD) and photosynthetic photon flux density (PPFD)on leaf elongation rate (LER) of maize, in the absence of waterdeficit in the soil have been quantified. This analysis wascarried out in a series of field experiments in northern andsouthern France over several seasons and years, and in growthchamber experiments. LER was measured with 10 min steps, togetherwith meristem temperature, VPD and PPFD at leaf level in threetypes of experiments: in growth chamber experiments with stepsin PPFD or VPD at constant meristem temperature, in growth chamberexperiments with several combinations of constant, but contrasting,PPFDs, VPDs and meristem temperatures, and in the field withfluctuating conditions, (i) When evaporative demand was low(night or day with low air VPD), LER was only linked to meristemtemperature, regardless of other climatic conditions, (ii) Lighthad no effect per se on LER in the range from 0 to 1500 molm^–2 s^–1 for time-scales longer than 2 h, providedthat its indirect effects on meristem temperature and on evaporativedemand were corrected (in the growth chamber) or taken intoaccount (in the field), and provided that cumulated PPFD overa weekly time-scale was compatible with field conditions, (iii)Evaporative demand sensed by growing leaves, as estimated bymeristem-to-air vapour pressure difference, markedly affectedLER in the range from 1–4 kPa, at all time-scales understudy, with a unique relationship in the growth chamber (constantconditions) and in the field (fluctuating conditions). Thiseffect was only observed when PPFD was high enough for stomatato open. The negative effect of evaporative demand on LER wasprobably not due to long distance root-to-shoot signalling,since soil was wet, calculated root water potential remainedclose to 0 MPa and concentration of ABA in the xylem sap wasvery low. Therefore, it is proposed to model maize LER witha two-step process, involving the calculation of the maximumLER at a given meristem temperature and then the calculationof the reduction in LER due to evaporative demand. Joint analysisof the whole set of data by using the two equations yieldeda r² of 0.75. This two-step process would be more accurate thanthe provision of LER from temperature only in cases where airVPD frequently exceeds 2 kPa. Key words: Leaf growth, light, evaporative demand, temperature, thermal time, water deficit, ABA, Zea mays L.     

Nitrate Accumulation and its Relation to Leaf Elongation in Spinach Leaves

The leaf elongation rate (LER) of spinach leaves during theday was twice that during the night when grown at a photon fluxdensity of 145 µmol m^–2 s^–1. All leaves showedthe same LER-pattern over 24 h. Due to low turgor, LER was lowin the afternoon and in the first hours of the night until wateruptake restored full turgor. Osmotic potential remained constantdue to increased nitrate uptake and starch degradation in thisperiod. LER increased to high rates in the second part of thenight and in the morning. The lower rate in the dark comparedto the light was not caused by the lower night temperatures,as increased photon flux density during growth resulted in equalrates in the light and the dark. Increased relative humiditydecreased LER and afternoon rates were most sensitive to waterstress. A ‘low light’ night period did not changeLER-pattern during the night or on the following day. We concludethat nitrate is not an obligatory osmoticum during the nightand can be exchanged for organic osmotica without decreasingLER. During the night the turgor is first restored by increasingwater uptake, nitrate uptake and starch degradation. This resultedin increased leaf fresh weight in this period. Thereafter, elongationincreased by simultaneous uptake of nitrate and water. Nitrateconcentration was, therefore, constant in the older leaves.In the younger leaves nitrate concentration increased to replacesoluble carbohydrates. The vacuoles of the old leaves were filledwith nitrate before those of the young leaves. Key words: Spinacia oleracea L., nitrate accumulation, osmotic potential, organic acids     

Root Growth Maintenance at Low Water Potentials (Increased Activity of Xyloglucan Endotransglycosylase and Its Possible Regulation by Abscisic Acid)

Previous work suggested that an increase in cell wall-loosening contributes to the maintenance of maize (Zea mays L.) primary root elongation at low water potentials ([psi]w). It was also shown that root elongation at low [psi]w requires increased levels of abscisic acid (ABA). In this study we investigated the effects of low [psi]w and ABA status on xyloglucan endotransglycosylase (XET) activity in the root elongation zone. XET is believed to contribute to wall-loosening by reversibly cleaving xyloglucan molecules that tether cellulose microfibrils. The activity of XET per unit fresh weight in the apical 10 mm (encompassing the elongation zone) was constant at high [psi]w but increased by more than 2-fold at a [psi]w of -1.6 MPa. Treatment with fluridone to decrease ABA accumulation greatly delayed the increase in activity at low [psi]w. This effect was largely overcome when internal ABA levels were restored by exogenous application. Spatial distribution studies showed that XET activity was increased in the apical 6 mm at low [psi]w whether expressed per unit fresh weight, total soluble protein, or cell wall dry weight, corresponding to the region of continued elongation. Treatment with fluridone progressively inhibited the increase in activity with distance from the apex, correlating with the pattern of inhibition of elongation. Added ABA partly restored activity at all positions. The increase in XET activity at low [psi]w was due to maintenance of the rate of deposition of activity despite decreased deposition of wall material. The loss of activity associated with decreased ABA was due to inhibition of the deposition of activity. The results demonstrate that increased XET activity is associated with maintenance of root elongation at low [psi]w and that this response requires increased ABA.     

