Exogenous zeatin accumulation in wheat root cells shows its role in regulation of cytokinin transport

Here we have shown that 24 hours after addition of zeatin to the nutrient solution the cytokinin content in xylem sap of wheat plants appears to be about 10 times lower that in the nutrient solution. Cytokinins accumulated mostly in roots and not in shoots of treated plants. These data demonstrate the existence of some barrier on cytokinin pathway from the nutrient solution to the plant shoot. With the help of Sudan III an increase in lignin and suberin deposition in the endodermis could be detected, being stronger with the increase in the distance from the root tip. The increase in deposition of suberin and lignin coincided with the decrease in cytokinin immunolabeling in root cells revealed with the help of monoclonal cytokinin antibodies and the second gold-labelled antibodies. Simultaneously exogenous cytokinins accumulated in root stele cells showing that the Casparian band was not only barrier on cytokinin pathway to plant shoot. It is concluded that high cytokinin immunolabe ling in the stele parenchyma cells around the stele vessels demonstrated accumulation of cytokinins by these cells, which could be important in regulation of cytokinin loading to the xylem vessels during there transport to the shoot. The role of cytokinin transporters is discussed.     

Hormonal Control of the Shoot-to-Root Ratio Is Not Related to Water Deficiency in Wheat Plants under Mineral Deficiency

We measured the content of hormones, the rate of growth, and some parameters of water regime (water content, transpiration, and stomatal and hydraulic conductivities) one and two days after wheat plant transfer from 10 to 1% Hoagland-Arnon nutrient medium. It was shown that, a day after dilution of nutrient solution, the content of various cytokinin forms decreased in the xylem sap, shoots, and roots. This decrease was most pronounced in the case of zeatin in the xylem sap and zeatin riboside in the mature zone of the first leaf. ABA was found to accumulate in shoots. A day after dilution of nutrient solution, we observed root elongation evidently induced by mineral nutrient deficiency, and this accelerated root growth was maintained later. Two days after dilution of nutrient solution, we observed the slowing of shoot weight accumulation, whereas root weight remained unchanged. Plant growth response could be related to ABA accumulation in shoots and cytokinin depletion in the whole plant. A reduced hydraulic conductivity and water content in the growing leaf zone was detected only two days after dilution of nutrient solution. Thus, changes in the growth rates and hormone contents could not result from disturbances in water regime induced by mineral nutrient deficiency.     

Ultrastructure of the haustorial interface and apoplastic continuum between host and the root hemiparasiteOlax phyllanthi (Labill.) R. Br. (Olacaceae)

Summary Structural features of haustorial interface parenchyma of the root hemiparasiteOlax phyllanthi are described. Walls contacting host xylem are thickened non-uniformly with polysaccharides, not lignin, and show only a thin protective wall layer when abutting pits in walls of host xylem vessels or tracheids. Lateral walls of interface parenchyma exhibit an expanded middle layer of open fibrillar appearance, sometimes with, but mostly lacking adjoining layers of dense wall material. Free ribosomes and rough endoplasmic reticulum are prominent and occasional wall ingrowths present. Experiments involving transpirational feeding of the apoplast tracers lanthanum nitrate or uranyl acetate to host roots cut below haustorial connections, indicate effective apoplastic transfer from host to parasite root via the haustorium. Deposits of the tracers suggest a major pathway for water flow through host xylem pits, across the thin protective wall layer, and thence into the haustorium via the electronopaque regions of the terminal and lateral walls of the contact parenchyma. Graniferous tracheary elements and walls of parenchyma cells of the body of the haustorium appear to participate in tracer flow as do walls of cortical cells, stele parenchyma and xylem conducting elements of the parasite root, suggesting that both vascular and non-vascular routes are involved in extracytoplasmic transfer of xylem sap from host to parasite. The Casparian strip of the endodermis and the suberin lamella of the exodermis of theOlax root act as barriers to flow within the system.     

