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.     

知母根的组织发育及其与生长环境的关系

观察知母根的组织发育,发现具有典型的单子叶植物根的组织特征,特别是在同一条根中初生木质部脊数可以随根的发育而增加,同一株知母不同根之间初生木质部脊数也不尽相同;内皮层细胞壁增厚由凯氏带发育为马蹄形,内皮层外侧1－3层皮层细胞内侧壁可以木栓化或强烈木化增厚,这种变化与其生长环境具有密切关系,当其移植于湿润环境中后,内皮层马蹄形增厚减弱,其外侧皮层细胞内侧壁仅发生木柱化增厚,且减弱。     

矢母根的组织发育及其与生长环境的关系

观察知母根的组织发育,发现具有典型的单子叶植物根的组织特征,特别是在同一条根中初生木质部脊数可以随机的发育而增加,同一株知母不同根之间初生木质部脊数也不尽相同;内皮层细胞壁增厚由凯氏带发育淡马蹄形,内皮层外侧１－３层皮层细胞内侧壁可以木栓化或强烈木化增厚。     

植物凯氏带形成分子机制及功能特点的研究进展

凯氏带位于被子植物初生根内皮层细胞,环绕细胞1周,是与质膜紧密结合的非极性带状增厚结构。凯氏带作为植物根中离子径向运输障碍,调节离子的质外体吸收途径,迫使土壤中的离子通过内皮层细胞膜,选择性地进入中柱。凯氏带发现于1865年,但直至拟南芥凯氏带蛋白的发现和凯氏带阻滞作用物质基础被揭示,凯氏带的形成机理和功能才逐渐为人们所认知。凯氏带的物质基础为木质素,其形成需要由凯氏带蛋白和受体激酶组成的合成平台。细胞内部的木质素单体经ABCG载体运输到凯氏带的形成区,经ESB1dirigent蛋白、RBOHF氧化酶和PER64过氧化物酶等催化,合成木质素。该文对近年来国内外有关凯氏带形成的分子机制和功能特点方面的研究进展进行综述,为进一步理解和解析凯氏带的形成机理和功能提供参考。     

大豆初生根凯氏带对铜离子的通透性

采用在根内生成有色铜沉淀的方法研究大豆（Glycine max）初生根凯氏带对铜离子的通透性。用真空泵抽取浓度为200μmol·L^–1的CuSO4溶液进入根中,然后在重力作用下从根基部灌注400μmol·L^–1的K4[Fe（CN）6]溶液,两种物质在根内相遇即可产生棕色的Cu2[Fe（CN）6]沉淀,根据沉淀的位置来确定铜离子所经过的途径。结果表明：Cu^2＋可以穿过内皮层凯氏带,在木质部导管壁以及凯氏带至木质部之间的细胞壁处产生棕色沉淀,侧根发生的部位也产生了大量的沉淀;当抽取K4[Fe（CN）6]溶液后再灌注CuSO4溶液,发现Cu^2＋仍然可以穿过凯氏带,并在凯氏带外侧以及外皮层细胞的细胞壁处产生棕色沉淀。研究结果证明凯氏带并不是一个可以完全阻止离子进出的完美屏障。     

单子叶植物根的内皮层及其观察方法

裸子植物和具有少量次生长的双子叶植物根的内皮层细胞通常都具有径向壁和横壁加厚而成的凯氏带结构。但多数单子叶植物和少数双子叶植物(没有次生生长的种类)根的内皮层细胞只有在发育的早期(幼根)具有凯氏带,发育的后期(较老的根)内皮层细胞壁内敷贴了一整层栓质层,随后细胞壁又经过了五面加厚(只有邻接皮层一面的壁没有加厚)或六面加厚(即全部细胞壁都加厚),加厚部分都经过了木质化,因此这时就看不到凯氏带了。由于内皮层细胞内敷贴了一整层栓质层,这类植物根的维管柱似乎被一层不透水的套子所隔开了。实际不然,因为并不是全部内皮层细胞都具有五面或六面加厚的壁,通常有一部分内皮层细胞,就是在根的横切面上靠近木质部角端的那一部分内皮层细胞,仍有未栓质化、未加厚、只有凯氏带的纤维素薄壁,这一部分内皮层细胞     

