Kinetics of Aluminum Uptake in Triticum aestivum L: Identity of the Linear Phase of Aluminum Uptake by Excised Roots of Aluminum-Tolerant and Aluminum-Sensitive Cultivars

The identity of a linear phase of aluminum (Al) uptake in Triticum aestivum was investigated by analysis of the kinetics of Al uptake by excised roots and purified cell wall fractions. Classical interpretation of kinetic data suggests that a linear phase of uptake with time reflects uptake across the plasma membrane; however, in studies with Al the possibility that the linear phase of uptake includes accumulation of Al in both the symplasm and the apoplasm has not been discounted. In our experiments, we observed a linear phase of Al uptake at both ambient and low temperatures, although the rate of uptake at 0°C was 53 to 72% less than at 23°C, depending on cultivars. This nonsaturable phase of uptake at low temperature suggests that a portion of the linear phase of Al uptake is nonmetabolic. Furthermore, analysis of Al in cell wall fractions isolated from excised roots pretreated with Al suggests that the linear phase of uptake includes a cell wall component. When excised roots were pretreated with Al, accumulation of Al in purified cell wall material included a linear phase that could not be desorbed with a 30 minute wash in citrate. The rates of linear-phase accumulation of Al by cell wall material and cell contents were similar. In contrast, the linear phase of in vitro uptake of Al by purified cell wall material was completely desorbed by a 30 minute wash with citrate. These results suggest that the linear phase of Al uptake observed in excised roots of T. aestivum included metabolism-dependent binding of Al in apoplasm.     

Aluminum Partitioning in Intact Roots of Aluminum-Tolerant and Aluminum-Sensitive Wheat (Triticum aestivum L.) Cultivars

Aluminum (Al) partitioning in intact roots of wheat (Triticum aestivum L.) cultivars that differ in sensitivity to Al was investigated. Roots of intact seedlings were exposed to Al for up to 24 hours and distribution of Al was assessed visually by hematoxylin staining or by direct measurement of concentration of Al by atomic absorption spectrophotometry or ion chromatography. Major differences in Al accumulation between Al-tolerant (Atlas 66) and Al-sensitive (Tam 105) cultivars were found in the growing regions 0 to 2 and 2 to 5 millimeters from the root apex. Al content was 9 to 13 times greater in the 0 to 2 millimeters root tips of cv Tam 105 than in the tips of cv Atlas 66 when exposed to 50 micromolar Al for 19 to 24 hours. The oxidative phosphorylation inhibitor carbonyl cyanide m-chlorophenylhydrazone and the protein synthesis inhibitor cycloheximide increased Al uptake by intact root tips of cv Atlas 66. Also, loss of Al from the roots of both cultivars was measured after the roots were “pulsed” with 50 micromolar Al for 2 hours and then placed in an Al-free nutrient solution for 6 hours. The 0 to 2 millimeter root tips of cv Tam 105 lost 30% of the absorbed Al, whereas the tips of cv Atlas 66 lost 60%. In light of these results, we conclude that the differential Al sensitivity in wheat correlates with the concentration of Al in the root meristems. The data support the hypothesis that part of the mechanism for Al tolerance in wheat is based on a metabolism-dependent exclusion of Al from the sensitive meristems.     

Differential Exudation of Polypeptides by Roots of Aluminum-Resistant and Aluminum-Sensitive Cultivars of Triticum aestivum L. in Response to Aluminum Stress