三角叶滨藜根吸水特点与其抗盐性的关系

采用压力室和冰点渗透压计测定了三角叶滨藜在不同浓度NaCl的根系环境溶液中根木质部的压力势和伤流液的渗透势,并利用原子吸收分光光度计测定了植株和伤流液以及环境溶液中Na 含量。结果表明:随着根环境溶液NaCl浓度的增加,三角叶滨藜植株和木质部伤流液中Na 含量虽呈上升趋势,但根系的过滤系数和体内Na 相对累积量逐渐降低,说明三角叶滨藜根细胞对盐分有很强的过滤作用;木质部伤流液的渗透势随着环境溶液渗透势的降低而降低,但根木质部溶液的水势则逐渐高出根外环境溶液的渗透势;表明三角叶滨藜能够利用较低的木质部负压来抵抗根外溶液的低渗透势而反渗透吸水,并利用根细胞对盐分的过滤作用来避免从环境摄取过量的盐分。     

Dependence of cytokinin distribution in plants on their physical and chemical properties and transpiration rate

The effect of transpiration on cytokinin accumulation and distribution in 7-day-old wheat (Triticum durum Desf.) seedlings grown on nutrient medium supplemented with zeatin or its riboside was studied. The content of cytokinins in plants and nutrient medium was measured by the immunoenzyme analysis; cytokinin distribution between root cells was assessed immunohistochemically using antibodies against zeatin derivatives. The rate of transpiration was reduced 20-fold by plant placing in humid chamber. At normal transpiration, after 6 h of plant incubation on the solution of zeatin, the level of cytokinins in plant tissues increased stronger than after incubation on the solution of zeatin riboside (by 7.3 and 3.5 times, respectively, as compared with control), although the rates of both cytokinin uptake were equal. Most portions of cytokinins were retained in the roots, which was stronger expressed in the case of free zeatin uptake. A decrease in the rate of transpiration did not affect substantially the zeatin absorption from nutrient medium and the total level of cytokinin accumulation in plants, but these indices were sharply decreased in the case of zeatin riboside. In the zone of absorption of both control roots and roots treated with cytokinins, more intense cytokinin immunostaining was observed in the cells of the central cylinder. The interrelation between cytokinin distribution between the cells and apoplast, their inactivation, and transport over the plant and their form (zeatin or zeatin riboside) used for treatment is discussed.     

Ultrastructure of xylem parenchyma cells of barley roots in relation to ion transport to the xylem

Summary The structure of xylem parenchyma cells is examined in relation to transport of ions through the root. Measurement of uptake of ⁸⁶Rb⁺ and its transport through the root at different distances from the apex show that this is a general activity along the length of the root and not confined to a limited region. Thus transport through the root is not stopped by removal of that part of the root tip containing metaxylem vessels with living contents. The structure of xylem parenchyma appears to be suitable for involvement in ion transport from the stele to the xylem. At 1 cm behind the tip, where metaxylem vessels have no living contents but ion uptake and transport are going on at high rates, xylem parenchyma cells are rich in cytoplasm with extensive rough endoplasmic reticulum and well-developed mitochondria. Their cell walls contain numerous plasmodesmata, establishing the possibility of a symplastic pathway across the stele up to the vessels. The results are discussed in relation to regulation of ion transport to the xylem vessels in roots.Dedicated to Professor O. Stocker on the occasion of his 85th birthday.     

Cytokinin content and immunolocalization in wheat primary root cells

Immunohystochemical localization of cytokinins in cells of different root zones of wheat plants showed intensive immunostaining of zeatin in the apical root zone and its subsequent decline with the increase in the distance from the root tip. More intensive labeling of metaxylem and parenchyma cells of the root central cylinder was observed on the sections of the zone where root hairs appeared. Above this zone the decline in immunostaining of the cells of the central cylinder was paralleled by the signs if finalization of differentiation of the xylem vessels shown by lignin deposition. The data of immunohystochemical staining were confirmed by the results of enzyme immunoassay of different cytokinin forms. Likely sources of zeatin accumulation are considered. Possibility of additional (alongside with that in apical root zone) synthesis of cytokinins in the vascular tissues of root and the role of cytokinins in stimulation oflignification are discussed.     

The Arabidopsis translatome cell-specific mRNA atlas: Mining suberin and cutin lipid monomer biosynthesis genes as an example for data application

Plants consist of distinct cell types distinguished by position, morphological features and metabolic activities. We recently developed a method to extract cell-type specific mRNA populations by immunopurification of ribosome-associated mRNAs. Microarray profiles of 21 cell-specific mRNA populations from seedling roots and shoots comprise the Arabidopsis Translatome dataset. This gene expression atlas provides a new tool for the study of cell-specific processes. Here we provide an example of how genes involved in a pathway limited to one or few cell-types can be further characterized and new candidate genes can be predicted. Cells of the root endodermis produce suberin as an inner barrier between the cortex and stele, whereas the shoot epidermal cells form cutin as a barrier to the external environment. Both polymers consist of fatty acid derivates, and share biosynthetic origins. We use the Arabidopsis Translatome dataset to demonstrate the significant cell-specific expression patterns of genes involved in those biosynthetic processes and suggest new candidate genes in the biosynthesis of suberin and cutin.Key words: cell-type specific expression, polysome immunopurification, translatome, suberin, cutin, endodermis, epidermis, arabidopsis     