菖蒲营养器官的解剖结构特征及其生态适应性

采用石蜡切片法和组织离析法观察湿地植物菖蒲Acorus calamus营养器官的解剖结构特征,并探讨与菖蒲生态适应性的关系。结果表明,菖蒲根状茎具有不定根,不定根的表皮由一层细胞组成,排列紧密;皮层由多层薄壁细胞组成,薄壁细胞破裂形成多数的通气道;内皮层存在细胞壁马蹄形加厚的凯氏带,维管柱具5原型初生木质部,具4~10个大型导管,形成发达的通气组织。根状茎外层表皮细胞类方形,外壁增厚;基本组织近表皮有厚壁细胞团,皮层薄壁细胞呈链状排列,中间有较大的通气组织,内皮层形成凯氏带,有大量导管附着在凯氏带周围;维管束散生于基本组织,中柱维管束为周木型。根状茎部分区域存在与内生菌共生形成的结瘤。叶是等面叶,叶肉组织薄壁细胞破裂形成大的通气组织,叶脉具有限外韧维管束,中间有一较大的导管。菖蒲的解剖结构特点与其湿地生长环境相适应,根状茎存在结瘤状内共生菌,具有净化污水作用,不定根发达适于分株繁殖。     

玉米气生根是观察通道细胞的好材料

在高等院校植物课本和实验中都要讲解和观察玉米根结构中的内皮层细胞.它具有两种细胞:一种是细胞壁的五面加厚,只有外切向壁较薄,在横切面上次生增厚的部分象马蹄形;另一种是少数位于木质部束处的内皮层细胞,仍保持初期发育阶段的结构,即除凯氏带外,薄     

大豆初生根凯氏带对铜离子的通透性

采用在根内生成有色铜沉淀的方法研究大豆(Glycine max)初生根凯氏带对铜离子的通透性。用真空泵抽取浓度为200 μmol·L^–1 的CuSO₄溶液进入根中, 然后在重力作用下从根基部灌注400 μmol·L^–1的K₄[Fe(CN)₆]溶液, 两种物质在根内相遇即可产生棕色的Cu₂[Fe(CN)₆]沉淀, 根据沉淀的位置来确定铜离子所经过的途径。结果表明: Cu²⁺可以穿过内皮层凯氏带, 在木质部导管壁以及凯氏带至木质部之间的细胞壁处产生棕色沉淀, 侧根发生的部位也产生了大量的沉淀; 当抽取K₄[Fe(CN)₆]溶液后再灌注CuSO₄溶液, 发现Cu²⁺仍然可以穿过凯氏带, 并在凯氏带外侧以及外皮层细胞的细胞壁处产生棕色沉淀。研究结果证明凯氏带并不是一个可以完全阻止离子进出的完美屏障。     

重金属镉(Cd)在植物体内的转运途径及其调控机制

重金属镉(Cd)的毒害效应与其由土壤向植物地上部分运输有关,揭示Cd~(2+)转运途径及其调控机制可为提高植物抗镉性以及镉污染的植物修复提供依据。对Cd~(2+)在植物体内的转运途径,特别是限制Cd~(2+)移动的细胞结构和分子调控机制研究进展进行了回顾。Cd~(2+)通过共质体和质外体途径穿过根部皮层进入木质部的过程中,大部分在皮层细胞间沉积,少部分抵达中柱后转移到地上部分。为了免受Cd~(2+)的危害,植物体产生了多种限制Cd~(2+)吸收和转移的生理生化机制:1)环绕在内皮层径向壁和横向壁上的凯氏带阻止Cd~(2+)以质外体途径进入木质部;2)螯合剂与进入根的Cd~(2+)螯合形成稳定化合物并区隔在液泡中;3)通过H+/Cd~(2+)离子通道等将Cd~(2+)逆向转运出根部。植物共质体和质外体途径转运重金属镉的能力以及两条途径的串扰尚待进一步明晰和阐明。     

