Characterization of the silicon uptake system and molecular mapping of the silicon transporter gene in rice

Rice (Oryza sativa L. cv Oochikara) is a typical silicon-accumulating plant, but the mechanism responsible for the high silicon uptake by the roots is poorly understood. We characterized the silicon uptake system in rice roots by using a low-silicon rice mutant (lsi1) and wild-type rice. A kinetic study showed that the concentration of silicon in the root symplastic solution increased with increasing silicon concentrations in the external solution but saturated at a higher concentration in both lines. There were no differences in the silicon concentration of the symplastic solution between the wild-type rice and the mutant. The form of soluble silicon in the root, xylem, and leaf identified by (29)Si-NMR was also the same in the two lines. However, the concentration of silicon in the xylem sap was much higher in the wild type than in the mutant. These results indicate that at least two transporters are involved in silicon transport from the external solution to the xylem and that the low-silicon rice mutant is defective in loading silicon into xylem rather than silicon uptake from external solution to cortical cells. To map the responsible gene, we performed a bulked segregant analysis by using both microsatellite and expressed sequence tag-based PCR markers. As a result, the gene was mapped to chromosome 2, flanked by microsatellite marker RM5303 and expressed sequence tag-based PCR marker E60168.     

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.     

OsNRAMP1 transporter contributes to cadmium and manganese uptake in rice

Rice is a major dietary source of the toxic metal, cadmium (Cd). Previous studies reported that the rice transporter, OsNRAMP1, (Natural resistance-associated macrophage protein 1) could transport iron (Fe), Cd and arsenic (As) in heterologous yeast assays. However, the in planta function of OsNRAMP1 remains unknown. Here, we showed that OsNRAMP1 was able to transport Cd and manganese (Mn) when expressed in yeast, but did not transport Fe or As. OsNRAMP1 was mainly expressed in roots and leaves and encoded a plasma membrane-localized protein. OsNRAMP1 expression was induced by Cd treatment and Fe deficiency. Immunostaining showed that OsNRAMP1 was localized in all root cells, except the central vasculature, and in leaf mesophyll cells. The knockout of OsNRAMP1 resulted in significant decreases in root uptake of Cd and Mn and their accumulation in rice shoots and grains, and increased sensitivity to Mn deficiency. The knockout of OsNRAMP1 had smaller effects on Cd and Mn uptake than knockout of OsNRAMP5, while knockout of both genes resulted in large decreases in the uptake of the two metals. Taken together, OsNRAMP1 contributes significantly to the uptake of Mn and Cd in rice, and the functions of OsNRAMP1 and OsNRAMP5 are similar but not redundant.     

Interactive effects of potassium and sodium on root growth and expression of K/Na transporter genes in rice

Sufficient supply of potassium (K) can alleviate the adverse effects of excess sodium (Na) on plant growth. However, it remains unclear if such a beneficial function is related to regulation of root growth and/or expression of K/Na transporters. Herein we report the responses of a rice cultivar, which was pretreated with normal nutrient solution for 1 month, to three levels of Na (0, 25, and 100 mM) without or with supply of K for 9 days. High Na (100 mM) significantly decreased plant growth, root activity, and total K uptake, and increased biomass ratio of roots to shoots. Short-term removal of K supply (9 days) did not affect root morphology and biomass ratio of roots to shoots, but decreased root activity of seedlings grown in high Na solution. K deficiency increased uptake of Na and transport of K from roots to shoots. Moreover, expression of OsHAK1, a putative K transporter gene, was upregulated by low Na (25 mM) and downregulated by high Na (100 mM) in roots. In leaves, its expression was suppressed by the Na treatments when K supply was maintained. Expression of OsHKT2;1, which encodes a protein that acts mainly as a Na transporter, was downregulated by high Na, but was enhanced by K deficiency both in roots and leaves. Expression of five other putative K/Na transporter or Na⁺/H⁺ genes, OsHKT1;1, OsHKT1;2, OsHKT2;3, OsNHX1, and OsSOS1, was not affected by the treatments. The results suggest that OsHAK1 and OsHKT2;1 were involved in the interactive effects of K and Na on their uptake and distribution in rice. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Lanthanum reduces the cadmium accumulation by suppressing expression of transporter genes involved in cadmium uptake and translocation in wheat

Plant and Soil - Lanthanum (La) is attracting increasing attention due to its effects in enhancing yields and reducing cadmium (Cd) accumulation in crops. Although a few transporter genes are...     

