The boron transporter BnaC4.BOR1;1c is critical for inflorescence development and fertility under boron limitation in Brassica napus

Boron (B) is an essential micronutrient for plants, but the molecular mechanisms underlying the uptake and distribution of B in allotetraploid rapeseed (Brassica napus) are unclear. Here, we identified a B transporter of rapeseed, BnaC4.BOR1;1c, which is expressed in shoot nodes and involved in distributing B to the reproductive organs. Transgenic Arabidopsis plants containing a BnaC4.BOR1;1c promoter‐driven GUS reporter gene showed strong GUS activity in roots, nodal regions of the shoots and immature floral buds. Overexpressing BnaC4.BOR1;1c in Arabidopsis wild type or in bor1‐1 mutants promoted wild‐type growth and rescued the bor1‐1 mutant phenotype. Conversely, knockdown of BnaC4.BOR1;1c in a B‐efficient rapeseed line reduced B accumulation in flower organs, eventually resulting in severe sterility and seed yield loss. BnaC4.BOR1;1c RNAi plants exhibited large amounts of disintegrated stigma papilla cells with thickened cell walls accompanied by abnormal proliferation of lignification under low‐B conditions, indicating that the sterility may be a result of altered cell wall properties in flower organs. Taken together, our results demonstrate that BnaC4.BOR1;1c is a AtBOR1‐homologous B transporter gene expressing in both roots and shoot nodes that is essential for the developing inflorescence tissues, which highlights its diverse functions in allotetraploid rapeseed compared with diploid model plant Arabidopsis.     

Improvement of seed yields under boron-limiting conditions through overexpression of BOR1, a boron transporter for xylem loading, in Arabidopsis thaliana

Soil fertilization is a common practice in modern agriculture, undertaken to prevent nutrient deficiency in crops. However, fertilization is costly and causes environmental pollution. The cultivation of plants that tolerate low nutrient supplies may circumvent this problem. Here, we report the generation of Arabidopsis thaliana plants that tolerate boron (B)-deficient conditions due to the overexpression of BOR1, an efflux B transporter that is required for efficient xylem loading of B. In several independently generated transgenic plants expressing BOR1 or BOR1-GFP under the control of the cauliflower mosaic virus 35S RNA promoter, root-to-shoot translocation of B was enhanced and shoot growth was greater under B-limiting conditions compared with wild-type plants. In addition, the transgenic plants showed increased translocation of B, especially to the shoot apex, and set seed normally under B-limiting conditions, under which wild-type plants failed to set seed. This study therefore reports plants that show improved seed yields compared with wild-type under nutrient-deficient conditions as a result of increased production of an essential mineral nutrient transporter.     

The effect of boron availability,CO<Subscript>2</Subscript>, and irradiance on relative accumulation of the major boron transport proteins,BOR1 and NIP5;1

Boron (B) is an essential plant micronutrient. Two major B-transport proteins have been recently identified and partially characterized: BOR1, a high-affinity B efflux transporter involved in xylem loading, and NIP5;1, a plasma-membrane boric-acid channel involved in B uptake. To date, studies of these B transporters have investigated their expression individually (mainly as mRNA), and only in response to variation in B availability (mostly B deficiency); the influence of other factors, such as plant resource status, has not been studied. To address this, we grew geranium (Pelargonium × hortorum cv. Maverick White) plants under ambient or elevated CO₂ concentration, different sub-saturating irradiances, and different B availability. For comparison we also grew three other species (Arabidopsis thaliana, Azolla caroliniana, and Hordeum vulgare) under broad range of B supply. Relative accumulation of BOR1 and NIP5;1 proteins were measured using protein-specific antibodies and Western blotting or ELISA. Elevated CO₂ significantly increased content of NIP5;1, while increases in irradiance increased BOR1 content, but decreased NIP5;1 content. Across species, content of both transporters often decreased with increasing B availability, but sometimes remained unchanged or even increased, depending on CO₂, irradiance, species, or transporter. Content of BOR1 and NIP5;1 was correlated with root proteins, B content, and sugar content (for high CO₂ only), as well as B uptake, but CO₂ and irradiance often affected these relationships. Thus, relative accumulation of BOR1 and NIP5;1 is influenced not only by B content, as expected, but by other environmental factors as well.     

Increased expression of OsPT1, a high-affinity phosphate transporter, enhances phosphate acquisition in rice

Most high-affinity phosphate transporter genes (OsPTs) in rice were highly induced in roots when phosphate was depleted. OsPT1, however, was highly expressed in primary roots and leaves regardless of external phosphate concentrations. This finding was confirmed histochemically using transgenic rice plants that express the GUS reporter gene under the control of the OsPT1 promoter, which exhibited high GUS activity even in the phosphate sufficient condition. Furthermore, transgenic rice plants overexpressing the OsPT1 gene accumulated almost twice as much phosphate in the shoots as did wild-type plants. As a result, transgenic plants had more tillers than did wild-type plants, which is a typical physiological indicator for phosphate status in rice.     