Chemical composition and ultrastructure of broad bean (<Emphasis Type="Italic">Vicia faba</Emphasis> L.) nodule endodermis in comparison to the root endodermis

Ultrastructure and development of apoplastic barriers within indeterminate root nodules formed by Vicia faba L. were examined by light and electron microscopy. The nodule outer cortex is separated from the inner cortex by a heavily suberized nodule endodermis, which matures in submeristematic regions and possesses suberin lamellae. Unsuberized passage cells are present near vascular strands, which are surrounded by a vascular endodermis attached on the inner side of the nodule endodermal cell walls. The vascular endodermis appears immediately below the meristematic apex in developmental state I (Casparian bands), gradually develops suberin lamellae, and attains developmental state II at the base of the nodule. For chemical analysis apoplastic barrier tissues were dissected after enzymatic digestion of non-impregnated tissues. Root epidermal and endodermal cell walls as well as nodule outer cortex could be isolated as pure fractions; nodule endodermal cell walls could not be separated from vascular endodermal cell walls and enclosed xylem vessels. Gas chromatography-flame ionization detection and gas chromatography-mass spectrometry were applied for quantitative and qualitative analysis of suberin and lignin in isolated cell walls of these tissues. The suberin content of isolated endodermal cell walls of nodules was approximately twice that of the root endodermal cell walls. The suberin content of the nodule outer cortex and root epidermal cell walls was less than one-tenth of that of the nodule endodermal cell wall. Substantial amounts of lignin could only be found in the nodule endodermal cell wall fraction. Organic solvent extracts of the isolated tissues revealed long-chain aliphatic acids, steroids, and triterpenoid structures of the lupeol type. Surprisingly, extract from the outer cortex consisted of 89% triterpenoids whereas extracts from all other cell wall isolates contained not more than 16% total triterpenoids. The results of ultrastructural and chemical composition are in good correspondence and underline the important role of the examined tissues as apoplastic barriers.     

植物对硅的吸收转运机制研究进展

硅(Si)能缓解生物与非生物胁迫对植物的毒害作用,Si的吸收转运是由Si转运蛋白介导的.最近,多个Si转运蛋白(Lsi)基因相继在水稻、大麦和玉米中被克隆出来,并在Si的吸收转运机制方面取得了很大进展.水稻OsLsi在根组织中呈极性分布,OsLsi1定位在根外皮层和内皮层凯氏带细胞外侧质膜,负责将外部溶液中的单硅酸转运到皮层细胞内.OsLsi2定位在凯氏带细胞内侧质膜,在外皮层中负责将Si输出到通气组织质外体中,在内皮层与OsLsi1协同作用将Si转运到中柱中.导管中的Si通过蒸腾流转运到地上部,再由定位在叶鞘和叶片木质部薄壁细胞靠近导管一侧的OsLsi6负责木质部Si的卸载和分配.在大麦和玉米中,ZmLsi1/HvLsi1定位在根表皮和皮层细胞外侧质膜负责Si的吸收,然后Si通过共质体途径被转运到内皮层凯氏带细胞中,再由ZmLsi2/HvLsi2输出转运到中柱中.ZmLsi6在细胞中的定位和活性与OsLsi6相似,推测其可能具有类似的功能,但大麦Lsi6至今未见报道.所以,Si转运机制仍需要进一步研究.     

Feedback regulation of xylem cytokinin content is conserved in pea and Arabidopsis

