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David A Day

Publications and source records attributed to David A Day.

25 records · Page 2Linked to original sources
PubMed2003

The soybean NRAMP homologue, GmDMT1, is a symbiotic divalent metal transporter capable of ferrous iron transport.

Iron is an important nutrient in N2-fixing legume root nodules. Iron supplied to the nodule is used by the plant for the synthesis of leghemoglobin, while in the bacteroid fraction, it is used as an essential cofactor for the bacterial N2-fixing enzyme, nitrogenase, and iron-containing proteins of the electron transport chain. The supply of iron to the bacteroids requires initial transport across the plant-derived peribacteroid membrane, which physically separates bacteroids from the infected plant cell cytosol. In this study, we have identified Glycine max divalent metal transporter 1 (GmDmt1), a soybean homologue of the NRAMP/Dmt1 family of divalent metal ion transporters. GmDmt1 shows enhanced expression in soybean root nodules and is most highly expressed at the onset of nitrogen fixation in developing nodules. Antibodies raised against a partial fragment of GmDmt1 confirmed its presence on the peribacteroid membrane (PBM) of soybean root nodules. GmDmt1 was able to both rescue growth and enhance 55Fe(II) uptake in the ferrous iron transport deficient yeast strain (fet3fet4). The results indicate that GmDmt1 is a nodule-enhanced transporter capable of ferrous iron transport across the PBM of soybean root nodules. Its role in nodule iron homeostasis to support bacterial nitrogen fixation is discussed.

Amino Acid Sequence↗
PubMed2002

Environmental stress causes oxidative damage to plant mitochondria leading to inhibition of glycine decarboxylase.

A cytotoxic product of lipid peroxidation, 4-hydroxy-2-nonenal (HNE), rapidly inhibited glycine, malate/pyruvate, and 2-oxoglutarate-dependent O2 consumption by pea leaf mitochondria. Dose- and time-dependence of inhibition showed that glycine oxidation was the most severely affected with a K(0.5) of 30 microm. Several mitochondrial proteins containing lipoic acid moieties differentially lost their reactivity to a lipoic acid antibody following HNE treatment. The most dramatic loss of antigenicity was seen with the 17-kDa glycine decarboxylase complex (GDC) H-protein, which was correlated with the loss of glycine-dependent O2 consumption. Paraquat treatment of pea seedlings induced lipid peroxidation, which resulted in the rapid loss of glycine-dependent respiration and loss of H-protein reactivity with lipoic acid antibodies. Pea plants exposed to chilling and water deficit responded similarly. In contrast, the damage to other lipoic acid-containing mitochondrial enzymes was minor under these conditions. The implication of the acute sensitivity of glycine decarboxylase complex H-protein to lipid peroxidation products is discussed in the context of photorespiration and potential repair mechanisms in plant mitochondria.

Aldehydes↗
PubMed2002

Regulation of alternative oxidase gene expression in soybean.

Soybean (Glycine max cv. Stevens) suspension cells were used to investigate the expression of the alternative oxidase (Aox) multigene family. Suspension cells displayed very high rates of cyanide-insensitive respiration, but Aox3 was the only isoform detected in untreated cells. Incubation with antimycin A, citrate, salicylic acid or at low temperature (10 degrees C) specifically induced the accumulation of the Aox1 isoform. Aox2 was not observed under any conditions in the cells. Increases in Aox1 protein correlated with increases in Aox1 mRNA. Treatment of soybean cotyledons with norflurazon also induced expression of Aox1. Reactive oxygen species (ROS) were detected upon incubation of cells with antimycin, salicylic acid or at low temperature, but not during incubation with citrate. Aox1 induction by citrate, but not by antimycin, was prevented by including the protein kinase inhibitor staurosporine in the medium. The results suggest that multiple pathways exist in soybean to regulate expression of Aox genes and that Aox1 specifically is induced by a variety of stress and metabolic conditions via at least two independent signal transduction pathways.

Antimycin A↗
PubMed2002

Reassessment of major products of N2 fixation by bacteroids from soybean root nodules.