Cultivars of Triticum aestivum differing in resistance to Al were grown under aseptic conditions in the presence and absence of Al and polypeptides present in root exudates were collected, concentrated, and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Upon exposure to 100 and 200 [mu]M Al, root elongation in Al-sensitive cultivars was reduced by 30 and 65%, respectively, whereas root elongation in resistant cultivars was reduced by only 15 and 30%. Accumulation of polypeptides in the growth medium increased with time for 96 to 120 h, with little additional accumulation thereafter. This pattern of exudation was virtually unaffected by exposure to 100 [mu]M Al in the Al-resistant cultivars Atlas 66 and Maringa, whereas total accumulation was reduced in sensitive cultivars. Changes in exudation were consistent with alterations in root elongation. Al-induced or Al-enhanced polypeptide bands were detected in Atlas 66 and Maringa after 72 h of exposure to Al. Increased accumulation of 12-, 22-, and 33-kD bands was observed at 75 [mu]M Al in Atlas 66 and 12-, 23-, and 43.5-kD bands started to appear at 50 [mu]M Al in Maringa. In the Al-sensitive cultivars Roblin and Katepwa, no significant effect on polypeptide profiles was observed at values up to 100 [mu]M Al. When root exudates were separated by ultrafiltration and the Al content was measured in both high molecular mass (HMM; >10 kD) and ultrafiltrate (<10 kD) fractions, approximately 2 times more Al was detected in HMM fractions from Al-resistant cultivars than from Al-sensitive cultivars. Dialysis of HMM fractions against water did not release this bound Al;digestion with protease released between 62 and 73% of total Al, with twice as much released from exudates of Al-resistant than of Al-sensitive cultivars. When plants were grown in the presence of 0 to 200 [mu]M Al, saturation of the Al-binding capacity of HMM exudates occurred at 50 [mu]M Al in Al-sensitive cultivars. Saturation was not achieved in resistant cultivars. Differences in exudation of total polypeptides in response to Al stress, enhanced accumulation of specific polypeptides, and the greater association of Al with HMM fractions from Al-resistant cultivars suggest that root exudate polypeptides may play a role in plant response to Al.     

Aluminum Effects on Calcium (45Ca2+) Translocation in Aluminum-Tolerant and Aluminum-Sensitive Wheat (Triticum aestivum L.) Cultivars (Differential Responses of the Root Apex versus Mature Root Regions)

The influence of Al exposure on long-distance Ca2+ translocation from specific root zones (root apex or mature root) to the shoot was studied in intact seedlings of winter wheat (Triticum aestivum L.) cultivars (Al-tolerant Atlas 66 and Al-sensitive Scout 66). Seedlings were grown in 100 [mu]M CaCl2 solution (pH 4.5) for 3 d. Subsequently, a divided chamber technique using 45Ca2+-labeled solutions (100 [mu]M CaCl2 with or without 5 or 20 [mu]M AlCl3, pH 4.5) was used to study Ca2+ translocation from either the terminal 5 to 10 mm of the root or a 10-mm region of intact root approximately 50 mm behind the root apex. The Al concentrations used, which were toxic to Scout 66, caused a significant inhibition of Ca2+ translocation from the apical region of Scout 66 roots. The same Al exposures had a much smaller effect on root apical Ca2+ translocation in Atlas 66. When a 10-mm region of the mature root was exposed to 45Ca2+, smaller genotypic differences in the Al effects effects on Ca2+ translocation were observed, because the degree of Al-induced inhibition of Ca2+ translocation was less than that at the root apex. Exposure of the root apex to Al inhibited root elongation by 70 to 99% in Scout 66 but had a lesser effect (less than 40% inhibition) in Atlas 66. When a mature root region was exposed to Al, root elongation was not significantly affected in either cultivar. These results demonstrate that genotypic differences in Al-induced inhibition of Ca2+ translocation and root growth are localized primarily in the root apex. The pattern of Ca2+ translocation within the intact root was mainly basipetal, with most of the absorbed Ca2+ translocated toward the shoot. A small amount of acropetal Ca2+ translocation from the mature root regions to the apex was also observed, which accounted for less than 5% of the total Ca2+ translocation within the entire root. Because Ca2+ translocation toward the root apex is limited, most of the Ca2+ needed for normal cellular function in the apex must be absorbed from the external solution. Thus, continuous Al disruption of Ca2+ absorption into cells of the root apex could alter Ca2+ nutrition and homeostasis in these cells and could play a pivotal role in the mechanisms of Al toxicity in Al-sensitive wheat cultivars.     

