Julian C. Theobald

Rhizosphere bacteria containing 1‐aminocyclopropane‐1‐carboxylate deaminase increase yield of plants grown in drying soil via both local and systemic hormone signalling

Decreased soil water availability can stimulate production of the plant hormone ethylene and inhibit plant growth. Strategies aimed at decreasing stress ethylene evolution might attenuate its negative effects. An environmentally benign (nonchemical) method of modifying crop ethylene relations - soil inoculation with a natural root-associated bacterium Variovorax paradoxus 5C-2 (containing the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase that degrades the ethylene precursor ACC), was assessed with pea (Pisum sativum) plants grown in drying soil. Inoculation with V. paradoxus 5C-2, but not with a transposome mutant with massively decreased ACC deaminase activity, improved growth, yield and water-use efficiency of droughted peas. Systemic effects of V. paradoxus 5C-2 included an amplified soil drying-induced increase of xylem abscisic acid (ABA) concentration, but an attenuated soil drying-induced increase of xylem ACC concentration. A local bacterial effect was increased nodulation by symbiotic nitrogen-fixing bacteria, which prevented a drought-induced decrease in nodulation and seed nitrogen content. Successfully deploying a single bacterial gene in the rhizosphere increased yield and nutritive value of plants grown in drying soil, via both local and systemic hormone signalling. Such bacteria may provide an easily realized, economic means of sustaining crop yields and using irrigation water more efficiently in dryland agriculture.

The Identification of Genes Involved in the Stomatal Response to Reduced Atmospheric Relative Humidity

Stomatal pores of higher plants close in response to decreases in atmospheric relative humidity (RH). This is believed to be a mechanism that prevents the plant from losing excess water when exposed to a dry atmosphere and as such is likely to have been of evolutionary significance during the colonization of terrestrial environments by the embryophytes. We have conducted a genetic screen, based on infrared thermal imaging, to identify Arabidopsis genes involved in the stomatal response to reduced RH. Here we report the characterization of two genes, identified during this screen, which are involved in the guard cell reduced RH signaling pathway. Both genes encode proteins known to be involved in guard cell ABA signaling. OST1 encodes a protein kinase involved in ABA-mediated stomatal closure while ABA2 encodes an enzyme involved in ABA biosynthesis. These results suggest, in contrast to previously published work, that ABA plays a role in the signal transduction pathway connecting decreases in RH to reductions in stomatal aperture. The identification of OST1 as a component required in stomatal RH and ABA signal transduction supports the proposition that guard cell signaling is organized as a network in which some intracellular signaling proteins are shared among different stimuli.

Biomass allocation in tomato (Lycopersicon esculentum) plants grown under partial rootzone drying: enhancement of root growth

Tomato (Lycopersicon esculentum Mill.) plants were grown in either a glasshouse (GH) or a controlled environment cabinet (CEC) to assess the effects of partial rootzone drying (PRD) on biomass allocation. Control and PRD plants received the same amounts of water. In control plants, water was equally distributed between two compartments of a split-root system. In PRD plants, only one compartment was watered while the other was allowed to dry. At the end of each drying cycle, wet and dry compartments were alternated. In the GH, total biomass did not differ between PRD and control plants after four cycles of PRD, but PRD increased root biomass by 55% as resources were partitioned away from shoot organs. In the CEC, leaf water potential did not differ between treatments at the end of either of two cycles of PRD, but stomatal conductance of PRD plants was 20% less at the end of the first cycle than at the beginning. After two cycles of PRD in the CEC, biomass did not differ between PRD and control plants, but PRD increased root biomass by 19% over the control plants. The promotion of root biomass in PRD plants was associated with the alternation of wet and dry compartments, with increased root biomass occurring in the re-watered compartment after previous exposure to soil drying. Promotion of root biomass in field-grown PRD plants may allow the root system to access resources (water and nutrients) that would otherwise be unavailable to control plants. This may contribute to the ability of PRD plants to maintain similar leaf water potentials to conventionally irrigated plants, even when smaller irrigation volumes are supplied.

Alternation of wet and dry sides during partial rootzone drying irrigation alters root-to-shoot signalling of abscisic acid.