Silicon suppresses zinc uptake through down-regulating zinc transporter gene in rice

One of the beneficial effects of silicon (Si) is to improve nutrient imbalance including deficiency and excess of nutrients, however the molecular mechanisms underlying this effect are still poorly understood. In this study, we investigated the interaction between Si and zinc (Zn) in rice by using a mutant (lsi1) defective in Si uptake and its wild-type (WT, cv. Oochikara) at different Zn levels. High Zn inhibited the root elongation of both WT and lsi1 mutant, but Si did not alleviate this inhibition in both lines. By contrast, Si supply decreased Zn concentration in both the roots and shoots of the WT, but not in the lsi1 mutant. A short-term (24 h) labeling experiment with stable isotope ⁶⁷Zn showed that Si decreased ⁶⁷Zn uptake, but did not affect the root-to-shoot translocation and distribution ratio to different organs of ⁶⁷Zn in the WT. Furthermore, Si accumulated in the shoots, rather than Si in the external solution, is required for suppressing Zn uptake, but this was not caused by Si-decreased transpiration. A kinetic study showed that Si did not affect K_m value of root Zn uptake, but decreased V_max value in the WT. Analysis of genes related with Zn transport showed that among ZIP family genes, the expression of only OsZIP1 implicated in Zn uptake, was down-regulated by Si in the WT, but not in the lsi1 mutant. These results indicate that Si accumulated in the shoots suppresses the Zn uptake through down-regulating the transporter gene involved in Zn uptake in rice.     

A method for expression cloning of transporter genes by screening yeast for uptake of radiolabelled substrate

A method has been developed for the cloning of plasma membrane transporters by screening yeast transformed with a cDNA library for the accumulation of radiolabelled substrate. The applicability of the method is demonstrated by cloning the amino acid permease AAP1. A yeast mutant defective in proline uptake was transformed with an Arabidopsis thaliana cDNA library and plated on medium supplemented with L-[U-(14)C]proline. Yeast colonies accumulating radiolabelled proline were identified by autoradiography. The plasmids of these colonies were reintroduced into the yeast mutant and restoration of proline uptake was confirmed by L-[U-(14)C]proline uptake measurements. Whereas cloning of transporters by functional complementation requires that the substrate taken up is metabolized by yeast to promote growth, the method described here can be used to isolate transporters of substrates which are not metabolized. The method has great potential for the isolation of transporters of various substrates such as secondary plant products.     

Spatial distribution and temporal variation of the rice silicon transporter Lsi1

Rice (Oryza sativa) is a typical silicon (Si) accumulator and requires a large amount of Si for high-yield production. Recently, a gene (Low silicon rice1 [Lsi1]) encoding a Si transporter was identified in rice roots. Here, we characterized Lsi1 in terms of spatial distribution and temporal variation using both physiological and molecular approaches. Results from a multicompartment transport box experiment showed that the major site for Si uptake was located at the basal zone (>10 mm from the root tip) of the roots rather than at the root tips (<10 mm from the root tip). Consistent with the Si uptake pattern, Lsi1 expression and distribution of the Lsi1 protein were found only in the basal zone of roots. In the basal zones of the seminal, crown, and lateral roots, the Lsi1 protein showed a polar localization at the distal side of both the exodermis and endodermis, where the Casparian bands are formed. This indicates that Lsi1 is required for the transport of Si through the cells of the exodermis and endodermis. Expression of Lsi1 displayed a distinct diurnal pattern. Furthermore, expression was transiently enhanced around the heading stage, which coincides with a high Si requirement during this growth stage. Expression was down-regulated by dehydration stress and abscisic acid, suggesting that expression of Lsi1 may be regulated by abscisic acid.     

