Gordon V. Johnson

The role of endomycorrhizal fungi in iron uptake by Hilaria jamesii

Abstract The semiarid warm season grass galleta [Hilaria jamesii (Torr.)] was used in a series of mycorrhizal experiments to determine the mechanism of enhanced iron (Fe) absorption under conditions of Fe deficiency. Galleta was grown from seed in sterilized soil which had either been infested with mycorrhizal fungi or left uninfested. The four fungal species used to infest the soil were Glomus albidum, Glomus fasciculatum, Glomus macrocarpum, and Glomus mosseae. After 85 days of growth, the nonmycorrhizal and mycorrhizal plants were removed from the soil and placed in complete Hoaglands hydroponic nutrient solutions with phosphorus (P) at 1/5 the normal concentration. Galleta, grown hydroponically from seed, was used as a second nonmycorrhizal control. At the end of ten days of growth, the plants were transferred to Hoaglands nutrient solution at 1/5 the phosphorus concentration and without Fe. The plants were grown for twenty days in this solution; at the end of the growth period, all of the treatment...

Inhibition of ferric chelate reductase in alfalfa roots by cobalt, nickel, chromium, and copper

Abstract This research was initiated to examine the influence of metal cations on ferric chelate reducing capacity of alfalfa grown in hydroponic culture under conditions of iron sufficient or iron limited conditions. In the absence of added metal ions, ferric chelate reductase was greater in plants cultivated in iron limited conditions than in plants where iron was not limiting. Ferric chelate reductase activity in alfalfa roots was inhibited in both iron limited and iron sufficient growth by 0.5 to 2.5 μM Co2+. While the addition of Cr3+ at 2.0 μM slightly inhibited ferric chelate reductase in roots of plants grown under iron limited conditions, Cr3+ at 10 μM stimulated ferric chelate reductase in roots from both iron limited and iron sufficient media. Cu2+ additions at 5 uM to hydroponic media greatly inhibits ferric chelate reductase in plants with iron limited growth while shoot growth was inhibited in both iron sufficient and iron limited growth. The addition of Ni2+ at levels greater than 5 μM to alfalfa cultures inhibits both root and shoot growth; however, at these concentrations of Ni2+ ferric chelate reductase is stimulated in iron sufficient media but reduced in iron stressed plants. Acidification of the media was observed only with iron stressed cultures and did not closely correlate with levels of ferric chelate reductase of the roots.

Reduction and transport of Fe from siderophores

Soils contain siderophores produced by bacteria and fungi; however, the role of siderophores in Fe nutrition of plants is uncertain. The Strategy I plant cucumber (Cucumis sativus L.) was used in an investigation of ferric chelate reduction activity and uptake and transport of Fe from ferric hydroxyethylethylenetriacetic acid (FeHEDTA) and ferric N,N′–di–(2–hydroxybenzoyl)–ethylenediamine– N,N′-diacetic acid (FeHBED) and the hydroxamate siderophores, ferric rhodotorulic acid (FeRA) and ferric ferrioxime B (FeFOB). Cucumber seedlings were grown in a hydroponic medium without Fe or supplied with 10 μM FeHEDTA. Iron-deficient cucumber roots readily reduced FeHEDTA, while Fe-sufficient roots had low levels of ferric chelate reduction activity. The siderophore FeRA was reduced by Fe-deficient roots at 8% of the rate of FeHEDTA, while FeFOB was not reduced. The highly stable synthetic chelate FeHBED was reduced at 16% the rate of FeHEDTA. Fe transport to shoots by Fe-deficient seedlings from the slowly reducible complexes 59FeRA and 59FeHBED was, respectively, 74% and 73% of that transported from 59FeHEDTA. The ferrous complexing agent, bathophenanthrolinedisulfonic acid (BPDS), had a strong inhibitory effect on uptake and transport of Fe from 59FeHEDTA or 59FeRA into shoots. An average of 11% as much Fe was transported to shoots of Fe-deficient seedlings from 59FeFOB as from 59FeHEDTA. Neither the Fe nutritional status of the seedlings nor the presence of BPDS influenced the uptake and transport of Fe from 59FeFOB. It is concluded that cucumber roots may take up substantial amounts of Fe from FeRA and FeHBED following reduction, while small amounts of Fe may be taken up from FeFOB by a mechanism not involving reduction of the ferric siderophore at the root surface.

The effect of azospirilluh brasilense on iron absorption and translocation by sorghum

Abstract Sorghum [Sorghum bicolor (L.) Moench] displays Fe chlorosis when grown in alkaline soils where iron is insoluble and unavailable as a plant nutrient. We were interested in evaluating the role of naturally produced Fe chelators of rhizospheric bacteria in the utilization of insoluble Fe by sorghum. Seedlings of sorghum were grown under hydroponlc conditions in a nutrient solution containing FeHEDTA or without Fe. All seedlings were transferred after two.weeks into fresh nutrient solution containing FeCl3 with CaCO3 added to adjust the pH to 7.4 and to precipitate Fe. Azospirillum brasilense, a nitrogen fixing bacterium often associated with roots of grasses, was introduced into several of the hydroponic cultures to determine if the siderophore produced by the bacterium would enable sorghum to use the precipitated inorganic Fe. Utilization of added 59FeCl3 was indicated by measurement of radiolabeled Fe in the roots and shoots or by determination of the chlorophyll content of the leaf blades. Addit...