Aluminum Toxicity in Roots : Correlation among Ionic Currents, Ion Fluxes, and Root Elongation in Aluminum-Sensitive and Aluminum-Tolerant Wheat Cultivars

The inhibition of root growth by aluminum (Al) is well established, yet a unifying mechanism for Al toxicity remains unclear. The association between cell growth and endogenously generated ionic currents measured in many different systems, including plant roots, suggests that these currents may be directing growth. A vibrating voltage microelectrode system was used to measure the net ionic currents at the apex of wheat (Triticum aestivum L.) roots from Al-tolerant and Al-sensitive cultivars. We examined the relationship between these currents and Al-induced inhibition of root growth. In the Al-sensitive cultivar, Scout 66, 10 micromolar Al (pH 4.5) began to inhibit the net current and root elongation within 1 to 3 hours. These changes occurred concurrently in 75% of experiments. A significant correlation was found between current magnitude and the rate of root growth when data were pooled. No changes in either current magnitude or growth rate were observed in similar experiments using the Al-tolerant cultivar Atlas 66. Measurements with ion-selective microelectrodes suggested that H⁺ influx was responsible for most of the current at the apex, with smaller contributions from Ca²⁺ and Cl⁻ fluxes. In 50% of experiments, Al began to inhibit the net H⁺ influx in Scott 66 roots at the same time that growth was affected. However, in more than 25% of cases, Al-induced inhibition of growth rate occurred before any sustained decrease in the current or H⁺ flux. Although showing a correlation between growth and current or H⁺ fluxes, these data do not suggest a mechanistic association between these processes. We conclude that the inhibition of root growth by Al is not caused by the reduction in current or H⁺ influx at the root apex.     

Boron Uptake by Excised Barley Roots

Active uptake of boron (B) by excised barley roots is linear with time for at least 1.5 h. Although no evidence was found for accumulation of B against a concentration gradient. this component of B uptake does satisfy other criteria for an active transport process. Transport is inhibited by 0.05 mM 2,4-dinitrophenol, 0.05 mM azide, 5 mM arsenate and 5 mM dicoumarol. Also, uptake is temperature-sensitive, being nil at 2°C and maximal at 34 to 38°C. Boron uptake by barley roots increases with time when they are washed in aerated 0.5 mM CaSO₄ solution. A double reciprocal plot of the B uptake data manifests a series of phases separated by sharp transitions or “jumps”, and is compatible with the concept of multiphasic uptake mechanisms. Kinetic constants and transition points for the various phases were calculated accordingly. The fit of these data was compared statistically to three other relevant models, viz, the dual model, the “single + diffusion” model (a Michaelis–Menten term and a diffusion term), and the negative cooperativity model. In each case, the data were better represented by the multiphasic model.     

Varietal Differences in the Kinetics of Iron Uptake by Excised Rice Roots

Iron uptake by excised rice roots can be described in kineticterms of active absorption and the formation of ion-carriercomplexes. Conventional interpretation indicates two carriers,one of which is responsible for iron absorption at low concentrations,with the second primarily functioning at high iron levels. Themaximum absorption of both carriers vanes greatly between varieties,the variety Pebifun having much greater absorptive capacitythan either Siam-29 or Paldal. Varietal differences in uptakecapacity primarily depend on total carrier concentration, 0.93,0.40,and 0.31 µ moles per g roots of Pebifun, Siam-29, andPaldal respectively. Although the kinetic treatment suggeststwo carriers, the chromatographic evidence indicates the presenceof the iron primarily in one major initial complex, with suggestionsof accumulation rather than turnover. An alternative to thecarrier hypothesis is therefore put forward which regards theaccumulation of iron as binding initially on to one or severalclosely related substances, followed by incorporation into secondaryproducts when a critical level of the initial product is exceeded.Manganese was found to competitively inhibit iron uptake byrice roots. Inhibition was more severe in Siam-29 than in Paldaland can be explained on the basis of carrier concentration andcompetition for the same carrier site. Copper was found notto be competitive for the iron absorption sites. The effectof copper was not significant with the low levels of iron butonly with the 5 x 10^–4 M Fe level. The effect varied betweenvarieties, in Pebifun both 5 x 10^–5 and 5 x 10^–6M Cu inhibited iron uptake compared with the control. In thecase of Siam-29 both copper levels accelerated iron uptake,5 x 10^–5 M Cu giving the greatest uptake. With 5 x 10^–5M Cu Pebifun took up much less iron than Siam-29. The mecharosmof copper inhibition/stimulation is highly speculative.     