NH3/ was the principal product from soybean bacteroids, prepared by various procedures, when assayed in solution in a flow chamber under N2 fixation conditions. In addition, small quantities of alanine were produced (reaching 20% of NH3/ under some conditions). Some 15N was assimilated by bacteroids purified from soybean root nodules on Percoll density gradients and shaken with 15N2 and 0.008 atm O2. Under these conditions, accounted for 93% of the (15)N fixed into the soluble fraction. This fraction contained no measurable [15N]alanine. Neither these bacteroids nor those prepared by the previously used differential centrifugation method, when incubated with exogenous alanine under non-N2-fixing conditions, gave rise to NH3 from alanine. Therefore, contamination of bacteroid preparations with enzymes of plant cytosolic origin and capable of producing NH3 from alanine cannot explain the failure to detect [15N]alanine [as reported elsewhere: Waters, J. K., Hughes, B. L., II, Purcell, L. C., Gerhardt, K. O., Mawhinney, T. P. & Emerich, D. W. (1998). Proc Natl Acad Sci USA 95, 12038-12042]. Cell-free extracts of the bacteroids as used in the 15N experiments contained alanine dehydrogenase and were able to produce alanine from pyruvate and. Other experiments with alanine dehydrogenase in extracts of cultured rhizobia and bacteroids are reported and discussed in relation to the 15N experiments. Possible reasons for the differences between laboratories regarding the role of alanine are discussed. It is concluded that NH3 is the principal soluble product of N2 fixation by suspensions of soybean bacteroids ex planta and that should continue to be considered the principal product of N2 fixation which is assimilated in vivo in soybean nodules.

Alanine↗
PubMed2001

GmZIP1 encodes a symbiosis-specific zinc transporter in soybean.

The importance of zinc in organisms is clearly established, and mechanisms involved in zinc acquisition by plants have recently received increased interest. In this report, the identification, characterization and location of GmZIP1, the first soybean member of the ZIP family of metal transporters, are described. GmZIP1 was found to possess eight putative transmembrane domains together with a histidine-rich extra-membrane loop. By functional complementation of zrt1zrt2 yeast cells no longer able to take up zinc, GmZIP1 was found to be highly selective for zinc, with an estimated K(m) value of 13.8 microm. Cadmium was the only other metal tested able to inhibit zinc uptake in yeast. An antibody raised against GmZIP1 specifically localized the protein to the peribacteroid membrane, an endosymbiotic membrane in nodules resulting from the interaction of the plant with its microsymbiont. The specific expression of GmZIP1 in nodules was confirmed by Northern blot, with no expression in roots, stems, or leaves of nodulated soybean plants. Antibodies to GmZIP1 inhibited zinc uptake by symbiosomes, indicating that at least some of the zinc uptake observed in isolated symbiosomes could be attributed to GmZIP1. The orientation of the protein in the membrane and its possible role in the symbiosis are discussed.

Amino Acid Sequence↗
PubMed2001

Supply of O2 regulates demand for O2 and uptake of malate by N2-fixing bacteroids from soybean nodules.

Bacteroids, prepared anaerobically from soybean root nodules by fractional centrifugation or by sucrose or Percoll density-gradient methods, were retained within a stirred, flow-through reaction chamber and used to determine rates of respiration and N2 fixation at various rates of O2 supply. Liquid reaction solutions containing malate, oxyleghaemoglobin, dissolved N2 and various levels of dissolved O2 were passed through the reaction chamber at measured rates of flow. The relative oxygenation of leghaemoglobin in the chamber was determined automatically by spectrophotometry of the effluent solution, and the concentrations of free, dissolved O2 ([O2(free)]) and the rates of O2 consumption were calculated. N2 fixation was measured by analysis of fractions of effluent. The principal finding was that stepwise increases in the flow rate (increasing the supply of O2 and malate) induced an increase in O2 demand (respiration) resulting in a decrease in [O2(free)] and increased N2 fixation. In some experiments, samples of bacteroids were withdrawn from the flow chamber during steady states and the rates of malate uptake were measured in standard, microaerobic assays. Progressive taking of samples from the flow chamber whilst maintaining constant flow rates (increasing the supply of O2 and malate per bacteroid) also resulted in increased O2 demand and declines in [O2(free)]. With increased bacteroid respiration, transport of malate into bacteroids (linear with time between 1 and 5 min after starting each assay) increased proportionally. This suggests that the rate of malate transport is tightly coupled with bacteroid respiration. Thus, bacteroid respiration, coupled with malate uptake, must be regulated by the rate of O2 supply, rather than by the [O2(free)] prevailing in the stirred chamber as found or assumed in previous work. These features are discussed in relation to N2 fixation by anaerobically isolated bacteroids.

Bradyrhizobium↗