In vivo colocalization of xyloglucan endotransglycosylase activity and its donor substrate in the elongation zone of Arabidopsis roots

We have developed a method for the colocalization of xyloglucan endotransglycosylase (XET) activity and the donor substrates to which it has access in situ and in vivo. Sulforhodamine conjugates of xyloglucan oligosaccharides (XGO-SRs), infiltrated into the tissue, act as acceptor substrate for the enzyme; endogenous xyloglucan acts as donor substrate. Incorporation of the XGO-SRs into polymeric products in the cell wall yields an orange fluorescence indicative of the simultaneous colocalization, in the same compartment, of active XET and donor xyloglucan chains. The method is specific for XET, as shown by competition experiments with nonfluorescent acceptor oligosaccharides, by negligible reaction with cello-oligosaccharide-SR conjugates that are not XET acceptor substrates, by heat lability, and by pH optimum. Thin-layer chromatographic analysis of remaining unincorporated XGO-SRs showed that these substrates are not extensively hydrolyzed during the assays. A characteristic distribution pattern was found in Arabidopsis and tobacco roots: in both species, fluorescence was most prominent in the cell elongation zone of the root. Proposed roles of XET that include cell wall loosening and integration of newly synthesized xyloglucans could thus be supported.     

Xyloglucan Endotransglycosylase Activity, Microfibril Orientation and the Profiles of Cell Wall Properties Along Growing Regions of Maize Roots

A series of physical and chemical analyses were made on theexpanding zone of maize seedling roots grown in hydroponics.Comparison of longitudinal profiles of local relative elementalgrowth rate and turgor pressure indicated that cell walls becomelooser in the apical 5 mm and then tighten 5–10 mm fromthe root tip. Immersion of roots in 200 mol m^–3 mannitol(an osmotic stress of 0·48 MPa) rapidly and evenly reducedturgor pressure along the whole growing region. Growth was reducedto a greater extent in the region 5–10 mm from the roottip than in the apical region. This indicated rapid wall-looseningin the root tip, but not in the more basal regions. Following 24 h immersion in 400 mol m^–3 mannitol (an osmoticstress of 0·96 MPa) turgor had recovered to pre-stressedvalues. Under this stress treatment, growth was reduced in theregion 4–10 mm from the root tip, despite the recoveryof turgor, indicating a tightening of the wall. In the rootapex, local relative elemental growth rate was unchanged incomparison to control tissue, showing that wall properties herewere similar to the control values. Cellulose microfibrils on the inner face of cortical cell wallsbecame increasingly more parallel to the root axis along thegrowth profile of both unstressed and stressed roots. Orientationdid not correlate with the wall loosening in the apical regionof unstressed roots, or with the tightening in the region 5–10mm from the root tip following 24 h of osmotic stress. Longitudinal profiles of the possible wall-loosening enzymexyloglucan endotransglycosylase (XET) had good correspondencewith an increase in wall loosening during development. In thezone of wall tightening following osmotic stress, XET activitywas decreased per unit dry weight (compared with the unstressedcontrol), but not per unit fresh weight. Key words: Osmotic stress, turgor, growth, cell wall properties, microfibrils, XET     

The relationship between xyloglucan endotransglycosylase and in-vitro cell wall extension in cucumber hypocotyls

It has been proposed that cell wall loosening during plant cell growth may be mediated by the endotransglycosylation of load-bearing polymers, specifically of xyloglucans, within the cell wall. A xyloglucan endotransglycosylase (XET) with such activity has recently been identified in several plant species. Two cell wall proteins capable of inducing the extension of plant cell walls have also recently been identified in cucumber hypocotyls. In this report we examine three questions: (1) Does XET induce the extension of isolated cell walls? (2) Do the extension-inducing proteins possess XET activity? (3) Is the activity of the extension-inducing proteins modulated by a xyloglucan nonasaccharide (Glc₄-Xyl₃-Gal₂)? We found that the soluble proteins from growing cucumber (cucumis sativum L.) hypocotyls contained high XET activity but did not induce wall extension. Highly purified wall-protein fractions from the same tissue had high extension-inducing activity but little or no XET activity. The XET activity was higher at pH 5.5 than at pH 4.5, while extension activity showed the opposite sensitivity to pH. Reconstituted wall extension was unaffected by the presence of a xyloglucan nonasaccharide (Glc₄-Xyl₃-Gal₂), an oligosaccharide previously shown to accelerate growth in pea stems and hypothesized to facilitate growth through an effect on XET-induced cell wall loosening. We conclude that XET activity alone is neither sufficient nor necessary for extension of isolated walls from cucumber hypocotyls.     

