Jan K. Schjoerring

Specific Aquaporins Facilitate the Diffusion of Hydrogen Peroxide across Membranes

The metabolism of aerobic organisms continuously produces reactive oxygen species. Although potentially toxic, these compounds also function in signaling. One important feature of signaling compounds is their ability to move between different compartments, e.g. to cross membranes. Here we present evidence that aquaporins can channel hydrogen peroxide (H2O2). Twenty-four aquaporins from plants and mammals were screened in five yeast strains differing in sensitivity toward oxidative stress. Expression of human AQP8 and plant Arabidopsis TIP1;1 and TIP1;2 in yeast decreased growth and survival in the presence of H2O2. Further evidence for aquaporin-mediated H2O2 diffusion was obtained by a fluorescence assay with intact yeast cells using an intracellular reactive oxygen species-sensitive fluorescent dye. Application of silver ions (Ag+), which block aquaporin-mediated water diffusion in a fast kinetics swelling assay, also reversed both the aquaporin-dependent growth repression and the H2O2-induced fluorescence. Our results present the first molecular genetic evidence for the diffusion of H2O2 through specific members of the aquaporin family.

Reflectance measurement of canopy biomass and nitrogen status in wheat crops using normalized difference vegetation indices and partial least squares regression

Abstract Hyperspectral reflectance (438 to 884 nm) data were recorded at five different growth stages of winter wheat in a field experiment including two cultivars, three plant densities, and four levels of N application. All two-band combinations in the normalized difference vegetation index ( λ 1− λ 2)/( λ 1+ λ 2) were subsequently used in a linear regression analysis against green biomass (GBM, g fresh weight m −2 soil), leaf area index (LAI, m 2 green leaf m −2 soil), leaf chlorophyll concentration (Chl conc , mg chlorophyll g −1 leaf fresh weight), leaf chlorophyll density (Chl density , mg chlorophyll m −2 soil), leaf nitrogen concentration (N conc , mg nitrogen g −1 leaf dry weight), and leaf nitrogen density (N density , g nitrogen m −2 soil). A number of grouped wavebands with high correlation ( R 2 >95%) were revealed. For the crop variables based on quantity per unit surface area, i.e. GBM, LAI, Chl density , and N density , these wavebands had in the majority (87%) of the cases a center wavelength in the red edge spectral region from 680 to 750 nm and the band combinations were often paired so that both bands were closely spaced in the steep linear shift between R red and R nir . The red edge region was almost absent for bands related to Chl conc and N conc , where the visible spectral range, mainly in the blue region, proved to be better. The selected narrow-band indices improved the description of the influence of all six-crop variables compared to the traditional broad- and short-band indices normally applied on data from satellite, aerial photos, and field spectroradiometers. For variables expressed on the basis of soil or canopy surface area, the relationship was further improved when exponential curve fitting was used instead of linear regression. The best of the selected narrow-band indices was compared to the results of a partial least square regression (PLS). This comparison showed that the narrow-band indices related to LAI and Chl conc , and to some extent also Chl density and N density , were optimal and could not be significantly improved by PLS using the information from all wavelengths in the hyperspectral region. However, PLS improved the prediction of GBM and N conc by lowering the RMSE with 22% and 24%, respectively, compared to the best narrow-band indices. It is concluded that PLS regression analysis may provide a useful exploratory and predictive tool when applied on hyperspectral reflectance data.

Aquaporin homologues in plants and mammals transport ammonia

Using functional complementation and a yeast mutant deficient in ammonium (NH4 +) transport (Δmep1–3), three wheat (Triticum aestivum) TIP2 aquaporin homologues were isolated that restored the ability of the mutant to grow when 2 mM NH4 + was supplied as the sole nitrogen source. When expressed in Xenopus oocytes, TaTIP2;1 increased the uptake of NH4 + analogues methylammonium and formamide. Furthermore, expression of TaTIP2;1 increased acidification of the oocyte‐bathing medium containing NH4 + in accordance with NH3 diffusion through the aquaporin. Homology modeling of TaTIP2;1 in combination with site directed mutagenesis suggested a new subgroup of NH3‐transporting aquaporins here called aquaammoniaporins. Mammalian AQP8 sharing the aquaammoniaporin signature also complemented NH4 + transport deficiency in yeast.