Increased-branching mutants of garden pea (Pisum sativum; ramosus [rms]) and Arabidopsis (Arabidopsis thaliana; more axillary branches) were used to investigate control of cytokinin export from roots in relation to shoot branching. In particular, we tested the hypothesis that regulation of xylem sap cytokinin is dependent on a long-distance feedback signal moving from shoot to root. With the exception of rms2, branching mutants from both species had greatly reduced amounts of the major cytokinins zeatin riboside, zeatin, and isopentenyl adenosine in xylem sap compared with wild-type plants. Reciprocally grafted mutant and wild-type Arabidopsis plants gave similar results to those observed previously in pea, with xylem sap cytokinin down-regulated in all graft combinations possessing branched shoots, regardless of root genotype. This long-distance feedback mechanism thus appears to be conserved between pea and Arabidopsis. Experiments with grafted pea plants bearing two shoots of the same or different genotype revealed that regulation of root cytokinin export is probably mediated by an inhibitory signal. Moreover, the signaling mechanism appears independent of the number of growing axillary shoots because a suppressed axillary meristem mutation that prevents axillary meristem development at most nodes did not abolish long-distance regulation of root cytokinin export in rms4 plants. Based on double mutant and grafting experiments, we conclude that RMS2 is essential for long-distance feedback regulation of cytokinin export from roots. Finally, the startling disconnection between cytokinin content of xylem sap and shoot tissues of various rms mutants indicates that shoots possess powerful homeostatic mechanisms for regulation of cytokinin levels.     

Enhanced formation of aerenchyma and induction of a barrier to radial oxygen loss in adventitious roots of Zea nicaraguensis contribute to its waterlogging tolerance as compared with maize (Zea mays ssp. mays)

Enhancement of oxygen transport from shoot to root tip by the formation of aerenchyma and also a barrier to radial oxygen loss (ROL) in roots is common in waterlogging‐tolerant plants. Zea nicaraguensis (teosinte), a wild relative of maize (Zea mays ssp. mays), grows in waterlogged soils. We investigated the formation of aerenchyma and ROL barrier induction in roots of Z. nicaraguensis, in comparison with roots of maize (inbred line Mi29), in a pot soil system and in hydroponics. Furthermore, depositions of suberin in the exodermis/hypodermis and lignin in the epidermis of adventitious roots of Z. nicaraguensis and maize grown in aerated or stagnant deoxygenated nutrient solution were studied. Growth of maize was more adversely affected by low oxygen in the root zone (waterlogged soil or stagnant deoxygenated nutrient solution) compared with Z. nicaraguensis. In stagnant deoxygenated solution, Z. nicaraguensis was superior to maize in transporting oxygen from shoot base to root tip due to formation of larger aerenchyma and a stronger barrier to ROL in adventitious roots. The relationships between the ROL barrier formation and suberin and lignin depositions in roots are discussed. The ROL barrier, in addition to aerenchyma, would contribute to the waterlogging tolerance of Z. nicaraguensis.     

Permeability of Plant Young Root Endodermis to Cu Ions and Cu-Citrate Complexes in Corn and Soybean

The non-selective apoplastic passage of Cu and Cu-citrate complexes into the root stele of monocotyledonous corn and dicotyledonous soybean was investigated using an inorganic-salt-precipitation technique. Either Cu ions or Cu-citrate complexes were drawn into root through the apoplast from the root growth medium, and K₄[Fe(CN)₆] was subsequently perfused through xylem vessels or the entire root cross section. Based on microscopic identification of the reddish-brown precipitates of copper ferrocyanide in the cell walls of the xylem of corn and soybean roots, Cu²⁺ passed through the endodermal barrier into the xylem of both species. When the solution containing 200 μM CuSO₄ and 400 μM sodium citrate (containing 199.98 μM Cu-citrate, 0.02 μM Cu²⁺) was drawn via differential pressure gradients into the root xylem while being perfused with K₄[Fe(CN)₆] through the entire root cross-section, reddish-brown precipitates were observed in the walls of the stele of soybean, but not corn root. However, when a CuSO₄ solution containing 0.02 or 0.2 μM free Cu²⁺ was used, no reddish-brown precipitates were detected in the stele of either of the two plants. Results indicated that endodermis was permeable to Cu-citrate complexes in primary roots of soybean, but not corn. The permeability of the endodermal barrier to the Cu-citrate complex may vary between dicotyledonous and monocotyledonous plants, which has considerable implications for chelant-enhanced phytoextraction.     

The Radial and Longitudinal Path of Ion Movement in Roots

The existence of a barrier to lateral outward diffusion of ions from roots was demonstrated quantitatively and by autoradiography. Ions applied to the apical zone were excreted through the basal cut end with no lateral outward diffusion from the central zone. The ions moving through the conducting tissues showed no leakage to the external solution regardless whether the treatment roots came from seedlings grown under low or high salt conditions. However, when dinitrophenol (DNP) was also applied to the central zone in the external salt solution, leakage of apically applied calcium occurred from the conducting tissues. Autoradiographic studies with labeled calcium suggested that the endodermal layer was an effective barrier preventing the lateral outward diffusion of ions. The ions moved longitudinally through the stele. In the stelar tissues the autoradiographic studies failed to detect the presence of any radioactive calcium in the central duct and in the mature xylem vessels, although high concentrations of labeled ions were found in the living cells of the stele, particularly the xylem parenchyma. It is suggested that xylem parenchyma cells may be involved in the longitudinal transport of ions.     