Identification of maize silicon influx transporters

Maize (Zea mays L.) shows a high accumulation of silicon (Si),but transporters involved in the uptake and distribution havenot been identified. In the present study, we isolated two genes(ZmLsi1 and ZmLsi6), which are homologous to rice influx Sitransporter OsLsi1. Heterologous expression in Xenopus laevisoocytes showed that both ZmLsi1 and ZmLsi6 are permeable tosilicic acid. ZmLsi1 was mainly expressed in the roots. By contrast,ZmLsi6 was expressed more in the leaf sheaths and blades. Differentfrom OsLsi1, the expression level of both ZmLsi1 and ZmLsi6was unaffected by Si supply. Immunostaining showed that ZmLsi1was localized on the plasma membrane of the distal side of rootepidermal and hypodermal cells in the seminal and crown roots,and also in cortex cells in lateral roots. In the shoots, ZmLsi6was found in the xylem parenchyma cells that are adjacent tothe vessels in both leaf sheaths and leaf blades. ZmLsi6 inthe leaf sheaths and blades also exhibited polar localizationon the side facing towards the vessel. Taken together, it canbe concluded that ZmLsi1 is an influx transporter of Si, whichis responsible for the transport of Si from the external solutionto the root cells and that ZmLsi6 mainly functions as a Si transporterfor xylem unloading.     

HvLsi1 is a silicon influx transporter in barley

Most plants accumulate silicon in their bodies, and this is thought to be important for resistance against biotic and abiotic stresses; however, the molecular mechanisms for Si uptake and accumulation are poorly understood. Here, we describe an Si influx transporter, HvLsi1, in barley. This protein is homologous to rice influx transporter OsLsi1 with 81% identity, and belongs to a Nod26-like major intrinsic protein sub-family of aquaporins. Heterologous expression in both Xenopus laevis oocytes and a rice mutant defective in Si uptake showed that HvLsi1 has transport activity for silicic acid. Expression of HvLsi1 was detected specifically in the basal root, and the expression level was not affected by Si supply. There was a weak correlation between Si uptake and the expression level of HvLsi1 in eight cultivars tested. In the seminal roots, HvLsi1 is localized on the plasma membrane on the distal side of epidermal and cortical cells. HvLsi1 is also located in lateral roots on the plasma membrane of hypodermal cells. These cell-type specificity of localization and expression patterns of HvLsi1 are different from those of OsLsi1. These observations indicate that HvLsi1 is a silicon influx transporter that is involved in radial transport of Si through the epidermal and cortical layers of the basal roots of barley.     

Identification and Characterization of Maize and Barley Lsi2-Like Silicon Efflux Transporters Reveals a Distinct Silicon Uptake System from That in Rice

Silicon (Si) uptake has been extensively examined in rice (Oryza sativa), but it is poorly understood in other gramineous crops. We identified Low Silicon Rice 2 (Lsi2)-like Si efflux transporters from two important gramineous crops: maize (Zea mays) and barley (Hordeum vulgare). Both maize and barley Lsi2 expressed in Xenopus laevis oocytes showed Si efflux transport activity. Furthermore, barley Lsi2 was able to recover Si uptake in a rice mutant defective in Si efflux. Maize and barley Lsi2 were only expressed in the roots. Expression of maize and barley Lsi2 was downregulated in response to exogenously applied Si. Moreover, there was a significant positive correlation between the ability of roots to absorb Si and the expression levels of Lsi2 in eight barley cultivars, suggesting that Lsi2 is a key Si transporter in barley. Immunostaining showed that maize and barley Lsi2 localized only at the endodermis, with no polarity. Protein gel blot analysis indicated that maize and barley Lsi2 localized on the plasma membrane. The unique features of maize and barley Si influx and efflux transporters, including their cell-type specificity and the lack of polarity of their localization in Lsi2, indicate that these crops have a different Si uptake system from that in rice.     