Manganese Uptake by Excised Oat Roots

Uptake of ⁵⁴Mn by excised oat roots from dilute manganese chloridesolutions has been investigated. The time-course of uptake hasbeen analysed into the customary but somewhat arbitrary fastand slow phases. Uptake is not metabolic in either of these.The fast phase (‘exchangeable’ manganese) is essentiallycomplete in about 30 minutes and represents the attainment ofequilibrium in a process of ion-exchange. It is shown that analysesappropriate for enzyme kinetics cannot be applied in this situation,and an alternative formulation is based on Donnan equilibration,taking account of the selectivity of the ion exchanger towardsdifferent counter-ions; the predictions of this latter theoryare compared with the experimentally determined uptake. Theslow phase (‘absorbed’ manganese) may also involveexchange sites, either chemically different from, or more difficultof access than, those involved in the fast phase, or both. Equilibriumwas certainly not reached in three hours in this slow-phaseprocess. Release of manganese, taken up by the roots from manganese chloridesolutions, into calcium chloride solutions does not seem tobe simply the reversal of uptake, particularly with very dilutesolutions. This is particularly shown by the kinetics of uptakeand release, uptake being a much faster process than release.Manganese may transfer from the first phase to the second phase,but there is no evidence that uptake by the roots proceeds inseries from first to second phase. It is considered more likelythat the two phases function independently, linked by the surroundingsolution.     

Uptake of Iodide by Excised Bean Roots

Mesophyll cells of leaf slices of bean (Phaseolus vulgaris L.) absorb six to ten times more K⁺ than Rb⁺ from 0.1 mM single chlorides of these cations. Absorption of ⁴²K⁺ from 0.1 mM⁴²KCl is much more inhibited by low concentrations of Rb₂SO₄ than by K₂SO₄. The isotherm for K⁺ absorption is biphasic in the range 0.1–1.1 mM, and K⁺ is more effective than Rb⁺ in causing transition from phase 1 to phase 2.     

Mechanisms of Sodium and Rubidium Uptake by Excised Barley Roots

Accumulation of sodium and rubidium by excised barley roots was investigated. The concentration isotherm yielded one absorption shoulder. Nevertheless, it is suggested that two mechanisms take part in the uptake of sodium and rubidium: One non-metabolic mechanism with an apparent participation at low external salt concentrations (< 1 mM) and at high concentrations (> 20 mM). Such a mechanism is almost unaffected by low temperature conditions and by metabolic inhibitors. Rubidium possesses a high affinity toward this non-metabolic system. The second mechanism is sensitive to metabolic inhibitors and to low temperature conditions. It dominates at intermediate external concentrations (1–20 mM). Sodium possesses high affinity towards this mechanism. The two mechanisms operate in a parallel manner beyond a diffusion barrier (= plasmamembrane) surrounding the cells. It is assumed that both the metabolic and the non-metabolic mechanisms operate in the entire concentration spectrum, but their relative contribution to the total uptake varies at different ranges.     

Aluminum Tolerance in Wheat (Triticum aestivum L.) (I. Uptake and Distribution of Aluminum in Root Apices)

We investigated the uptake and distribution of Al in root apices of near-isogenic wheat (Triticum aestivum L.) lines differing in Al tolerance at a single locus (Alt1: aluminum tolerance). Seedlings were grown in nutrient solution that contained 100 [mu]M Al, and the roots were subsequently stained with hematoxylin, a compound that binds Al in vitro to form a colored complex. Root apices of Al-sensitive genotypes stained after short exposures to Al (10 min and 1 h), whereas apices of Al-tolerant seedlings showed less intense staining after equivalent exposures. Differential staining preceded differences observed in either root elongation or total Al concentrations of root apices (terminal 2-3 mm of root). After 4 h of exposure to 100 [mu]M Al in nutrient solution, Al-sensitive genotypes accumulated more total Al in root apices than Al-tolerant genotypes, and the differences became more marked with time. Analysis of freeze-dried root apices by x-ray microanalysis showed that Al entered root apices of Al-sensitive plants and accumulated in the epidermal layer and in the cortical layer immediately below the epidermis. Long-term exposure of sensitive apices to Al (24 h) resulted in a distribution of Al coinciding with the absence of K. Quantitation of Al in the cortical layer showed that sensitive apices accumulated 5- to 10-fold more Al than tolerant apices exposed to Al solutions for equivalent times. These data are consistent with the hypothesis that Alt1 encodes a mechanism that excludes Al from root apices.     