Ammonia Emission from Mineral Fertilizers and Fertilized Crops

Abstract A thorough understanding of the physical and chemical processes involved in NH 3 emission from inorganic N fertilizers and fertilized crops is required if reliable and operational NH 3 emission factors and decision support systems for inorganic fertilizers are to be developed, taking into account the actual soil properties, climatic conditions and management factors. For this reason, the present review focuses on processes involved in NH 3 volatilization from inorganic nitrogen fertilizers and the exchange of ammonia between crop foliage and the atmosphere. The proportion of nitrogen lost from N fertilizers due to NH 3 volatilization may range from ≈0 to >50%, depending on fertilizer type, environmental conditions (temperature, wind speed, rain), and soil properties (calcium content, cation exchange capacity, acidity). The risk for high NH 3 losses may be reduced by proper management strategies including, e.g., incorporation of the fertilizer into the soil, use of acidic fertilizers on calcareous soils, use of fertilizers with a high content of carbonate-precipitating cations, split applications to rice paddies or application to the soil surface beneath the crop canopy. The latter takes advantage of the relatively low wind speed within well-developed canopies, reducing the rate of vertical NH 3 transport and increasing foliar NH 3 absorption. Conversely, NH 3 is emitted from the leaves when the internal NH 3 concentration is higher than that in the ambient atmosphere as may often be the case, particularly during periods with rapid N absorption by the roots or during senescence induced N-remobilization from leaves. Between 1 and 4% of shoot N may be lost in this way.

Functions of Macronutrients

Publisher Summary This chapter focuses on the role played by various macronutrients such as nitrogen (N), sulfur (S), phosphorus (P), magnesium (Mg), calcium (Ca), and potassium (K) in plant metabolism and growth and describes the symptoms of deficiency and toxicity of these macronutrients. N is the most essential element required after carbon, and it plays a central role in plant metabolism as a constituent of proteins, nucleic acids, chlorophyll, coenzymes, phytohormones, and secondary metabolites. When it is taken as ammonium or nitrate, it is assimilated into amino acids either in the roots or shoots and within the plant, it is translocated as nitrate or amino acids. Sulfur is taken up as sulphate and assimilated into S-containing amino acids such as cysteine that are used to synthesize S-containing enzymes and coenzymes as well as secondary compounds such as phytochelatins (detoxification of metals) or aliins and glucosinolates (feeding deterrents). Phosphorus is a structural element in nucleic acids, and as a component of adenosine phosphates, it plays an important role in energy transfer, and it is also essential for transfer of carbohydrates in leaf cells. Magnesium is a component of chlorophyll, and it is required for photosynthesis and protein synthesis. Calcium is important for cell wall and membrane stabilization, osmoregulation, and as second messenger, thereby allowing plants to regulate developmental processes in response to environmental stimuli. The main role of K is osmoregulation, which is important for cell extension and stomata movement, and it affects loading of sucrose and the rate of mass flow-driven solute movement within the plant.

