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T2 tobacco lines overexpressing soybean ferritin in the plastids (+TPs) or apoplasm (AFs) under the regulation of CaMV 35S promoter were grown on MS nutrient solution. After 1 month growth, statuses of six major divalent‐metals (Ca, Cu, Fe, Mg, Mn, and Zn) were measured in leaves and roots. Both +TPs and AFs showed enhanced growth (max. 1.7×) in leaves than the control line. The Fe contents in leaves of +TPs and AFs were significantly larger (1.9–2.8×) than that of the control line. The other metal contents in leaves of +TPs and AFs were almost the same as or less than those of the control line. In contrast to the result of leaves, the growth enhancement in roots was not clear in +TPs, but in AFs. Also, some of the non‐ferrous metal contents in roots of +TPs and AFs were dramatically increased compared with those of the control line (Mn, 1.9–10.4×; Zn, 1.6–2.3×), whereas the differences in content of Fe, Cu, Ca, and Mg were insignificant. These results demonstrated that the ferritin overexpressio...

The phytotoxicity of aluminum (Al) in relation to preculture with phosphates was examined in the rice cultivar Arkansas fortuna. In plants precultured with phosphates, Al did not inhibit shoot growth, while Al retarded shoot growth in plants precultured without phosphates. In contrast, Al inhibited root elongation, irrespective of the presence of phosphates in the preculture solution. A large proportion of the Al in roots was in unknown, insoluble forms. In phosphate‐precultured plants, Al deposition was slightly increased, presumably due to the formation of aluminum phosphates in the roots, and phosphorus levels in shoots were markedly increased. Binding with phosphates may ameliorate the toxicity of Al when it enters the shoots and account for the uninhibited shoot growth in the presence of Al in plants precultured with phosphates.

S‐adenosyl‐L‐methionine (SAM) synthetase, which catalyzes methionine to SAM, is one of the enzymes involved in the biosynthesis of the mugineic acid‐family phytosiderophores (MAs). The MAs are secreted from grass roots under iron (Fe)‐deficient conditions. We compared the SAM synthetase activities between Fe‐deficient and Fe‐sufficient barley roots. However, no difference was observed. Then we performed Northern hybridization analysis between Fe‐deficient and Fe‐sufficient barley tissues by a cDNA fragment of SAM synthetase isolated from barley. No increase in the amount of mRNA was detected in the Fe‐deficient barley roots as compared to the Fe‐sufficient ones. It was suggested that the first step in the phytosiderophore biosynthesis induced under the Fe‐deficiency is not SAM synthetase, but NA synthase and the second induced step is NA aminotransferase.

Nicotianamine is proposed to play a key role in iron (Fe) metabolism in plants. In graminaceous plants nicotianamine is the precursor for phytosiderophores, and the activity of the nicotianamine synthase is strongly enhanced under Fe deficiency. The tomato mutant ‘chloronerva’ (Lycopersicon esculentum Mill, cv. chloronerva) lacks nicotianamine and has a disturbed Fe homeostasis. Our objective was to compare nicotianamine synthase activity of the mutant ‘chloronerva’ with the wild form ('Bonner Beste'). In contrast to the wild form which as high nicotianamine synthase activity, no activity of the enzyme could be detected in roots or shoots of ‘chloronerva’, regardless of the iron nutritional status of the plants. This finding might allow comparison of promoter regions of the genes of ‘chloronerva’, ‘Bonner Beste’, and graminaceous species to clarify the regulatory mechanism of gene expression by the Fe nutritional status in graminaceous species.

The expression of two barley genes, Ids1 and Ids2, that were induced specifically by iron (Fe) deficiency stress in solution culture, was examined in two barley genotypes differing in manganese (Mn) efficiency. Plants were grown in a calcareous soil supplied with two levels of Mn (15 and 100 mg/kg soil). Ids1 was expressed at equal levels in the roots of both genotypes, and this expression was not affected by Mn supply. These results suggest that the expression of Ids1 probably does not contribute to Mn efficiency. A contrasting result was obtained for Ids2, which was expressed at a higher level in the roots of the Mn‐inefficient genotype than in the Mn‐efficient genotype. However, the expression levels also were not affected by Mn supply. The differential expression of Ids2 may indicate that this gene plays a role both in the Fe deficiency response and in the Mn efficiency mechanism. An interesting observation made on the time course of expression of the two genes. Initially, both genes had low ...