Natural abundance of 15N in actinorhizal plants and nodules

Substantial enrichment of some plant parts in 15N relative to the rest of the plant is unusual, but is found in the nitrogen-fixing nodules of many legumes. A range of actinorhizal plants was surveyed to determine whether the nodules of any of them are also substantially enriched in 15N. The nonlegume Parasponia, nodulated by a rhizobium, was also included. Four of the actinorhizal genera and Parasponia were grown in N-free culture, and three actinorhizal genera were collected from the field. Nodules of Parasponia, Casuarina and Alnus were15N enriched relative to other plant parts, but only Parasponia approached the degree of enrichment found in some legume nodules. The nodules of Datisca, Myrica, Elaeagnus, Shepherdia, and Coriaria were depleted in 15N. Thus many actinorhizal nodules are depleted in 15N compared to other plant parts and enrichment is modest when it does occur. Whole plant 15N content (δ15N) in four actinorhizal plants and Parasponia showed a relatively narrow range of −1.41 to −1.90. Hence regardless of the degree of nodule enrichment or depletion, whole plant 15N content appears to vary little in plants grown in N-free culture.

Siderophore‐mediated iron metabolism in growth and nitrogen fixation by alfalfa nodulated with Rhizobium meliloti

Abstract This research was initiated to examine the significance of the high‐affinity iron (Fe) transport system in the nitrogen (N)‐fixing activity of Rhizobium meliloti bacteroides in nodules of alfalfa, Medicago sativa L. Iron was supplied as Fe‐chelate at a concentration of 10μM for nodulation and plant growth. Nitrogen‐fixation measurements by the acetylene reduction assay were conducted on plants with adequate Fe in the hydroponic medium. Mutants of R. meliloti strain 1021 that were defective in rhizobactin 1021 synthesis produced nodules on alfalfa but fixed insignificant amounts of dinitrogen. Mutants of R. meliloti constitutive forrhizobactin 1021 synthesis also produced nodules; however, the levels of dinitrogen fixed were low. Nodules produced by a strain of R. meliloti defective in rhizobactin‐Fe uptake and subject to siderophore induction were found to fix dinitrogen at levels similar to nodules produced by wild type bacteria. Iron availability to R. meliloti in developing nodules was critica...

Application of high performance liquid chromatography in the characterization of iron stress response

Abstract When Fe‐efficient plants become Fe deficient, physiological responses occur which may increase the availability of Fe in the root environment. Iron‐stress responses of the sunflower (Helianthus annus L.) include a lowering of the medium pH, an increase in the reducing capacity of the root, and a release of riboflavin into the root medium. In this investigation, compounds released into the root medium during Fe‐stress were characterized using high performance liquid chrbmatography (HPLC). Samples of nutrient solution from hydroponically grown sunflowers (cultivar Large Russian) were analyzed by HPLC using an acetonitrile : water gradient.. Absorbance at 254 nm was recorded as compounds were separated. Negligible concentrations of compounds absorbing at 254 nm were found to be released by non‐stressed sunflower plants; however, nutrient solutions from Fe‐stressed plants contained riboflavin and at least two other compounds not present in control nutrient solutions. The concentration of these compou...

Selection of iron‐efficient soybean cell lines using plant cell suspension culture

Abstract Plant cell suspension culture may prove to be useful in the selection and isolation of cell lines which are efficient in mineral utilization. We have successfully used plant cell suspension culture to select for iron‐efficient soybean cell lines. Some of the iron utilization characteristics of iron‐efficient, Hawkeye (HA) and iron‐inefficient, PI‐54619–5–1 (PI) soybean [Glycine max (L.) Merrill] cell lines in plant cell suspension culture were examined to determine their suitability as markers for identifying selected iron‐efficient cell lines. The growth response of soybean cell lines to low iron levels can be used to identify iron‐efficient plant cells. Cell suspension cultures of Pi‐soybeans grown on Gamborgs B5C medium required a higher concentration of iron as FeEDTA than HA‐soybeans. Both cell lines adapted to growth at sub‐optimal iron concentrations and yields increased after prolonged culture with sub‐optimal iron. Short term iron absorption experiments failed to differentiate iron‐effi...

Responses of nitrogen-fixing and nitrate-supplied alfalfa (Medicago sativa L.) to iron chelates in an alkaline hydroponic medium

Ferric ethylenediamine di-(o-hydroxyphenylacetate) (FeEDDHA) and ferric hydroxyethylethylenediaminetriacetic acid (FeHEDTA) were evaluated as Fe sources for hydroponic growth of alfalfa (Medicago sativa L., cv. Mesilla), either dependent on N2 fixation or supplied with NO3. The hydroponic medium was maintained at pH 7.5 by addition of CaCO3. Nitrogen-fixing cultures were inoculated with Rhizobium meliloti 102 F51 and grown in medium without added nitrogen. After five to seven weeks of growth under greenhouse conditions, plants were harvested. Nitrogen fixation was measured by the acetylene reduction method.

Characterization of iron uptake by iron‐efficient and iron‐inefficient soybeans in cell suspension culture

Abstract Cell suspension cultures of Fe‐efficient Hawkeye (HA) and Fe‐inefficient Pl‐54619–5–1 (PI) soybeans [Glycine max (L.) Merrill] respond differently to Fe‐stress and take up different quantities of Fe, but reflect differences observed at the whole plant level. Short‐term Fe uptake experiments showed that HA‐soybean cells respond to Fe‐stress by increasing their ability for Fe uptake. Pi‐soybean cells have a significantly decreased ability for Fe uptake when Fe‐stressed. Kinetic studies indicated that HA‐soybean cells had greater Km and Vmax values for Fe uptake than PI‐soybean cells. The data indicate that HA‐soybean cells have an inducible Fe transport system that is stimulated under conditions of Fe‐stress. It appears that when Fe‐stressed, HA‐soybean cells may increase the number of Fe reducing and/or Fe transporting sites, thus increasing their ability for Fe uptake. Iron is taken up as the reduced ferrous (Fe2+) form by a system showing a high specificity for Fel as indicated by competing cati...