Aluminum Interactions with Voltage-Dependent Calcium Transport in Plasma Membrane Vesicles Isolated from Roots of Aluminum-Sensitive and -Resistant Wheat Cultivars

The role of Al interactions with root-cell plasma membrane (PM) Ca2+ channels in Al toxicity and resistance was studied. The experimental approach involved the imposition of a transmembrane electrical potential (via K+ diffusion) in right-side-out PM vesicles derived from roots of two wheat (Triticum aestivum L.) cultivars (Al-sensitive Scout 66 and Al-resistant Atlas 66). We previously used this technique to characterize a voltage-dependent Ca2+ channel in the wheat root PM (J.W. Huang, D.L. Grunes, L.V. Kochian [1994] Proc Natl Acad Sci USA 91: 3473-3477). We found that Al3+ effectively blocked this PM Ca2+ channel; however, Al3+ blocked this Ca2+ channel equally well in both the Al-sensitive and -resistant cultivars. It was found that the differential genotypic sensitivity of this Ca2+ transport system to Al in intact roots versus isolated PM vesicles was due to Al-induced malate exudation localized to the root apex in Al-resistant Atlas but not in Al-sensitive Scout. Because malate can effectively chelate Al3+ in the rhizosphere and exclude it from the root apex, the differential sensitivity of Ca2+ influx to Al in intact roots of Al-resistant versus Al-sensitive wheat cultivars is probably due to the maintenance of lower Al3+ activities in the root apical rhizosphere of the resistant cultivar.     

The effect of Aging on Ion Uptake by Excised Barley Roots

When excised barley (Hodeum Vulgare L.) roots were aged, the rate of uptake of K and C1 increased raching a maximum in about 14 to 18 h. At its maximum the uptake rate was approxiamately twice that of the freshly excised root materila. Respiratory activity declined markedly during the aging period. Although excision was an essential requirement for uptake enhancement, washing was not. Both the uptake and aging processes were shown ot be unresponsive to the presence or absence of Ca in the tratment solutions. Because of cation exchange properties, the change in the total cation content of the root material was a more meaningful measure of the metabolically mediated cation uptake than was the change in content of the test cation itself. The pluggin of xylem vessels with protein and pectin-like substances was observed to increase with aging. It is proposed that the occlusion of the vessels may account for an apparent increaase in uptake since the obstruction could reduce a concurrent loss of ions through the cut ends of the root segments.     

部分耐盐小麦品种(系)SSR位点遗传多样性研究

选择有多态性的32对SSR引物对80个小麦耐盐品种(系)进行遗传差异研究,共检测出155个等位变异,平均每个位点上有4.75个等位变异;供试80份耐盐小麦品种(系)来源广泛,遗传基础丰富,表现出较高的遗传多样性,遗传相似系数范围在0.26～0.81;聚类分析结果显示,冬性小麦品种(系)聚为一大类;春性小麦品种(系)也聚为一大类;一些系谱相同或相近的品种(系)遗传相似系数较大;A、B、D基因组中SSR位点平均等位变异差异不大,以B基因组较高.     

Interaction between Aluminum Toxicity and Calcium Uptake at the Root Apex in Near-Isogenic Lines of Wheat (Triticum aestivum L.) Differing in Aluminum Tolerance

Aluminum (Al) is toxic to plants at pH < 5.0 and can begin to inhibit root growth within 3 h in solution experiments. The mechanism by which this occurs is unclear. Disruption of calcium (Ca) uptake by Al has long been considered a possible cause of toxicity, and recent work with wheat (Triticum aestivum L. Thell) has demonstrated that Ca uptake at the root apex in an Al-sensitive cultivar (Scout 66) was inhibited more than in a tolerant cultivar (Atlas 66) (J.W. Huang, J.E. Shaff, D.L. Grunes, L.V. Kochian [1992] Plant Physiol 98: 230-237). We investigated this interaction further in wheat by measuring root growth and Ca uptake in three separate pairs of near-isogenic lines within which plants exhibit differential sensitivity to Al. The vibrating calcium-selective microelectrode technique was used to estimate net Ca uptake at the root apex of 6-d-old seedlings. Following the addition of 20 or 50 [mu]M AlCl3, exchange of Ca for Al in the root apoplasm caused a net Ca efflux from the root for up to 10 min. After 40 min of exposure to 50 [mu]M Al, cell wall exchange had ceased, and Ca uptake in the Al-sensitive plants of the near-isogenic lines was inhibited, whereas in the tolerant plants it was either unaffected or stimulated. This provides a general correlation between the inhibition of growth by Al and the reduction in Ca influx and adds some support to the hypothesis that a Ca/Al interaction may be involved in the primary mechanism of Al toxicity in roots. In some treatments, however, Al was able to inhibit root growth significantly without affecting net Ca influx. This suggests that the correlation between inhibition of Ca uptake and the reduction in root growth may not be a mechanistic association. The inhibition of Ca uptake by Al is discussed, and we speculate about possible mechanisms of tolerance.     