Functional Characterization of a Silicon Transporter Gene Implicated in Silicon Distribution in Barley

Silicon (Si) is a beneficial element for plant growth. In barley (Hordeum vulgare), Si uptake by the roots is mainly mediated by a Si channel, Low Silicon1 (HvLsi1), and an efflux transporter, HvLsi2. However, transporters involved in the distribution of Si in the shoots have not been identified. Here, we report the functional characterization of a homolog of HvLsi1, HvLsi6. HvLsi6 showed permeability for Si and localized to the plasma membrane. At the vegetative growth stage, HvLsi6 was expressed in both the roots and shoots. The expression level was unaffected by Si supply. In the roots, HvLsi6 was localized in epidermis and cortex cells of the tips, while in the leaf blades and sheaths, HvLsi6 was only localized at parenchyma cells of vascular bundles. At the reproductive growth stage, high expression of HvLsi6 was also found in the nodes. HvLsi6 in node I was polarly located at the transfer cells surrounding the enlarged vascular bundles toward the numerous xylem vessels. These results suggest that HvLsi6 is involved in Si uptake in the root tips, xylem unloading of Si in leaf blade and sheath, and intervascular transfer of Si in the nodes. Furthermore, HvLsi2 was found to be localized at the parenchyma cell layer adjacent to the transfer cells with opposite polarity of HvLsi6, suggesting that the coupling of HvLsi6 and HvLsi2 is involved in the intervascular transfer of Si at the nodes. Si translocated via the enlarged vascular bundles is unloaded to the transfer cells by HvLsi6, followed by HvLsi2 to reload Si to the diffuse vascular bundles, which are connected to the upper part of the plant, especially the panicles, the ultimate Si sink.Silicon (Si) is a beneficial element for plant growth. It enhances the resistance of plants to various biotic and abiotic stresses (Epstein, 1999; Ma and Takahashi, 2002; Ma and Yamaji, 2006). For example, Si reduces the epidemics of both leaf and panicle blast in rice (Oryza sativa; Datnoff and Rodrigues, 2005) and decreases the incidence of powdery mildew in cucumber (Cucumis sativus), barley (Hordeum vulgare), and wheat (Triticum aestivum; Fauteux et al., 2005). Si also suppresses insect pests such as stem borer (Chilo suppressalis), brown planthopper (Nilaparvata lugens), and rice green leafhopper (Nephotettix cincticeps; Savant et al., 1997). Resistance to the damage by wild rabbit in wheat is also enhanced by an increased amount of Si in leaf tissue (Cotterill et al., 2007). Si is also able to alleviate lodging, drought, and low- and high-temperature stresses (Ma, 2004). The beneficial effects of Si under phosphate deficiency, phosphate excess, and manganese and salt toxicity stresses have been observed in many plants (Ma and Takahashi, 2002). Usually, the more Si that accumulates in the shoots, the greater its effect in enhancing the plant’s response. This is because most effects of Si are expressed through the formation of silica gel, which is deposited on leaves, stems, and other organs of plants (Ma and Yamaji, 2006). Therefore, for the plant to benefit from Si, a high accumulation is required. However, Si accumulation greatly varies with plant species, and this difference has been attributed to the ability of plants to take up Si.Transporters responsible for Si uptake by roots have been identified in several plant species, including barley, maize (Zea mays), pumpkin (Cucurbita moschata), rice, wheat (Ma et al., 2011), and most recently in horsetail (Equisetum arvense; EaNIP3s [for Nod26-like major intrinsic protein3]; Grégoire et al., 2012). Two different types of transporter, Si-permeable channel and efflux transporter, are involved in the Si-uptake process. Low Silicon1 (Lsi1) belongs to a NIP subfamily of aquaporin-like proteins and functions as a Si-permeable channel. Lsi1 in rice is localized in the distal side of root exodermis and endodermis (Ma et al., 2006), but Lsi1 in barley, maize, and pumpkin is localized in the epidermis and cortex (Chiba et al., 2009; Mitani et al., 2009b, 2011). On the other hand, Lsi2 functions as an efflux Si transporter and belongs to a putative anion transporter family without any similarity to Lsi1. Lsi2 in rice is also localized at the root exodermis and endodermis as Lsi1, but it is polarly localized at the proximal side (Ma et al., 2007). By contrast, Lsi2 in barley and maize is localized only to the endodermis of roots. Furthermore, these transporters do not show polar localization in barley and maize (Mitani et al., 2009a). Therefore, Si uptake mediated by Lsi1 and Lsi2 shows different pathways between rice and other plant species (Ma et al., 2011).Following uptake by the roots through Lsi1 and Lsi2, Si is translocated to the aboveground part and distributed in different tissues. Lsi6, a homolog of Lsi1, is involved in xylem unloading of Si in rice (Yamaji et al., 2008). Lsi6 is localized on the adaxial side of the xylem parenchyma cells in the leaf sheaths and leaf blades. Knockout of Lsi6 resulted in altered distribution of Si in the leaf cells. Furthermore, at the reproductive growth stage of rice, Lsi6 is also highly expressed at the nodes (Yamaji and Ma, 2009). At node I below the panicle, Lsi6 is mainly localized at the xylem transfer cells with polarity facing toward the xylem vessel (Yamaji and Ma, 2009). Knockout of Lsi6 decreased Si accumulation in the panicle but increased Si accumulation in the flag leaf. These findings indicate that Lsi6 is also required for the intervascular transfer of Si in rice, transferring Si from the enlarged vascular bundles coming from the roots to the diffuse vascular bundles connected to the panicle.Barley is a Si-accumulating species, although the accumulation extent is lower than that of rice. Transporters responsible for Si uptake in barley roots have been identified (Chiba et al., 2009; Mitani et al., 2009a); however, transporters for Si distribution in aboveground plant tissues are unknown. In this study, we functionally characterized a rice Lsi6 homolog gene in barley, HvLsi6, in terms of transport activity and expression pattern, as well as cellular and subcellular localizations. We found that HvLsi6 is probably involved in Si uptake in the root tip, xylem unloading in the leaf, and intervascular transfer of Si at the nodes in barley. We further found that HvLsi2 was also expressed in the nodes and involved in the intervascular transfer by coupling with HvLsi6.     