An experiment with different chelators involving plant made mugineic acid (MA), deferrioxamine B (FOB) and DTPA along with deionized water as a control was performed to evaluate the Fe extraction ability of the chelators from Indian calcareous soils having high pH, low Fe content, and rich in calcium carbonate. MA manifested the highest Fe extraction efficiency in all the calcareous soils. On the contrary, other chelators displayed rather poor activity of Fe extraction. MA also mobilized Cu, Zn, Mn, Ca, and Mg. However, DTPA solubilized Ca more efficiently than MA. Different calcareous soils also varied with respect to Fe extraction ability. In a green house experiment, Indian barley cultivar (cv) NDB‐209 under calcareous soils, however, exhibited very low Fe solubility in the soil rhizosphere indicating the possibility of simultaneous and periodical release and high Fe(III)‐MAs uptake rate by the plants in quick succession to optimize Fe acquisition efficiency and minimize the microbial decompos...

For the purpose of gene cloning for HAs‐synthesis or Fe(III)‐MAs transporter a ? gt10 cDNA library was constructed from mRNA isolated from Fe‐deficient barley roots. The library was then differentially screened between cDNA probes made from mRNA isolated from barley roots treated with +Fe and ‐Fe. Seven clones which hybridized specifically to the probe of Fe‐deficiency were selected. Their inserts however were too short and not likely to include full length of mRNA. On the basis of these results we decided to screen a newly constructed ? zapII cDNA library with one of the seven clones as a probe and selected a clone presumably having the full length of mRNA compared with northern hybridization. We named this cone as Ids1. The sequenced Ids1 consists of 503 nucleotides containing a putative open reading frame of 222 bp. It encodes a protein of 74 residues (7500 Da) having two cysteine rich domains like animal MT (class I MT).

The peptides appearing in barley roots related to phytosiderophore synthesis or absorption under Fe‐deficiency were analyzed. Two varieties of barley were used. ‘Ehimehadaka NO.11 secreted 3‐epihydroxymugineic acid and traces of 3‐hydroxymugineic acid, mugineic acid and 2'‐deoxymugineic acid. ‘Minorimugi’ secreted mugineic acid, and traces of 3‐epihydroxymugineic acid and 2'‐deoxymugineic acid. Peptides in supernatant fractions of Fe‐deficient roots were detected by 35s‐fluorography and/or by silver‐stain on two dimensional electrophoresis. In the case of ‘Ehimehadaka NO.1’, two peptides (m.w. 53kD, pI 6.6) and (53kD, 6.7) by fluorography, and (52kD, 6.9), (31kD, 6.6) and (30kD, 6.8) by silver‐stain were detected. In the case of ‘Minorimugi’, two peptides (53kD, 6.6) and (53kD, 6.7) by fluorography, (31kD, 6.6) and (30kD, 6.8) by silver‐stain were detected. The synthesis of the 53kD peptide was strongly related to the secretion amount of phytosiderophores. Whereas the 52kD related to 3‐epihydroxy...

An HPLC method for MAs (mugineic acid and its analogues) analysis was established. By this method, time course change of MAs secreted from the roots of Fe‐deficient barley were traced. After about one week of ‐Fe treatment, MAs secreted from the roots gradually increased and after 40 days it became plateau. The several MAs were synchronously secreted from the roots throughout a day; their amounts were maximum at noon. The pattern of MAs secretion was different from that of amino acids. MAI was found to be trace in roots, xylem sap and shoots of the control (+Fe) plant. However, MAs concentrations remarkably increased in those tissues when Fe‐deficiency chlorosis progressed due to ‐Fe treatment. From experiments of 14CO2‐fixation by photosynthesis, 14C‐MA1 secreted from the roots was found to be biosynthesized with a half‐life of 24 h.

The changes in the ultrastructure of barley root cells induced by Fe‐deficiency was investigated in relation to secretion of mugineic acid and its analogues (MAS). We found that the particular vesicles were present in barley root cells under both conditions of sufficient and deficient iron. These particular vesicles were increased in number and volume under Fe‐deficient condition. These vesicles contained fibrous materials. Their limiting membrane was thinner than that of Golgi vesicles, and they had ribosomes around their surface. Their structure suggests that they originated from rough endoplasmic reticulum. The vesicle volume was expanded especially before sunrise when MAs were enriched in Fe‐deficient barley roots. In the midday, after secretion of MAs, the volume of the vesicles decreased.