Identification of Arabidopsis subtilisin-like serine protease specifically expressed in root stele by gene trapping

Xylem plays a role not only in the transport of water and nutrients but also in the regulation of growth and development through the transport of biologically active substances. In addition to mineral salts, xylem sap contains hormones, organic nutrients and proteins. However, the physiological functions of most of those substances remain unclear. To explore genes involved in xylem sap production, we identified Arabidopsis genes expressed in the root stele of the root hair zone from gene-trap lines by randomly inserting the β-glucuronidase gene into the genome. Among 26 000 gene-trap lines, we found that 10 lines had β-glucuronidase (GUS) staining predominantly in the root stele of the root hair zone and no GUS staining in the shoots. Of these 10 lines, 2 lines showed that gene-trap tags inserted into the promoter region of the same gene, denoted Arabidopsis thaliana subtilase 4.12( AtSBT4.12 ). Analysis of AtSBT4.12 promoter using an pAtSBT4.12 ::β-glucuronidase transgenic line showed that the AtSBT4.12 gene was expressed only in the root stele of the root hair zone. AtSBT4.12 expression in roots was increased by application of methyl jasmonate. Subtilase proteins are commonly detected in proteomic analyses of xylem sap from various plant species, including Brassica napus , a relative of Arabidopsis . These results suggest that AtSBT4.12 may be a protein localized in the apoplast of root stele including xylem vessel and involved in stress responses in Arabidopsis roots.     

植物钙素吸收和运转

近年来,钙素在植物体内的吸收和运输研究主要集中在细胞和分子水平,但整株水平上的研究也同样重要.整株水平上的钙吸收和运输包括根细胞的钙吸收、钙离子横向穿过根系并进入木质部、在木质部运输、从木质部移出并进入叶片或果实及在叶片或果实中运转分配等环节,既经过质外体也穿越共质体.钙离子通道、Ca2 -ATP酶和Ca2 /H 反向转运器等参与根细胞的钙吸收.在钙离子横向穿根进入木质部的过程中,需要穿越内皮层和木质部薄壁细胞组织.根系内皮层凯氏带阻挡了Ca2 沿质外体途径由内皮层外侧向内侧的移动,部分Ca2 由此通过离子通道流进内皮层细胞而转入共质体并到达木质部薄壁细胞组织,而由木质部薄壁细胞组织进入中柱质外体可能需要Ca2 -ATP酶驱动;还有一些Ca2 由内皮层细胞运出,沿内皮层内侧的质外体途径进入木质部导管,并通过导管运向枝干.钙离子以螯合态的形式在枝干导管运输;水流速率是影响钙离子沿导管运输的关键因子.钙离子在果实和叶片中的运输和分配不仅通过质外体途径也通过共质体途径.     

Functions of passage cells in the endodermis and exodermis of roots

Passage cells frequently occur in the endodermis and exodermis but are not ubiquitous in either layer. Passage cells occur in the form of short cells in the dimorphic type of exodermis. In both layers, Casparian bands are formed in all cells, but the subsequent development of suberin lamellae and thick, cellulosic walls are delayed or absent in the passage cells. Available evidence suggests that passage cells of the endodermis are important for the transfer of calcium and magnesium into the stele and thus into the transpiration stream. They become the only cells which present a plasmalemma surface to the soil solution (and are thus capable of ion uptake) when the epidermis and central cortex die. This occurs naturally in some herbaceous and woody species and is known to be promoted by drought. Most evidence indicates that the development of suberin lamellae in both the endodermis and exodermis increases the resistance of the root to the radial flow of water. Passage cells thus provide areas of low resistance for the movement of water, and the position of these cells in the endodermis (i.e., in close proximity to the xylem) is explained in terms of function. Exodermal passage cells have a cytoplasmic structure suggesting an active role in ion uptake. This may be related to the tendency of the epidermis to die, leaving the passage cells as the only ones with their membranes exposed to the soil solution. Passage cells in the exodermis attract endomycorrhizal fungi while those in the endodermis do not. It is clear that passage cells of the endodermis and exodermis play a variety of roles in the plant root system.