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OsTZF1 is a member of the CCCH-type zinc finger gene family in rice (Oryza sativa). Expression of OsTZF1 was induced by drought, high-salt stress, and hydrogen peroxide. OsTZF1 gene expression was also induced by abscisic acid, methyl jasmonate, and salicylic acid. Histochemical activity of β-glucuronidase in transgenic rice plants containing the promoter of OsTZF1 fused with β-glucuronidase was observed in callus, coleoptile, young leaf, and panicle tissues. Upon stress, OsTZF1-green fluorescent protein localization was observed in the cytoplasm and cytoplasmic foci. Transgenic rice plants overexpressing OsTZF1 driven by a maize (Zea mays) ubiquitin promoter (Ubi:OsTZF1-OX [for overexpression]) exhibited delayed seed germination, growth retardation at the seedling stage, and delayed leaf senescence. RNA interference (RNAi) knocked-down plants (OsTZF1-RNAi) showed early seed germination, enhanced seedling growth, and early leaf senescence compared with controls. Ubi:OsTZF1-OX plants showed improved tolerance to high-salt and drought stresses and vice versa for OsTZF1-RNAi plants. Microarray analysis revealed that genes related to stress, reactive oxygen species homeostasis, and metal homeostasis were regulated in the Ubi:OsTZF1-OX plants. RNA-binding assays indicated that OsTZF1 binds to U-rich regions in the 3' untranslated region of messenger RNAs, suggesting that OsTZF1 might be associated with RNA metabolism of stress-responsive genes. OsTZF1 may serve as a useful biotechnological tool for the improvement of stress tolerance in various plants through the control of RNA metabolism of stress-responsive genes.

Gold nanoparticles have particular properties distinct from those of bulk gold crystals, and such nanoparticles are used in various applications in optics, catalysis, and drug delivery. Many reports on microbial synthesis of gold nanoparticles have appeared. However, the molecular details (reduction and dispersion) of such synthesis remain unclear. In the present study, we studied gold nanoparticle synthesis by Lactobacillus casei . A comparison of L. casei components before and after addition of an auric acid solution showed that the level of unsaturated lipids decreased significantly after addition. NMR and mass spectrum analysis showed that the levels of diglycosyldiacylglycerol (DGDG) and triglycosyldiacylglycerol (TGDG) bearing unsaturated fatty acids were much reduced after formation of gold nanoparticles. DGDG purified from L. casei induced the synthesis of gold nanoparticles in vitro . These results suggested that glycolipids, such as DGDG, play important roles in reducing Au(III) to Au(0) and in ensuring that the nanoparticles synthesized remain small in size. Our work will lead to the development of novel, efficient methods by which gold nanoparticles may be produced by, and accumulated within, microorganisms.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Iron and steel slag (ISS) is a byproduct of iron refining processes. The lack of iron in seawater can cause barren grounds where algae cannot grow. To improve the barren grounds of the sea, a supply of iron to the seawater is necessary. This study focused on bacteria interacting with ISS and promoting iron elution in seawater. Sulfitobacter sp. (TO1A) and Pseudomonas sp. (TO1B) were isolated from Tokyo Bay and Sagami Bay. The co-culture of both bacteria promoted more iron elution than individual cultures. After the incubation of both bacteria with ISS, quartz and vaterite appeared on the surface of the ISS. To maintain continuous iron elution from the ISS in the seawater, we also isolated Pseudoalteromonas sp. (TO7) that formed a yellow biofilm on the ISS. Iron was eluted by TO1A and TO1B, and biofilm was synthesized by TO7 continuously in the seawater. The present research is expected to contribute to the improvement of ISS usage as a material for the construction of seaweed forests.

CdSe quantum dots are often used in industry as fluorescent materials. In this study, CdSe quantum dots were synthesized using Fusarium oxysporum. The cadmium and selenium concentration, pH, and temperature for the culture of F. oxysporum (Fusarium oxysporum) were optimized for the synthesis, and the CdSe quantum dots obtained from the mycelial cells of F. oxysporum were observed by transmission electron microscopy. Ultra-thin sections of F. oxysporum showed that the CdSe quantum dots were precipitated in the intracellular space, indicating that cadmium and selenium ions were incorporated into the cell and that the quantum dots were synthesized with intracellular metabolites. To reveal differences in F. oxysporum metabolism, cell extracts of F. oxysporum, before and after CdSe synthesis, were compared using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The results suggested that the amount of superoxide dismutase (SOD) decreased after CdSe synthesis. Fluorescence microscopy revealed that cytoplasmic superoxide increased significantly after CdSe synthesis. The accumulation of superoxide may increase the expression of various metabolites that play a role in reducing Se4+ to Se2− and inhibit the aggregation of CdSe to make nanoparticles.

Nicotianamine (NA), a chelator of metals, is ubiquitously present in higher plants. Nicotianamine aminotransferase (NAAT) catalyzes the amino group transfer of NA in the biosynthetic pathway of phytosiderophores and is essential for iron acquisition in graminaceous plants. The gene that encodes NAAT from barley was introduced into the nongraminaceous plant tobacco, which produces NA but not phytosiderophores. Transgenic tobacco plants (naat tobacco) that constitutively expressed the NAAT gene had young leaves with interveinal chlorosis and flowers that were abnormally shaped and sterile. Endogenous NA was consumed as a result of NAAT overproduction in naat tobacco. The resulting NA shortage caused disorders in internal metal transport, leading to these abnormal phenotypes. In addition to its role in long-distance metal transport, NA may be involved in the regulation of metal transfer within the cells. These results suggest that a shortage of NA impaired the functions of metal-requiring proteins, including transcription factors.