Auxin and ethylene are involved in the responses of root system architecture to low boron supply in Arabidopsis seedlings

Changes in root architecture are one of the adaptive strategies used by plants to compensate for nutrient deficiencies in soils. In this work, the temporal responses of Arabidopsis (Arabidopsis thaliana) root system architecture to low boron (B) supply were investigated. Arabidopsis Col-0 seedlings were grown in 10 μM B for 5 days and then transferred to a low B medium (0.4 μM) or control medium (10 μM) for a 4-day period. Low B supply caused an inhibition of primary root (PR) growth without altering either the growth or number of lateral roots (LRs). In addition, low B supply induced root hair formation and elongation in positions close to the PR meristem not observed under control conditions. The possible role of auxin and ethylene in the alteration of root system architecture elicited by low B supply was also studied by using two Arabidopsis reporter lines (DR5:GUS and EBS:GUS) and two Arabidopsis mutants with impaired auxin and ethylene signaling (aux1-22 and ein2-1). Low B supply increased auxin reporter DR5:GUS activity in PR tip, suggesting that low B alters the pattern of auxin distribution in PR tip. Moreover, PR elongation in aux1-22 mutant was less sensitive to low B treatment than in wild-type plants, which suggests that auxin resistant 1 (AUX1) participates in the inhibition of PR elongation under low B supply. From all these results, a hypothetical model to explain the effect of low B treatment on PR growth is proposed. We also show that ethylene, via ethylene-insensitive 2 (EIN2) protein, is involved in the induction of root hair formation and elongation under low B treatment.     

Two rice phosphate transporters, OsPht1;2 and OsPht1;6, have different functions and kinetic properties in uptake and translocation

Plant phosphate (Pi) transporters mediate the uptake and translocation of this nutrient within plants. A total of 13 sequences in the rice ( Oryza sativa ) genome can be identified as belonging to the Pi transporter (Pht1) family. Here, we report on the expression patterns, biological properties and the physiological roles of two members of the family: OsPht1;2 ( OsPT2 ) and OsPht1;6 ( OsPT6 ). Expression of both genes increased significantly under Pi deprivation in roots and shoots. By using transgenic rice plants expressing the GUS reporter gene, driven by their promoters, we detected that OsPT2 was localized exclusively in the stele of primary and lateral roots, whereas OsPT6 was expressed in both epidermal and cortical cells of the younger primary and lateral roots. OsPT6, but not OsPT2, was able to complement a yeast Pi uptake mutant in the high-affinity concentration range. Xenopus oocytes injected with OsPT2 mRNA showed increased Pi accumulation and a Pi-elicited depolarization of the cell membrane electrical potential, when supplied with mM external concentrations. Both results show that OsPT2 mediated the uptake of Pi in oocytes. In transgenic rice, the knock-down of either OsPT2 or OsPT6 expression by RNA interference significantly decreased both the uptake and the long-distance transport of Pi from roots to shoots. Taken together, these data suggest OsPT6 plays a broad role in Pi uptake and translocation throughout the plant, whereas OsPT2 is a low-affinity Pi transporter, and functions in translocation of the stored Pi in the plant.     

Expression of the Arabidopsis borate efflux transporter gene, AtBOR4, in rice affects the xylem loading of boron and tolerance to excess boron

Boron is an essential nutrient for plants, but it is toxic in excess. Transgenic rice plants expressing an Arabidopsis thaliana borate efflux transporter gene, AtBOR4, at a low level exhibited increased tolerance to excess boron. Those lines with high levels of expression exhibited reduced growth. These findings suggest a potential of the borate transporter BOR4 for the generation of high-boron tolerant rice.     

Rice OsHKT2;1 transporter mediates large Na+ influx component into K+-starved roots for growth

Excessive accumulation of sodium in plants causes toxicity. No mutation that greatly diminishes sodium (Na+) influx into plant roots has been isolated. The OsHKT2;1 (previously named OsHKT1) transporter from rice functions as a relatively Na+-selective transporter in heterologous expression systems, but the in vivo function of OsHKT2;1 remains unknown. Here, we analyzed transposon-insertion rice lines disrupted in OsHKT2;1. Interestingly, three independent oshkt2;1-null alleles exhibited significantly reduced growth compared with wild-type plants under low Na+ and K+ starvation conditions. The mutant alleles accumulated less Na+, but not less K+, in roots and shoots. OsHKT2;1 was mainly expressed in the cortex and endodermis of roots. (22)Na+ tracer influx experiments revealed that Na+ influx into oshkt2;1-null roots was dramatically reduced compared with wild-type plants. A rapid repression of OsHKT2;1-mediated Na+ influx and mRNA reduction were found when wild-type plants were exposed to 30 mM NaCl. These analyses demonstrate that Na+ can enhance growth of rice under K+ starvation conditions, and that OsHKT2;1 is the central transporter for nutritional Na+ uptake into K+-starved rice roots.     