Effect of silicon on the growth and phosphorus uptake of rice

A pot experiment was conducted to measure the effect of silicon on phosphorus uptake and on the growth of rice at different P levels. Rice (Oryza sativa L. cv. Akebono) was cultured in Kimura B nutrient solution without and with silicon (1.66 mM Si) and with three phosphorus levels (0.014 mM P, low; 0.21 mM, medium; and 0.70 mM, high).Shoot dry weight with Si (+Si) in solution increased with increasing P level, while shoot weight without Si (–Si) was maximum at 0.21 mM P, suggesting that +Si raised the optimum P level for rice. +Si increased shoot weight more when P was low or high than when P was medium.The concentration and amount of inorganic P in shoots increased with increasing P level. +Si did not significantly decrease P uptake by rice at 0.014 mM P, however, uptake at 0.21 and 0.70 mM P was 27 and 30 percent less than uptake with –Si, respectively. In –Si with 0.21 and 0.70 mM P, inorganic P in shoots was more than double the concentration in shoots grown in +Si solutions.The Si concentration in shoots decreased slightly with increasing P level, although Si uptake was not significantly affected by P. +Si decreased the uptake of Fe and Mn by an average of 20 and 50 percent, respectively, thus P/Mn and P/Fe ratios increased in the shoot when P was low.From the results above, the beneficial effect of Si on the growth of rice was clearly shown when P was low or high. This effect may have resulted from decreased Mn and Fe uptake, and thus increased P availability within P deficient plants, or from reduced P uptake when P was high.     

Overexpression of rice phosphate transporter gene OsPT6 enhances phosphate uptake and accumulation in transgenic rice plants

Background and aims

Whereas the expression patterns and kinetic properties of the rice (Oryza sativa) phosphate transporter gene OsPht1; 6 (OsPT6) are well documented, little is known about the biological functions of this gene. The aim of this study was to investigate the roles of OsPT6 in inorganic phosphate (Pi) acquisition and mobilization, and examine its potential to enhance agricultural production.

Methods

Here, we generated OsPT6 overexpression transgenic plants using Wuyujing 7, a widely cultivated variety of japonica rice, and then treated transgenic lines and wild type with different Pi supply in hydroponic and soil experiments to explore the functions of OsPT6 in rice.

Results

The OsPT6-overexpressing rice lines grew better and accumulated more biomass than wild-type plants, and exhibited significant increases in P accumulation in various tissues, including reproductive tissues under both hydroponic and soil culture conditions. Phosphate-uptake experiment using radiolabeled Pi (³³P) showed that the rate of Pi uptake was 75 % and 73 % greater in transgenic plants grown under Pi-sufficient and -deficient conditions, respectively, than the wild-type controls, and that the shoot/root ratio of ³³P was 104 % and 42 % greater, respectively. In addition, the grain yield per transgenic plant was much higher than that of the wild-type plants under field conditions.

Conclusions

Taken together, our results demonstrate that OsPT6 plays a vital role in Pi acquisition and mobilization in rice and suggest that this gene may be used for genetic engineering rice plants that require less Pi fertilizer.     

Role of the iron transporter OsNRAMP1 in cadmium uptake and accumulation in rice

The heavy metal cadmium (Cd) is toxic to humans, and its accumulation in rice grains is a major agricultural problem. Rice has seven putative metal transporter NRAMP genes, but microarray analysis showed that only OsNRAMP1 is highly up-regulated by iron (Fe) deficiency. OsNRAMP1 localized to the plasma membrane and transported Cd as well as Fe. OsNRAMP1 expression was observed mainly in roots and was higher in the roots of a high-Cd-accumulating cultivar (Habataki) than in those of a low-Cd-accumulating cultivar (Sasanishiki). The amino acid sequence of OsNRAMP1 in the Sasanishiki and Habataki cultivars was found to be 100% identical. These results suggest that OsNRAMP1 participates in cellular Cd uptake and that the differences observed in Cd accumulation among cultivars are because of differences in OsNRAMP1 expression levels in roots.     

不同硅吸收效率水稻品种根系对硅素水平的响应

为明确硅对水稻根系生长发育的影响,以4个硅吸收效率不同的水稻品种（高效吸收品种TN1、白香粳和低效吸收品种卷叶粳、一目惚）为材料,采用国际水稻研究所的营养液配方水培试验,设置0(T₁)、1.25 (T₂)和2 (T₃) mmol·L^-1 3个硅素水平,研究了不同硅素水平对不同基因型水稻根系和地上部干物质量、根条数、侧根数、根总长和根直径等的影响.结果表明：随硅素水平的提高,水稻各品种均表现为根系干物质量、根冠比、侧根数和根总长逐渐减少,地上部干物质量、根条数和根直径逐渐增大.较高的硅素水平有利于水稻不定根的分化发育,而不利于侧根的分化发育.在较低的硅素水平下,硅吸收效率高的基因型水稻TN1和白香粳的根干物质量和根冠比显著高于硅吸收效率低的品种卷叶粳和一目惚,其中白香粳的侧根数和根总长均显著高于卷叶粳和一目惚.可见,根总长和侧根数是引起水稻硅素吸收差异的主要原因.     