Apoplastic pH and Ammonium Concentration in Leaves of Brassica napus L

A vacuum infiltration technique was developed that enabled the extraction of apoplastic solution with very little cytoplasmic contamination as evident from a malate dehydrogenase activity of less than 1% in the apoplastic solution relative to that in bulk leaf extracts. The volume of apoplastic water, a prerequisite for determination of the concentration of apoplastic solutes, was determined by vacuum infiltration of indigo carmine with subsequent analysis of the dilution of the dye in apoplastic extracts. Indigo carmine was neither transported across the cell membrane nor significantly adsorbed to the cell walls, ensuring reproducible (SE < 2%) and precise determination of apoplastic water. Analysis of leaves from four different positions on senescing Brassica napus plants showed a similar apoplastic pH of 5.8, while apoplastic NH4+ increased from 1.1 mM in lower leaves to 1.3 mM in upper leaves. Inhibition of glutamine synthetase in young B. napus plants resulted in increasing apoplastic pH from 6.0 to 6.8 and increasing apoplastic NH4+ concentration from 1.0 to 25.6 mM, followed by a marked increase in NH3 emission. Calculating NH3 compensation points for B. napus plants on the basis of measured apoplastic H+ and NH4+ concentrations gave values ranging from 4.3 to 5.9 nmol NH3 mol-1 air, consistent with an estimate of 5.3 [plus or minus] 3.6 nmol NH3 mol-1 air obtained by NH3 exchange experiments in growth chambers. A strong linear relationship was found between calculated NH3 compensation points and measured NH3 emission rates in glutamine synthetase-inhibited plants.

Rhizodeposition of nitrogen by red clover, white clover and ryegrass leys

Correct assessment of the rhizodeposition of N in grassland is essential for the evaluation of biological N2-fixation of legumes, for the total N balance of agro-ecosystems, and for the pre-cropping value of grasslands. Using a leaf-feeding technique by which plants were 15 N labelled while growing in mezotrons in the field, the rhizodeposition of N by unfertilised red clover, white clover and perennial ryegrass growing in pure stands was shown to amount to 64, 71 and 9 g N m 22 , respectively, over two complete growing seasons. The corresponding values for red clover and white clover growing in mixtures with ryegrass were 89 and 32 g N m 22 , respectively. The rhizodeposited N compounds, including fine roots, constituted more than 80% of the total plant-derived N in the soil, and in all cases exceeded the amount of N present in stubble. In the mixtures of red clover‐ryegrass and white clover‐ryegrass and the pure stands of red clover, white clover and ryegrass, respectively, the rhizodeposition constituted a 1.05, 1.52, 1.26, 2.21 and 2.77 fold increase over the total N in the shoots harvested during the two production years. In pure stands and mixtures of clover, 84 and 92%, respectively, of this N derived from biological N2 fixation. It is concluded that rhizodeposition provides a very substantial input of N to the legume-based grassland systems with great consequences for ecosystem N balance and turnover. Furthermore, the amount of atmospheric-derived N in the rhizodeposits may exceed that in the harvested shoots. q 2001 Elsevier Science Ltd. All rights reserved.

Interactions between white clover and ryegrass under contrasting nitrogen availability: N2 fixation, N fertilizer recovery, N transfer and water use efficiency

Seasonal variation in N2 fixation, N transfer from clover to ryegrass, and soil N absorption in white clover–ryegrass swards were investigated under field conditions over three consecutive years. The plots were established with different seeding ratios of clover and ryegrass and contrasting fertilizer N ranging from 3 to 72 kg ha-1 year-1.An initially poor clover population needed at least one growing season to reach the same yield output as an initially well established clover population. The clover content of the sward decreased by the annual application of 72 kg N ha-1 but not by smaller N dressings.The total amount of atmospherically derived N in clover growing in mixture with ryegrass was, on average over the three years equal to 83, 71, 68 and 60 kg N ha-1 for the treatments of 3, 24, 48 and 72 kg N ha-1, respectively. The proportion of atmospherically derived N declined with increasing N application, but never became smaller than 80% of total clover N. The proportion of atmospherically derived N in a pure stand white clover amounted to 60–80% of the total N content, equivalent to 109, 110, 103 and 90 kg N ha-1 for the treatments of 3, 24, 48 and 72 kg N ha-1, respectively.Only small amounts of atmospherically derived N was transferred to the associated ryegrass during the first production year, while in each of the following years up to 21 kg ha-1 was transferred. The average amount of N transferred from clover to ryegrass was equivalent to 3, 16 and 31% of the N accumulated in ryegrass in the first, second and third production year, respectively. Expressed relative to the total amount of fixed N2 in the clover–ryegrass mixture, the transfer amounted to 3, 17 and 22% in the first, second and third production year, respectively. Thus transfer of atmospherically derived N from clover contributed significantly to the N economy of the associated ryegrass.The clover–ryegrass mixture absorbed constantly higher amount of soil derived N than the pure stands of the two species. Only 11% of the total accumulated fertilizer N and soil derived N in the mixture was contained within the clover component. Lower water use efficiencies for the plants grown in mixture compared to pure stands were mainly related to the increased N uptake in the mixture, with the subsequent increase in growth compared to the pure stands.It is concluded that positive interactions between clover and ryegrass growing in mixture ensure a more efficient fixation of atmospheric N2 and absorption of fertilizer N and soil derived N than pure stands of the same species.