A constitutive expressed phosphate transporter, OsPht1;1, modulates phosphate uptake and translocation in phosphate-replete rice

A number of phosphate (Pi) starvation- or mycorrhiza-regulated Pi transporters belonging to the Pht1 family have been functionally characterized in several plant species, whereas functions of the Pi transporters that are not regulated by changes in Pi supply are lacking. In this study, we show that rice (Oryza sativa) Pht1;1 (OsPT1), one of the 13 Pht1 Pi transporters in rice, was expressed abundantly and constitutively in various cell types of both roots and shoots. OsPT1 was able to complement the proton-coupled Pi transporter activities in a yeast mutant defective in Pi uptake. Transgenic plants of OsPT1 overexpression lines and RNA interference knockdown lines contained significantly higher and lower phosphorus concentrations, respectively, compared with the wild-type control in Pi-sufficient shoots. These responses of the transgenic plants to Pi supply were further confirmed by the changes in depolarization of root cell membrane potential, root hair occurrence, (33)P uptake rate and transportation, as well as phosphorus accumulation in young leaves at Pi-sufficient levels. Furthermore, OsPT1 expression was strongly enhanced by the mutation of Phosphate Overaccumulator2 (OsPHO2) but not by Phosphate Starvation Response2, indicating that OsPT1 is involved in the OsPHO2-regulated Pi pathway. These results indicate that OsPT1 is a key member of the Pht1 family involved in Pi uptake and translocation in rice under Pi-replete conditions.     

Expression in Arabidopsis and cellular localization reveal involvement of rice NRAMP,OsNRAMP1, in arsenic transport and tolerance

Irrigation of paddy fields to arsenic (As) containing groundwater leads to As accumulation in rice grains and causes serious health risk to the people worldwide. To reduce As intake via consumption of contaminated rice grain, identification of the mechanisms for As accumulation and detoxification in rice is a prerequisite. Herein, we report involvement of a member of rice NRAMP (Natural Resistance‐Associated Macrophage Protein) transporter, OsNRAMP1, in As, in addition to cadmium (Cd), accumulation through expression in yeast and Arabidopsis. Expression of OsNRAMP1 in yeast mutant (fet3fet4) rescued iron (Fe) uptake and exhibited enhanced accumulation of As and Cd. Expression of OsNRAMP1 in Arabidopsis provided tolerance with enhanced As and Cd accumulation in root and shoot. Cellular localization revealed that OsNRAMP1 resides on plasma membrane of endodermis and pericycle cells and may assist in xylem loading for root to shoot mobilization. This is the first report demonstrating role of NRAMP in xylem mediated loading and enhanced accumulation of As and Cd in plants. We propose that genetic modification of OsNRAMP1 in rice might be helpful in developing rice with low As and Cd content in grain and minimize the risk of food chain contamination to these toxic metals.     

Interactive effects of mycorrhization and elevated atmospheric CO2 on sulphur nutrition of young pedunculate oak (Quercus robur L.) trees

Pedunculate oak (Quercus robur L.) was germinated and grown at ambient CO₂ concentration and 650 μmol mol^?1 CO₂ in the presence and absence of the ectomycorrhizal fungus Laccaria laccata for a total of 22 weeks under nonlimiting nutrient conditions. Sulphate uptake, xylem loading and exudation were analysed in excised roots. Despite a relatively high affinity for sulphate (K_M= 1.6 mmol m^?3), the rates of sulphate uptake by excised lateral roots of mycorrhizal oak trees were low as compared to herbaceous plants. Rates of sulphate uptake were similar in mycorrhizal and non-mycorrhizal roots and were not affected by growth of the trees at elevated CO₂. However, the total uptake of sulphate per plant was enhanced by elevated CO₂ and further enhanced by elevated CO₂ and mycorrhization. Sulphate uptake seemed to be closely correlated with biomass accumulation under the conditions applied. The percentage of the sulphate taken up by mycorrhizal oak roots that was loaded into the xylem was an order of magnitude lower than previously observed for herbaceous plants. The rate of xylem loading was enhanced by mycorrhization and, in roots of mycorrhizal trees only, by growth at elevated CO₂. On a whole-plant basis this increase in xylem loading could only partially be explained by the increased growth of the trees. Elevated CO₂ and mycorrhization appeared to increase greatly the sulphate supply of the shoot at the level of xylem loading. For all treatments, calculated rates of sulphate exudation were significantly lower than the corresponding rates of xylem loading of sulphate. Radiolabelled sulphate loaded into the xylem therefore seems to be readily diluted by unlabelled sulphate during xylem transport. Allocation of reduced sulphur from oak leaves was studied by flap-feeding radiolabelled GSH to mature oak leaves. The rate of export of radioactivity from the fed leaves was 4–5 times higher in mycorrhizal oak trees grown at elevated CO₂ than in those grown at ambient CO₂. Export of radiolabel proceeded almost exclusively in a basipetal direction to the roots. From these experiments it can be concluded that, in mycorrhizal oak trees grown at elevated CO₂, the transport of sulphate to the shoot is increased at the level of xylem loading to enable increased sulphate reduction in the leaves. Increased sulphate reduction seems to be required for the enhanced allocation of reduced sulphur to the roots which is observed in trees grown at elevated CO₂. These changes in sulphate and reduced sulphur allocation may be a prerequisite for the positive effect of elevated CO₂ on growth of oak trees previously observed.     