A rice phenolic efflux transporter is essential for solubilizing precipitated apoplasmic iron in the plant stele

Iron deficiency is one of the major agricultural problems, as 30% of the arable land of the world is too alkaline for optimal crop production, rendering plants short of available iron despite its abundance. To take up apoplasmic precipitated iron, plants secrete phenolics such as protocatechuic acid (PCA) and caffeic acid. The molecular pathways and genes of iron uptake strategies are already characterized, whereas the molecular mechanisms of phenolics synthesis and secretion have not been clarified, and no phenolics efflux transporters have been identified in plants yet. Here we describe the identification of a phenolics efflux transporter in rice. We identified a cadmium-accumulating rice mutant in which the amount of PCA and caffeic acid in the xylem sap was dramatically reduced and hence named it phenolics efflux zero 1 (pez1). PEZ1 localized to the plasma membrane and transported PCA when expressed in Xenopus laevis oocytes. PEZ1 localized mainly in the stele of roots. In the roots of pez1, precipitated apoplasmic iron increased. The growth of PEZ1 overexpression lines was severely restricted, and these lines accumulated more iron as a result of the high solubilization of precipitated apoplasmic iron in the stele. We show that PEZ1 is responsible for an increase of PCA concentration in the xylem sap and is essential for the utilization of apoplasmic precipitated iron in the stele.     

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.     