ERRATA

p. 1284. Fig. 1, values on the ordinate (Leaf extension rate,mm h^–1): scale should be moved down so that zero lieson the x-axis ... LER = 3.2 {1 – exp [–4 (S – 0.41)]}...     

Kinetics of maize leaf elongation IV. Effects of (+)- and (-)-abscisic acid

Abscisic acid (ABA) is involved in many of the responses of plants to environmental stress. This study focuses on the inhibitory effect of ABA on leaf expansion. In addition, the effects of (+)-ABA, the natural form of ABA, were compared to the effects of (-)-ABA. Leaf elongation rates (LER) were measured for the 3rd leaf of maize plants. ABA concentrations were measured by RIA for total ABA and an ELISA specific for (+)-ABA. ABA was added to the hydroponic solution and changes in the LER were measured over time. ABA could inhibit LER within 30 min ad reached steady-state LER within 4 h. Internal ABA concentrations in the growing zone of the leaf also reached steady-state concentrations after 4 h. This effect of ABA was reversible, because LER was fully restored upon removal of externally applied ABA, and internal concentrations of ABA in the growing zone returned to normal levels, whereas ABA concentrations remained elevated in mature tissue. Thus, steady-state LER was highly correlated with the steady-state internal ABA concentration of the growing zone. ABA inhibited leaf expansion by increasing the apparent cell wall yield threshold; no other growth parameters were affected. The (-)-enantiomer of ABA had much less effect on LER than (+)-ABA when compared upon an external concentration basis. Internal ABA concentrations rationalized the response, showing that (-)-ABA accumulation was very low, most likely due to low uptake rates. From this analysis, it was determined that LER was equally sensitive to internal concentrations of (+)- or (-)-ABA.     

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.     

The Growth Rate of Wheat Leaves in Relation to the Extension Zone Sugar Concentration Manipulated by Shading

The main objective of this study was to determine the relationshipbetween the relative rate of growth of emerging wheat leavesand the hexose sugar concentration of the extension zone. Shortperiods of intense shading (to 20 or 5% of full sun for up to14 d) were used to decrease hexose concentrations. Shading decreased hexose concentrations to a fraction of thatof controls and also resulted in thin and narrow leaves thatwere less in dry weight than control leaves of the same length.Shading did however increase the length of the zone of extendingtissue at the leaf base by 30%. The effect of hexose concentrations on the relative rate ofleaf growth was evaluated by determining the ratio between growthrates of shaded and control leaves. This ratio declined as hexoseconcentrations declined and the relationship was described bya rectangular hyperbola (r > 0.95, P < 0.01). Combineddata from many leaves on the main shoot and its tillers fromtwo irrigated wheat crops all conformed to the same relationship.The hexose concentrations where the ratio of growth rates washalf the maximum rate were 0.42 mg g^–1 fr. wt. for extensiongrowth and 1.74 mg g^–1 fr. wt. for dry weight growth.These values were significantly (P < 0.01) different. These results were compared with data from emerging leaves offield crops and it was concluded that hexose concentrationshad not limited leaf growth rates, the lowest values recordedbeing 2.5–3.0 mg g^–1 fr. wt. It was further suggestedas unlikely that leaf growth rates of wheat crops in the fieldwould be limited by hexose concentrations.     

Analysis of a xyloglucan endotransglycosylase/hydrolase (XTH) from the lycopodiophyte Selaginella kraussiana suggests that XTH sequence characteristics and function are highly conserved during the evolution of vascular plants

A tissue print followed by a xyloglucan endotransglycosylase assay revealed that XET activity is present at sites of cell elongation in both roots and shoots of the lycopodiophyte Selaginella kraussiana. This paper provides the first report and analysis of a xyloglucan endotransglycosylase/hydrolase (XTH) cDNA sequence, isolated from a club moss. In silico analysis of the deduced amino acid sequence revealed a strong conservation of the XET-domain described in higher plants. The catalytic site (DEIDLEFLG) varies in only one amino acid compared with the consensus sequence and was shown to be functional after recombinant expression of Sk-XTH1 in Pichia pastoris. Sk-XTH1 displays xyloglucan endotransglycosylase activity over a broad pH (4.5-7.5) and temperature range (4-30 degrees C), but it shows no hydrolase activity. The catalytic site is followed by a consensus sequence for N-linked glycosylation. Four terminal cysteines were shown to stabilize a putative XET-C terminal extension region, which includes conserved amino acids, involved in the recognition and binding of the substrates. The N-linked sugar interactions as well as the disulphide bridges were shown to be necessary to perform XET activity. The presence of a highly conserved XTH sequence and function in a microphyllophyte suggests that XTHs were present before the divergence of lycopodiophytes and euphyllophytes. It also points to a possible key role for XTHs in the production of a cell wall that allowed the further evolution of land plants.