A high activity zinc transporter OsZIP9 mediates zinc uptake in rice

Zinc (Zn) is an essential micronutrient for most organisms including humans, and Zn deficiency is widespread in human populations, particularly in underdeveloped regions. Cereals such as rice (Oryza sativa) are the major dietary source of Zn for most people. However, the molecular mechanism underlying Zn uptake in rice is still not fully understood. Here, we report that a member of the ZIP (ZRT, IRT‐like protein) family, OsZIP9, contributes to Zn uptake in rice. It was expressed in the epidermal and exodermal cells of lateral roots, localized in the plasma membrane and induced during Zn deficiency. Yeast‐expressed OsZIP9 showed much higher Zn influx transport activity than other rice ZIP proteins in a wide range of Zn concentrations. OsZIP9 knockout rice plants showed a significant reduction in growth at low Zn concentrations, but could be rescued by a high Zn supply. Compared with the wild type, accumulation of Zn in root, shoot and grain was much lower in knockout lines, particularly with a low supply of Zn under both hydroponic and paddy soil conditions. OsZIP9 also showed Co uptake activity. Natural variation of OsZIP9 expression level is highly associated with Zn content in milled grain among rice varieties in the germplasm collection. Taken together, these results show that OsZIP9 is an important influx transporter responsible for the take up of Zn and Co from external media into root cells.     

Genotypic difference in canopy diffusive conductance measured by a new remote-sensing method and its association with the difference in rice yield potential

There have been few practical ways of measuring physiological determinants of rice yield. Rapid evaluation of yield determination traits may expedite breeding of high-yielding rice. Here, we report a new remote-sensing technique for the evaluation of canopy ecophysiological status under field conditions developed based on simultaneous measurements of sunlit and suddenly shaded canopy temperatures. This technique has the advantage of instantaneous estimation of aerodynamic resistance (r(a)) and canopy diffusive resistance (r(c) without measuring wind velocity. Canopy diffusive conductance (1 / r(c)) estimated by the remote sensing method was closely related to leaf stomatal conductance (g(s)) measured with a portable gas exchange system. This result supported the validity of this new method for quantitative estimation of canopy physiological characteristics. Significant genotypic differences were obtained in canopy-air temperature difference (Tc-Ta), r(c) and 1 / r(c) during the 2-week period preceding full heading for two years, and 1 / r(c) was highly correlated with crop growth rate (CGR), which was closely related to the final yield. These results suggest that 1 / r(c) can be an effective criterion for the selection of high-yielding rice genotypes, and the remote sensing technique proposed here can be a powerful tool for the rapid evaluation of 1 / r(c) under field conditions.     

Alteration of nutrient allocation and transporter genes expression in rice under N,P, K,and Mg deficiencies

Nitrogen (N), phosphorus (P), potassium (K), and magnesium (Mg) have essential physiological functions in plants. Their interactions in plants are not fully understood especially at the molecular level. In this study, we detected the physiological and molecular responses of rice plants at the vegetative growth phase to N, P, K, and Mg starvations. Deficiencies of N and P resulted in accumulation of soluble sugar and starch in the leaves. The root to shoot ratio increased under N and P deficiencies, but decreased under K and Mg deficiencies. In addition, deficiency of either K or Mg resulted in accumulation of the other cation in shoots. Moreover, K starvation decreased both K and soluble sugar contents in the roots pronouncedly. RT-PCR analysis showed that several sugar transporter genes in the leaves orchestrated with sugar accumulation induced by the nutrient shortages. Expression of a high affinity K transporter gene (OsHAK1) and a putative Mg transporter gene (OsMGT) showed opposite down- and up-regulation in the roots by K starvation. These findings suggest that deficiencies of the major nutrients suppressed the export of carbohydrates from source leaves. The regulated sugar and nutrient transporter genes investigated in this study could be used for elucidating the molecular mechanism of plants in their adaptation to varied nutrient supply.