A transporter regulating silicon distribution in rice shoots

Rice (Oryza sativa) accumulates very high concentrations of silicon (Si) in the shoots, and the deposition of Si as amorphous silica helps plants to overcome biotic and abiotic stresses. Here, we describe a transporter, Lsi6, which is involved in the distribution of Si in the shoots. Lsi6 belongs to the nodulin-26 intrinsic protein III subgroup of aquaporins and is permeable to silicic acid. Lsi6 is expressed in the leaf sheath and leaf blades as well as in the root tips. Cellular localization studies revealed that Lsi6 is found in the xylem parenchyma cells of the leaf sheath and leaf blades. Moreover, Lsi6 showed polar localization at the side facing toward the vessel. Knockdown of Lsi6 did not affect the uptake of Si by the roots but resulted in disordered deposition of silica in the shoots and increased excretion of Si in the guttation fluid. These results indicate that Lsi6 is a transporter responsible for the transport of Si out of the xylem and subsequently affects the distribution of Si in the leaf.     

OsFRDL1 Is a Citrate Transporter Required for Efficient Translocation of Iron in Rice

Multidrug and toxic compound extrusion (MATE) transporters represent a large family in plants, but their functions are poorly understood. Here, we report the function of a rice (Oryza sativa) MATE gene (Os03g0216700, OsFRDL1), the closest homolog of barley (Hordeum vulgare) HvAACT1 (aluminum [Al]-activated citrate transporter 1), in terms of metal stress (iron [Fe] deficiency and Al toxicity). This gene was mainly expressed in the roots and the expression level was not affected by either Fe deficiency or Al toxicity. Knockout of this gene resulted in leaf chlorosis, lower leaf Fe concentration, higher accumulation of zinc and manganese concentration in the leaves, and precipitation of Fe in the root's stele. The concentration of citrate and ferric iron in the xylem sap was lower in the knockout line compared to the wild-type rice. Heterologous expression of OsFRDL1 in Xenopus oocytes showed transport activity for citrate. Immunostaining showed that OsFRDL1 was localized at the pericycle cells of the roots. On the other hand, there was no difference in the Al-induced secretion of citrate from the roots between the knockout line and the wild-type rice. Taken together, our results indicate that OsFRDL1 is a citrate transporter localized at the pericycle cells, which is necessary for efficient translocation of Fe to the shoot as a Fe-citrate complex.     

The aromatic/arginine selectivity filter of NIP aquaporins plays a critical role in substrate selectivity for silicon, boron, and arsenic

Nodulin-26-like intrinsic proteins (NIPs) of the aquaporin family are involved in the transport of diverse solutes, but the mechanisms controlling the selectivity of transport substrates are poorly understood. The purpose of this study was to investigate how the aromatic/arginine (ar/R) selectivity filter influences the substrate selectivity of two NIP aquaporins; the silicic acid (Si) transporter OsLsi1 (OsNIP2;1) from rice and the boric acid (B) transporter AtNIP5;1 from Arabidopsis; both proteins are also permeable to arsenite. Native and site-directed mutagenized variants of the two genes were expressed in Xenopus oocytes and the transport activities for Si, B, arsenite, and water were assayed. Substitution of the amino acid at the ar/R second helix (H2) position of OsLsi1 did not affect the transport activities for Si, B, and arsenite, but that at the H5 position resulted in a total loss of Si and B transport activities and a partial loss of arsenite transport activity. Conversely, changes of the AtNIP5;1 ar/R selectivity filter and the NPA motifs to the OsLsi1 type did not result in a gain of Si transport activity. B transport activity was partially lost in the H5 mutant but unaffected in the H2 mutant of AtNIP5;1. In contrast, both the single and double mutations at the H2 and/or H5 positions of AtNIP5;1 did not affect arsenite transport activity. The results reveal that the residue at the H5 position of the ar/R filter of both OsLsi1 and AtNIP5;1 plays a key role in the permeability to Si and B, but there is a relatively low selectivity for arsenite.     