Elevated Nicotianamine Levels in Arabidopsis halleri Roots Play a Key Role in Zinc Hyperaccumulation

The hyperaccumulation of micronutrients and toxic metals (such as zinc and cadmium, respectively) represents an extreme trait of metallophytes adapted to metal-rich environments. This work demonstrates that elevated production of the metal chelator nicotianamine, specifically in roots of the metallophyte Arabidopsis halleri, is important for efficient root-to-shoot translocation of zinc. Zn deficiency is among the leading health risk factors in developing countries. Breeding of Zn-enriched crops is expected to be facilitated by molecular dissection of plant Zn hyperaccumulation (i.e., the ability of certain plants to accumulate Zn to levels >100-fold higher than normal plants). The model hyperaccumulators Arabidopsis halleri and Noccaea caerulescens share elevated nicotianamine synthase (NAS) expression relative to nonaccumulators among a core of alterations in metal homeostasis. Suppression of Ah-NAS2 by RNA interference (RNAi) resulted in strongly reduced root nicotianamine (NA) accumulation and a concomitant decrease in root-to-shoot translocation of Zn. Speciation analysis by size-exclusion chromatography coupled to inductively coupled plasma mass spectrometry showed that the dominating Zn ligands in roots were NA and thiols. In NAS2-RNAi plants, a marked increase in Zn-thiol species was observed. Wild-type A. halleri plants cultivated on their native soil showed elemental profiles very similar to those found in field samples. Leaf Zn concentrations in NAS2-RNAi lines, however, did not reach the Zn hyperaccumulation threshold. Leaf Cd accumulation was also significantly reduced. These results demonstrate a role for NAS2 in Zn hyperaccumulation also under near-natural conditions. We propose that NA forms complexes with Zn(II) in root cells and facilitates symplastic passage of Zn(II) toward the xylem.

Below-ground nitrogen transfer between different grassland species : Direct quantification by 15N leaf feeding compared with indirect dilution of soil 15N

Nitrogen (N) transfer from one species to another is important for the N cycling in low-input grassland. In the present work, estimates obtained by an indirect 15N dilution technique were compared with estimates obtained by a direct 15N leaf feeding technique over two complete growing seasons in red clover-ryegrass and white clover-ryegrass mixtures under field conditions.The direct technique confirmed that N transfer between clovers and ryegrass is a bi-directional process. The transfer of N from both clovers to ryegrass occurred within 25 days upon the first labelling event. A very high N transfer occurred from white clover to the associated ryegrass, 4.5 and 7.5 g m−2 in the 1st and 2nd production year, respectively. The corresponding values for transfer from red clover to the associated ryegrass were 1.7 and 3.6 g m−2. Quantified relatively to the total above-ground N content of white clover- ryegrass and red clover-ryegrass mixtures, the N transfer exceeded 50% and 10%, respectively, in three out of seven harvests. The N transfer from 15N labelled grass to associated clovers constituted a relatively constant proportion of approx. 8% of the above-ground N content of the mixtures.Estimates based on the soil 15N dilution technique generally underestimated the net N transfer by more than 50% compared to the direct 15N labelling technique. Furthermore, the indirect 15N dilution technique estimated only marginal differences between red and white clover in the quantities of N transferred, whereas the direct 15N labelling technique showed the N transfer from white clover to the associated ryegrass to be significantly higher than that involving red clover. It is concluded that N transfer is a much more dynamic and quantitatively important process in grassland than previously recognised.