Partial rootzone drying (PRD) is an irrigation technique where water is distributed unevenly to the root system such that part is irrigated while the remainder is allowed to dry the soil. Tomato (Lycopersicon esculentum Mill.) plants were grown with their roots in two soil columns to compare the physiological consequences of alternation of wet and dry columns during PRD irrigation (alternate PRD, PRD-A) with retention of the same wet and dry columns (fixed PRD, PRD-F). When PRD plants received 50% less water than well-watered (WW) plants, xylem ABA concentration ([X-ABA]) increased and stomatal conductance decreased relative to WW plants. Although both sets of PRD plants received the same amount of water, [X-ABA] of PRD-A plants increased up to 2-fold above that of PRD-F plants, which further decreased stomatal conductance. Differences in [X-ABA] were detected within an hour of alternation, but did not persist beyond the photoperiod of alternation. [X-ABA] increased linearly as whole-pot soil water content (θpot) and leaf water potential (Ψleaf) declined, but the difference in [X-ABA] between the two sets of PRD plants was not due to differences in either θpot or Ψleaf. In PRD-F plants, the unwatered part of the root system contributes proportionally less to the transpiration stream as the soil progressively dries (Yao et al. 2001, Plant, Cell & Environment 24, 227-235). In PRD-A plants, we hypothesise that re-watering the dry part of the root system allows these roots to contribute proportionally more to total sap flux, thus liberating a pulse of ABA to the transpiration stream as the root ABA pool accumulated during soil drying is depleted. Since the enhancement of [X-ABA] caused by PRD-A increased as θpot and Ψleaf declined, an optimal frequency of alternation to maximise the cumulative physiological effects of this ABA pulse must consider possible negative impacts of leaf water deficit as soil water status declines.

Partial phenotypic reversion of ABA-deficient flacca tomato (Solanum lycopersicum) scions by a wild-type rootstock: normalizing shoot ethylene relations promotes leaf area but does not diminish whole plant transpiration rate

To evaluate the role of root-synthesized ABA in regulating growth and stomatal behaviour under well-watered conditions, isogenic wild-type (WT) and ABA-deficient flacca (flc) tomato (Solanum lycopersicum) were reciprocally and self-grafted just below the cotyledonary node. Since flc scions had lower leaf water potentials due to higher transpiration rates, a subset of all graft combinations was grown under a shoot misting treatment to minimize differences in shoot water status. Misting did not alter the relative effects of the different graft combinations on leaf area. WT scions had the greatest leaf area and lowest whole plant transpiration rate irrespective of the rootstock, implying that shoot ABA biosynthesis was sufficient to account for a WT shoot phenotype. In WT scions, the rootstock had no effect on detached leaf ethylene evolution or xylem concentrations of ABA or the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC). In flc scions, although the WT rootstock suppressed stomatal conductance of individual leaves, there was no detectable effect on whole plant transpiration rate. However, leaf area of flc/WT (scion/rootstock) plants increased 1.6-fold compared to flc self-grafts. WT rootstocks increased xylem ABA concentration in flc scions (relative to flc self-grafts) up to 3-fold, and resulted in xylem ACC concentrations and detached leaf ethylene evolution similar to WT scions. Since the WT rootstock normalized shoot ethylene relations but only partially restored the leaf area of flc scions (relative to that of WT scions), shoot ABA biosynthesis can directly promote leaf area via an unknown, ethylene-independent, mechanism.

The rhizobacterium Variovorax paradoxus 5C-2, containing ACC deaminase, promotes growth and development of Arabidopsis thaliana via an ethylene-dependent pathway

Many plant-growth-promoting rhizobacteria (PGPR) associated with plant roots contain the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase and can metabolize ACC, the immediate precursor of the plant hormone ethylene, thereby decreasing plant ethylene production and increasing plant growth. However, relatively few studies have explicitly linked ethylene emission and/or action to growth promotion in these plant–microbe interactions. This study examined effects of the PGPR Variovorax paradoxus 5C-2 containing ACC deaminase on the growth and development of Arabidopsis thaliana using wild-type (WT) plants and several ethylene-related mutants (etr1-1, ein2-1, and eto1-1). Soil inoculation with V. paradoxus 5C-2 promoted growth (leaf area and shoot biomass) of WT plants and the ethylene-overproducing mutant eto1-1, and also enhanced floral initiation of WT plants by 2.5 days. However, these effects were not seen in ethylene-insensitive mutants (etr1-1 and ein2-1) even though bacterial colonization of the root system was similar. Furthermore, V. paradoxus 5C-2 decreased ACC concentrations of rosette leaves of WT plants by 59% and foliar ethylene emission of both WT plants and eto1-1 mutants by 42 and 37%, respectively. Taken together, these results demonstrate that a fully functional ethylene signal transduction pathway is required for V. paradoxus 5C-2 to stimulate leaf growth and flowering of A. thaliana.

Photosynthetic responses in spring wheat grown under elevated CO2 concentrations and stress conditions in the European, multiple-site experiment 'ESPACE-wheat'

Spring wheat cv. Minaret crop stands were grown under ambient and elevated CO2 concentrations at seven sites in Germany, Ireland, the UK, Belgium and the Netherlands. Six of the sites used open-top chambers and one used a controlled environment mimicking field conditions. The effect of elevated CO2 for a range of N application regimes, O3 concentrations, and growth temperatures on flag leaf photosynthesis was studied. Before anthesis, flag leaf photosynthesis was stimulated about 50% by 650 compared with 350 mmol mol−1 CO2 at all sites, regardless of other treatments. Furthermore, there was no evidence of a decrease in photosynthetic capacity of flag leaves due to growth at elevated CO2 before anthesis, even for low N treatments. However, photosynthetic capacity, particularly carboxylation capacity, of flag leaves was usually decreased by growth at elevated CO2 after anthesis, especially in low N treatments. Acclimation of photosynthesis to elevated CO2 therefore appears to occur only slowly, consistent with a response to changes in sink–source relationships, rather than a direct response. Effect of elevated CO2 on stomatal conductance was much more variable between sites and treatments, but on average was decreased by ~10% at 650 compared with 350 mmol mol−1 CO2. Carboxylation capacity of flag leaves was decreased by growth at elevated O3 both before and after anthesis, regardless of CO2 concentration.