An iron (Fe)-binding substance (FeBS) was sought for the marine diatom Thalassiosira pseudonana, because the organism is devoid of a gene that encodes the predominant Fe storage protein ferritin. High-performance liquid chromatography-inductively coupled plasma/emission mass spectrometry (HPLC-ICP/MS) analysis of an extract of diatom cells indicated the presence of an FeBS specific to the cells grown in Fe-replete medium. Heat treatment followed by anion-exchange chromatography with monitoring of Fe(III)-dissolving activity was used to isolate the FeBS from the cell extract. Structural analysis using 1H NMR spectrometry demonstrated that the FeBS was composed of citrate. In addition, HPLC-ICP/MS together with ESI-MS demonstrated that the FeBS was likely a triferric tricitrate complex. Although the citrate content of the diatom cells was enhanced according to the Fe nutritional status of the medium, this phenomenon seemed to occur by an indirect mechanism, as a previous report demonstrated a reduced citrate content in T. pseudonana cells grown in Fe-replete medium (Bromke et al. in Plos One doi: 10.1371/journal.pone.0067340, 2013 ). It is most likely that, based on the extended Redfield ratio 121(C):16(N):1(P):0.0075(Fe), the culture used in the current study may have suffered from nutrient deficiency of a major element, such as nitrogen. Such a situation may occur frequently for this open sea diatom. As windblown dust is a prevailing Fe source for the open oceanic diatom T. pseudonana, a rapid elevation in Fe concentration in the seawater can render the cells nitrogen deficient, causing enhanced citrate synthesis and resulting in the formation of a triferric tricitrate complex as Fe storage machinery.

Transmission electron microscopy revealed the presence of electron-dense bodies (EDB) in the cytosol of the acidophilic, thermophilic red alga Cyanidium caldarium. These bodies contain almost exclusively Fe, P, and O and can play a role in Fe storage. ³¹P-nuclear magnetic resonance analysis identified a sharp signal at -23.3 ppm, which was attributed to the phosphate groups of the inner portions of polyphosphate chains. From this evidence, as well as that of a previous ESR study (Nagasaka et al., BioMetals 16:465-470, 2003), it can be concluded that polyphosphates are the major anionic constituents of the EDB. Omission of Fe from the culture medium resulted in substantially decreased polyphosphate levels, demonstrating the control of cellular polyphosphate content by the Fe status of the culture medium.

Osmotic adjustment plays a fundamental role in water stress responses and growth in plants; however, the molecular mechanisms governing this process are not fully understood. Here, we demonstrated that the KUP potassium transporter family plays important roles in this process, under the control of abscisic acid (ABA) and auxin. We generated Arabidopsis thaliana multiple mutants for K⁺ uptake transporter 6 (KUP6), KUP8, KUP2/SHORT HYPOCOTYL3, and an ABA-responsive potassium efflux channel, guard cell outward rectifying K⁺ channel (GORK). The triple mutants, kup268 and kup68 gork, exhibited enhanced cell expansion, suggesting that these KUPs negatively regulate turgor-dependent growth. Potassium uptake experiments using ⁸⁶radioactive rubidium ion (⁸⁶Rb⁺) in the mutants indicated that these KUPs might be involved in potassium efflux in Arabidopsis roots. The mutants showed increased auxin responses and decreased sensitivity to an auxin inhibitor (1-N-naphthylphthalamic acid) and ABA in lateral root growth. During water deficit stress, kup68 gork impaired ABAmediated stomatal closing, and kup268 and kup68 gork decreased survival of drought stress. The protein kinase SNF1-related protein kinases 2E (SRK2E), a key component of ABA signaling, interacted with and phosphorylated KUP6, suggesting that KUP functions are regulated directly via an ABA signaling complex. We propose that the KUP6 subfamily transporters act as key factors in osmotic adjustment by balancing potassium homeostasis in cell growth and drought stress responses.

Two free flavin-independent enzymes were purified by detecting the NAD(P)H oxidation in the presence of Fe(III)-EDTA and t-butyl hydroperoxide from E. coli. The enzyme that requires NADH or NADPH as an electron donor was a 28 kDa protein, and N-terminal sequencing revealed it to be oxygen-insensitive nitroreductase (NfnB). The second enzyme that requires NADPH as an electron donor was a 30 kDa protein, and N-terminal sequencing revealed it to be ferredoxin-NADP⁺ reductase (Fpr). The chemical stoichiometry of the Fenton activities of both NfnB and Fpr in the presence of Fe(III)-EDTA, NAD(P)H and hydrogen peroxide was investigated. Both enzymes showed a one-electron reduction in the reaction forming hydroxyl radical from hydrogen peroxide. Also, the observed Fenton activities of both enzymes in the presence of synthetic chelate iron compounds were higher than their activities in the presence of natural chelate iron compounds. When the Fenton reaction occurs, the ferric iron must be reduced to ferrous iron. The ferric reductase activities of both NfnB and Fpr occurred with synthetic chelate iron compounds. Unlike NfnB, Fpr also showed the ferric reductase activity on an iron storage protein, ferritin, and various natural iron chelate compounds including siderophore. The Fenton and ferric reductase reactions of both NfnB and Fpr occurred in the absence of free flavin. Although the k cat/K m value of NfnB for Fe(III)-EDTA was not affected by free flavin, the k cat/K m value of Fpr for Fe(III)-EDTA was 12-times greater in the presence of free FAD than in the absence of free FAD.