植物硼营养研究的重要进展与展望

硼是高等植物必需的矿质营养元素,但是人们对硼行使生理功能及其分子机理的认识远落后于其它必需营养元素。近几年国际上植物硼营养的研究取得了一些重要的突破。首先是进一步明确了B-RG-II复合物的形成及其影响细胞壁结构和功能的分子机制,并且发现B-RG-II的形成及其含量与陆生植物的进化密切相关。其次是在拟南芥中克隆了第一个植物硼转运子基因BOR1,并揭示了它的作用机理;通过转基因实验证明了植物硼的高效吸收与水通道基因NIP5;1密切相关。进而通过大量的种质筛选,从油菜、小麦、大麦及棉花等农作物中获得一批硼高效吸收利用的优异种质材料,并开展了硼高效QTL定位和克隆。本文详细综述了以上几个方面的研究进展,并对进一步的研究做了展望。     

植物硼营养研究的重要进展与展望

硼是高等植物必需的矿质营养元素, 但是人们对硼行使生理功能及其分子机理的认识远落后于其它必需营养元素。近几年国际上植物硼营养的研究取得了一些重要的突破。首先是进一步明确了B-RG-II复合物的形成及其影响细胞壁结构和功能的分子机制, 并且发现B-RG-II的形成及其含量与陆生植物的进化密切相关。其次是在拟南芥中克隆了第一个植物硼转运子基因 BOR1, 并揭示了它的作用机理; 通过转基因实验证明了植物硼的高效吸收与水通道基因NIP5;1密切相关。进而通过大量的种质筛选, 从油菜、小麦、大麦及棉花等农作物中获得一批硼高效吸收利用的优异种质材料, 并开展了硼高效QTL定位和克隆。本文详细综述了以上几个方面的研究进展, 并对进一步的研究做了展望。     

AtNRAMP3, a multispecific vacuolar metal transporter involved in plant responses to iron deficiency

Metal homeostasis is critical for the survival of living organisms, and metal transporters play central roles in maintaining metal homeostasis in the living cells. We have investigated the function of a metal transporter of the NRAMP family, AtNRAMP3, in Arabidopsis thaliana. A previous study showed that AtNRAMP3 expression is upregulated by iron (Fe) starvation and that AtNRAMP3 protein can transport Fe. In the present study, we used AtNRAMP3 promoter beta-glucoronidase (GUS) fusions to show that AtNRAMP3 is expressed in the vascular bundles of roots, stems, and leaves under Fe-sufficient conditions. This suggests a function in long-distance metal transport within the plant. Under Fe-starvation conditions, the GUS activity driven by the AtNRAMP3 promoter is upregulated without any change in the expression pattern. We analyze the impact of AtNRAMP3 disruption and overexpression on metal accumulation in plants. Under Fe-sufficient conditions, AtNRAMP3 overexpression or disruption does not lead to any change in the plant metal content. Upon Fe starvation, AtNRAMP3 disruption leads to increased accumulation of manganese (Mn) and zinc (Zn) in the roots, whereas AtNRAMP3 overexpression downregulates Mn accumulation. In addition, overexpression of AtNRAMP3 downregulates the expression of the primary Fe uptake transporter IRT1 and of the root ferric chelate reductase FRO2. Expression of AtNRAMP3::GFP fusion protein in onion cells or Arabidopsis protoplasts shows that AtNRAMP3 protein localizes to the vacuolar membrane. To account for the results presented, we propose that AtNRAMP3 influences metal accumulation and IRT1 and FRO2 gene expression by mobilizing vacuolar metal pools to the cytosol.