Xylem sap collection and extraction methodologies to determine in vivo concentrations of ABA and its bound forms by gas chromatography-mass spectrometry (GC-MS)

BackgroundAccurate quantification of xylem sap ABA concentrations is important to underpin models of root-to-shoot ABA signalling to predict the physiological effects of soil drying. Growing tomato plants in a whole plant pressure chamber allowed sequential xylem sap collection from a detached leaf, the petiole stub of an otherwise intact plant and finally the de-topped root system of the same plant, to determine the impact of xylem sap sampling methodology on xylem ABA concentration. Since xylem sap can contain bound forms of ABA, a novel gas chromatography-mass spectrometry (GC-MS) procedure was developed to chemically separate free ABA from two in planta bound ABA forms known as Adducts I and II and ABA-glucose-ester (ABA-GE).ResultsXylem sap ABA concentrations were highly dependent on the sampling methodology used: the highest concentrations were detected in sap collected by applying an overpressure to detached leaves following the measurement of leaf water potential. Irrespective of xylem sap source, the wild-type cultivars Ailsa Craig and Rheinlands Ruhm had higher free ABA concentrations than a range of ABA-deficient mutants (notabilis, flacca and sitiens). However, in the mutants, concentrations of bound forms of ABA were similar to wild-type plants, and similar to free ABA concentrations.ConclusionsAlthough xylem concentrations of these bound ABA forms and ABA-GE suggest they have a limited physiological impact on ABA homeostasis in tomato, the methods developed here will allow a more complete understanding of ABA biochemistry and root-to-shoot signalling in species known to have higher concentrations of these compounds.

Egg load dynamics and the risk of egg and time limitation experienced by an aphid parasitoid in the field.

Insect parasitoids and herbivores must balance the risk of egg limitation and time limitation in order to maximize reproductive success. Egg and time limitation are mediated by oviposition and egg maturation rates as well as by starvation risk and other determinants of adult lifespan. Here, we assessed egg load and nutritional state in the soybean aphid parasitoid Binodoxys communis under field conditions to estimate its risk of becoming either egg- or time-limited. The majority of female B. communis showed no signs of egg limitation. Experimental field manipulations of B. communis females suggested that an average of 4–8 eggs were matured per hour over the course of a day. Regardless, egg loads remained constant over the course of the day at approximately 80 eggs, suggesting that egg maturation compensates for oviposition. This is the first case of such “egg load buffering” documented for a parasitoid in the field. Despite this buffering, egg loads dropped slightly with increasing host (aphid) density. This suggests that egg limitation could occur at very high host densities as experienced in outbreak years in some locations in the Midwestern USA. Biochemical analyses of sugar profiles showed that parasitoids fed upon sugar in the field at a remarkably high rate. Time limitation through starvation thus seems to be very low and aphid honeydew is most likely a source of dietary sugar for these parasitoids. This latter supposition is supported by the fact that body sugar levels increase with host (aphid) density. Together, these results suggest that fecundity of B. communis benefits from both dynamic egg maturation strategies and sugar-feeding.

An Arabidopsis mutant able to green after extended dark periods shows decreased transcripts of seed protein genes and altered sensitivity to abscisic acid

An Arabidopsis mutant showing an altered ability to green on illumination after extended periods of darkness has been isolated in a screen for genomes uncoupled (gun) mutants. Following illumination for 24 h, 10-day-old dark-grown mutant seedlings accumulated five times more chlorophyll than wild-type seedlings and this was correlated with differences in plastid morphology observed by transmission electron microscopy. The mutant has been named greening after extended darkness 1 (ged1). Microarray analysis showed much lower amounts of transcripts of genes encoding seed storage proteins, oleosins, and late embryogenesis abundant (LEA) proteins in 7-day-old seedlings of ged1 compared with the wild type. RNA gel-blot analyses confirmed very low levels of transcripts of seed protein genes in ged1 seedlings grown for 2–10 d in the dark, and showed higher amounts of transcripts of photosynthesis-related genes in illuminated 10-day-old dark-grown ged1 seedlings compared with the wild type. Consensus elements similar to abscisic acid (ABA) response elements (ABREs) were detected in the upstream regions of all genes highly affected in ged1. Germination of ged1 seeds was hypersensitive to ABA, although no differences in ABA content were detected in 7-day-old seedlings. This suggests the mutant may have an altered responsiveness to ABA, affecting expression of ABA-responsive genes and plastid development during extended darkness.