Silicon efflux transporters isolated from two pumpkin cultivars contrasting in Si uptake

The accumulation of silicon (Si) differs greatly with plant species and cultivars due to different ability of the roots to take up Si. In Si accumulating plants such as rice, barley and maize, Si uptake is mediated by the influx (Lsi1) and efflux (Lsi2) transporters. Here we report isolation and functional analysis of two Si efflux transporters (CmLsi2-1 and CmLsi2-2) from two pumpkin (Cucurbita moschata Duch.) cultivars contrasting in Si uptake. These cultivars are used for rootstocks of bloom and bloomless cucumber, respectively. Different from mutations in the Si influx transporter CmLsi1, there was no difference in the sequence of either CmLsi2 between two cultivars. Both CmLsi2-1 and CmLsi2-2 showed an efflux transport activity for Si and they were expressed in both the roots and shoots. These results confirm our previous finding that mutation in CmLsi1, but not in CmLsi2-1 and CmLsi2-2 are responsible for bloomless phenotype resulting from low Si uptake.Key words: silicon, efflux transporter, pumpkin, cucumber, bloomSilicon (Si) is the second most abundant elements in earth''s crust.¹ Therefore, all plants rooting in soils contain Si in their tissues. However Si accumulation in the shoot differs greatly among plant species, ranging for 0.1 to 10% of dry weight.^1–3 In higher plants, only Poaceae, Equisetaceae and Cyperaceae show a high Si accumulation.^2,3 Si accumulation also differs with cultivars within a species.^4,5 These differences in Si accumulation have been attributed to the ability of the roots to take up Si.^6,7Genotypic difference in Si accumulation has been used to produce bloomless cucumber (Cucumis sativus L.).⁸ Bloom (white and fine powders) on the surface of cucumber fruits is primarily composed of silica (SiO₂).⁹ However, nowadays, cucumber without bloom (bloomless cucumber) is more popular in Japan due to its more attractive and distinctly shiny appearance. Bloomless cucumber is produced by grafting cucumber on some specific pumpkin (Cucurbita moschata Duch.) cultivars. These pumpkin cultivars used for bloomless cucumber rootstocks have lower silicon accumulation compared with the rootstocks used for producing bloom cucumber.⁹Our study showed that the difference in Si accumulation between bloom and bloomless root stocks of pumpkin cultivars results from different Si uptake by the roots.¹⁰ Si uptake has been demonstrated to be mediated by two different types of transporters (Lsi1 and Lsi2) in rice, barley and maize.^11–15 Lsi1 is an influx transporter of Si, belonging to a NIP subfamily of aquaporin family.^10,11,13,14 This transporter is responsible for transport of Si from external solution to the root cells.¹¹ On the other hand, Lsi2 is an efflux transporter of Si, belonging to putative anion transporter.¹² Lsi2 releases Si from the root cells towards the xylem. Both Lsi1 and Lsi2 are required for Si uptake by the roots.^11,12 To understand the mechanism underlying genotypic difference in Si uptake, we have isolated and functionally characterized an influx Si transporter CmLsi1 from two pumpkin cultivars used for rootstocks of bloomless and bloom cucumber.¹⁰ Sequence analysis showed only two amino acids difference of CmLsi1 between two pumpkin cultivars. However, CmLsi1 from bloom rootstock [CmLsi1(B⁺)] showed transport activity for Si, whereas that from bloomless rootstock [CmLsi1(B⁻)] did not.¹⁰ Furthermore, we found that loss of Si transport activity was caused by one amino acid mutation at the position of 242 (from proline to leucine).¹⁰ This mutation resulted in failure to be localized at the plasma membrane, which is necessary for functioning as an influx transporter. The mutated protein was localized at the ER.¹⁰ Here, we report isolation and expression analysis of Si efflux transporters from two pumpkin cultivars contrasting in Si uptake and accumulation to examine whether Si efflux transporter is also involved in the bloom and bloomless phenotypes.