Light-dependent Proton Excretion of Wheat (Triticum aestivum L.) and Maize (Zea mays L.) Roots

We investigated the change of root net proton excretion of seedlings of Triticum aestivum L. and Zea mays L. with daily variation of illumination using a multi-channel pH-stat system. We found an increase of net proton excretion during darkness and a drop after the beginning of illumination. Inhibition of carotenoid biosynthesis by norflurazone and photooxidation of chlorophylls did not change the periodicity or its induction. The induction of diurnal periodicity was possible with blue, green and red light. After induction the oscillation of net proton excretion continued for at least two cycles under constant light. We conclude that net H⁺ excretion of wheat and maize roots may be regulated by an endogenous clock or by a signal from the leaves. The nature of such a hypothetical signal remains unknown.     

The Direction of Growth of Seminal Roots of Triticum aestivum L. and Experimental Modification Thereof

The seminal root system of wheat (Triticum aestivum L.) consistsof a primary seminal root and the first and second pair of seminalroots, counting upwards. These roots are plagiotropic. The directionof growth was estimated as the angle from the vertical for eachof the three types of seminal roots that protruded from a hemispherical,soil-filled basket buried in the field. The angle of growthvaried with cultivar. It was smallest in the primary seminalroot and largest in the second pair of roots in all 12 cultivarsgrown in the field. Attempts to modify the angle of growth weremade under controlled environmental conditions. When the grainwas sown with its embryo face down, the angle of growth of thefirst pair of roots became smaller in the cultivars with inherentlylarger angles. The excision of the primary seminal root affectedthe first pair of roots and the excision of the first pair affectedthe second pair. These effects comprised a decrease in the angleof growth and an increase in root diameter. These changes inthe angle of growth caused by root excision is interpreted asa kind of compensatory growth. The direction of root growthand its impact on shaping wheat root system is discussed.Copyright1994, 1999 Academic Press Compensatory growth, direction of growth, gravitropism, liminal angle, plagiotropism, seminal roots, Triticum aestivum L., wheat     

The Effects of pH and Carbon Dioxide on Ion Uptake by Excised Barley Roots

The effects of carbon dioxide concentrations up to 8 per centin air on uptake of potassium and chloride at two pH levels,nominally pH 6 and 8, werestudied. In all experiments, enhanced uptake of potassium occurred atthe higher pH level with carbon dioxidefree air, but chlorideuptake was generally unaffected. At nominal pH 6, 1 per cent carbon dioxide reduced and 6 percent increased potassium uptake. There was no effect on chlorideuptake except with 1 per cent carbon dioxide where a markedenhancement was recorded. At nominal pH 8, l and 2 per cent carbon dioxide increased potassiumuptake whereas 6 and 8 per cent were inhibitory. Chloride uptakewas favoured by 1 and 8 per cent concentrations of the gas.     

Kinetics of Potassium and Magnesium Uptake by Intact Soybean Roots

The kinetics of uptake of K⁺ and Mg²⁺ were studied by using intact soybean [Glycine max (L.) Merr. cv. Amsoy] roots. Uptake of K⁺ in the concentration range 1.29 × 10^?5 to 1.82 × 10^?3 M can be represented by two phases of a single, multiphasic mechanism. Similarly, uptake of Mg²⁺ in the concentration range 4.10 × 10^?6 to 2.49 × 10^?4M was biphasic.