Route and Regulation of Zinc, Cadmium, and Iron Transport in Rice Plants (Oryza sativa L.) during Vegetative Growth and Grain Filling: Metal Transporters, Metal Speciation, Grain Cd Reduction and Zn and Fe Biofortification.

Zinc (Zn) and iron (Fe) are essential but are sometimes deficient in humans, while cadmium (Cd) is toxic if it accumulates in the liver and kidneys at high levels. All three are contained in the grains of rice, a staple cereal. Zn and Fe concentrations in rice grains harvested under different levels of soil/hydroponic metals are known to change only within a small range, while Cd concentrations show greater changes. To clarify the mechanisms underlying such different metal contents, we synthesized information on the routes of metal transport and accumulation in rice plants by examining metal speciation, metal transporters, and the xylem-to-phloem transport system. At grain-filling, Zn and Cd ascending in xylem sap are transferred to the phloem by the xylem-to-phloem transport system operating at stem nodes. Grain Fe is largely derived from the leaves by remobilization. Zn and Fe concentrations in phloem-sap and grains are regulated within a small range, while Cd concentrations vary depending on xylem supply. Transgenic techniques to increase concentrations of the metal chelators (nicotianamine, 2'-deoxymugineic acid) are useful in increasing grain Zn and Fe concentrations. The elimination of OsNRAMP5 Cd-uptake transporter and the enhancement of root cell vacuolar Cd sequestration reduce uptake and root-to-shoot transport, respectively, resulting in a reduction of grain Cd accumulation.

Concentrations of metals and potential metal-binding compounds and speciation of Cd, Zn and Cu in phloem and xylem saps from castor bean plants (Ricinus communis) treated with four levels of cadmium.

We examined the concentrations of metals (Cd, Zn, Cu, Fe and Mn) and potential metal-binding compounds [nicotianamine (NA), thiol compounds and citrate] in xylem and phloem saps from 4-week-old castor bean plants (Ricinus communis) treated with 0 (control), 0.1, 1.0, and 10 µM Cd for 3 weeks. Treatment with 0.1 and 1 µM Cd produced no visible damage, while 10 µM Cd retarded growth. Cadmium concentrations in both saps were higher than those in the culture solution at 0.1 µM, similar at 1.0 µM and lower at 10 µM. Cd at 10 µM reduced Cu and Fe concentrations in both saps. NA concentrations measured by capillary electrophoresis-mass spectrometry (MS) in xylem sap (20 µM) were higher than the Cu concentrations, and those in phloem sap (150 µM) were higher than those of Zn, Fe and Cu combined. Reduced glutathione concentrations differed in xylem and phloem saps (1-2 and 30-150 µM, respectively), but oxidized glutathione concentrations were similar. Phloem sap phytochelatin 2 concentration increased from 0.8 µM in controls to 8 µM in 10 µM Cd. Free citrate was 2-4 µM in xylem sap and 70-100 µM in phloem sap. Total bound forms of Cd in phloem and xylem saps from 1 µM Cd-treated plants were 54 and 8%, respectively. Treatment of phloem sap with proteinaseK reduced high-molecular compounds while increasing fractions of low-molecular Cd-thiol complexes. Zinc-NA, Fe-NA and Cu-NA were identified in the phloem sap fraction of control plants by electrospray ionization time-of-flight MS, and the xylem sap contained Cu-NA.

Nano scale proteomics revealed the presence of regulatory proteins including three FT-Like proteins in phloem and xylem saps from rice.

The main physiological roles of phloem and xylem in higher plants involve the transport of water, nutrients and metabolites. They are also involved, however, in whole plant events including stress responses and long-distance signaling. Phloem and xylem saps therefore include a variety of proteins. In this study, we have performed a shotgun analysis of the proteome of phloem and xylem saps from rice, taking advantage of the complete and available genomic information for this plant. Xylem sap was prepared using the root pressure method, whereas phloem sap was prepared with a unique method with the assistance of planthoppers to ensure the robustness of the detected proteins. The technical difficulties caused by the very limited availability of rice samples were overcome by the use of nano-flow liquid chromatography linked to a mass spectrometer. We identified 118 different proteins and eight different peptides in xylem sap, and 107 different proteins and five different peptides in phloem sap. Signal transduction proteins, putative transcription factors and stress response factors as well as metabolic enzymes were identified in these saps. Interestingly, we found the presence of three TERMINAL FLOWER 1/FLOWERING LOCUS T (FT)-like proteins in phloem sap. The detected FT-like proteins were not rice Hd3a (OsFTL2) itself that acted as a non-cell-autonomous signal for flowering control, but they were members of distinct subfamilies of the FT family with differential expression patterns. These results imply that proteomics on a nano scale is a potent tool for investigation of biological processes in plants.

Regulation of sulfur-responsive gene expression by exogenously applied cytokinins in Arabidopsis thaliana.

Effects of plant hormones on a sulfur-deficiency responsive element (betaSR) from the promoter region of the beta subunit gene of beta-conglycinin, a major seed storage protein of soybean, were investigated using transgenic Arabidopsis thaliana. Among the hormones tested, the cytokinins, trans-zeatin (Z) and trans-zeatin riboside, upregulated gene expression directed by the betaSR element both in the presence and in the absence of sulfate in the medium. Z also increased transcript accumulation of two endogenous sulfur-responsive genes, the adenosine 5'-phosphosulfate reductase (APR1) and the Sultr2;2, a sulfate transporter. Concentrations of cytokinins were unaltered during early stages of sulfur starvation when expression of these genes was upregulated. Z did not alter concentrations of O-acetyl-L-serine, a positive regulator of gene expression in sulfur starvation response. Concentrations of sucrose, which is known to upregulate expression of APR1, were increased in rosette leaves by Z. Sucrose application to the medium also increased expression directed by the betaSR element, although sucrose concentrations in tissues were not significantly altered by sulfur availability. These results suggest that exogenously applied cytokinins positively regulate expression of these sulfur responsive genes through a pathway independent of that from sulfur starvation, possibly through increasing sucrose concentrations in tissues.