The barrier to radial oxygen loss impedes the apoplastic entry of iron into the roots of Urochloa humidicola.

Lack of O2 and high concentrations of iron (Fe) and manganese (Mn) commonly occur in waterlogged soils. The development of a barrier to impede radial O2 loss (ROL) is a key trait improving internal O2 transport and waterlogging tolerance in plants. We evaluated the ability of the barrier to ROL to impede the entry of excess Fe into the roots of the waterlogging-tolerant grass Urochloa humidicola. Plants were grown in aerated or stagnant deoxygenated nutrient solution with 5 µM or 900 µM Fe. Quantitative X-ray microanalysis was used to determine cell-specific Fe concentrations at two positions behind the root apex in relation to ROL and the formation of apoplastic barriers. At a mature zone of the root, Fe was 'excluded' at the exodermis where a suberized lamella was evident, a feature also associated with a strong barrier to ROL. In contrast, the potassium (K) concentration was similar in all root cells, indicating that K uptake was not affected by apoplastic barriers. The hypothesis that the formation of a tight barrier to ROL impedes the apoplastic entry of toxic concentrations of Fe into the mature zones of roots was supported by the significantly higher accumulation of Fe on the outer side of the exodermis.

Sensitivity of chickpea and faba bean to root-zone hypoxia, elevated ethylene, and carbon dioxide.

During soil waterlogging, plants experience O2 deficits, elevated ethylene, and high CO2 in the root-zone. The effects on chickpea (Cicer arietinum L.) and faba bean (Vicia faba L.) of ethylene (2 µL L-1 ), CO2 (2-20% v/v) or deoxygenated stagnant solution were evaluated. Ethylene and high CO2 reduced root growth of both species, but O2 deficiency had the most damaging effect and especially so for chickpea. Chickpea suffered root tip death when in deoxygenated stagnant solution. High CO2 inhibited root respiration and reduced growth, whereas sugars accumulated in root tips, of both species. Gas-filled porosity of the basal portion of the primary root of faba bean (23%, v/v) was greater than for chickpea (10%), and internal O2 movement was more prominent in faba bean when in an O2 -free medium. Ethylene treatment increased the porosity of roots. The damaging effects of low O2 , such as death of root tips, resulted in poor recovery of root growth upon reaeration. In conclusion, ethylene and high CO2 partially inhibited root extension in both species, but low O2 in deoxygenated stagnant solution had the most damaging effect, even causing death of root tips in chickpea, which was more sensitive to the low O2 condition than faba bean.

Tolerance to partial and complete submergence in the forage legume Melilotus siculus: an evaluation of 15 accessions for petiole hyponastic response and gas-filled spaces, leaf hydrophobicity and gas films, and root phellem.

Background and Aims: Submergence is a severe stress for most plants. Melilotus siculus is a waterlogging- (i.e. root zone hypoxia) tolerant annual forage legume, but data were lacking for the effects of partial and full submergence of the shoots. The aim was to compare the tolerance to partial and full submergence of 15 M. siculus accessions and to assess variation in traits possibly contributing to tolerance. Recovery ability post-submergence was also evaluated. Methods: A factorial experiment imposed treatments of water level [aerated root zone with shoots in air as controls, stagnant root zone with shoots in air, stagnant root zone with partial (75 %) or full shoot submergence] on 15 accessions, for 7 d on 4-week-old plants in a 20/15 °C day/night phytotron. Measurements included: shoot and root growth, hyponastic petiole responses, petiole gas-filled spaces, leaflet sugars, leaflet surface hydrophobicity, leaflet gas film thickness and phellem area near the base of the main root. Recovery following full submergence was also assessed. Key Results: Accessions differed in shoot and root growth during partial and full shoot submergence. Traits differing among accessions and associated with tolerance were leaflet gas film thickness upon submergence, gas-filled spaces in petioles and phellem tissue area near the base of the main root. All accessions were able to re-orientate petioles towards the vertical under both partial and full submergence. Petiole extension rates were maintained during partial submergence, but decreased during full submergence. Leaflet sugars accumulated during partial submergence, but were depleted during full submergence. Growth resumption after full submergence differed among accessions and was positively correlated with the number of green leaves retained at desubmergence. Conclusions: Melilotus siculus is able to tolerate partial and full submergence of at least 7 d. Leaflet surface hydrophobicity and associated gas film retention, petiole gas-filled porosity and root phellem abundance are important traits contributing to tolerance. Post-submergence recovery growth differs among accessions. The ability to retain green leaves is essential to succeed during recovery.

Leaf gas films, underwater photosynthesis and plant species distributions in a flood gradient.

Traits for survival during flooding of terrestrial plants include stimulation or inhibition of shoot elongation, aerenchyma formation and efficient gas exchange. Leaf gas films form on superhydrophobic cuticles during submergence and enhance underwater gas exchange. The main hypothesis tested was that the presence of leaf gas films influences the distribution of plant species along a natural flood gradient. We conducted laboratory experiments and field observations on species distributed along a natural flood gradient. We measured presence or absence of leaf gas films and specific leaf area of 95 species. We also measured, gas film retention time during submergence and underwater net photosynthesis and dark respiration of 25 target species. The presence of a leaf gas film was inversely correlated to flood frequency and duration and reached a maximum value of 80% of the species in the rarely flooded locations. This relationship was primarily driven by grasses that all, independently of their field location along the flood gradient, possess gas films when submerged. Although the present study and earlier experiments have shown that leaf gas films enhance gas exchange of submerged plants, the ability of species to form leaf gas films did not show the hypothesized relationship with species composition along the flood gradient.

Efficient use of energy in anoxia-tolerant plants with focus on germinating rice seedlings.

Anoxia tolerance in plants is distinguished by direction of the sparse supply of energy to processes crucial to cell maintenance and sometimes to growth, as in rice seedlings. In anoxic rice coleoptiles energy is used to synthesise proteins, take up K(+) , synthesise cell walls and lipids, and in cell maintenance. Maintenance of electrochemical H(+) gradients across the tonoplast and plasma membrane is crucial for solute compartmentation and thus survival. These gradients sustain some H(+) -solute cotransport and regulate cytoplasmic pH. Pyrophosphate (PPi ), the alternative energy donor to ATP, allows direction of energy to the vacuolar H(+) -PPi ase, sustaining H(+) gradients across the tonoplast. When energy production is critically low, operation of a biochemical pHstat allows H(+) -solute cotransport across plasma membranes to continue for at least for 18 h. In active (e.g. growing) cells, PPi produced during substantial polymer synthesis allows conversion of PPi to ATP by PPi -phosphofructokinase (PFK). In quiescent cells with little polymer synthesis and associated PPi formation, the PPi required by the vacuolar H(+) -PPi ase and UDPG pyrophosphorylase involved in sucrose mobilisation via sucrose synthase might be produced by conversion of ATP to PPi through reversible glycolytic enzymes, presumably pyruvate orthophosphate dikinase. These hypotheses need testing with species characterised by contrasting anoxia tolerance.

Gas film retention and underwater photosynthesis during field submergence of four contrasting rice genotypes.

Floods can completely submerge some rice (Oryza sativa L.) fields. Leaves of rice have gas films that aid O2 and CO2 exchange under water. The present study explored the relationship between gas film persistence and underwater net photosynthesis (PN) as influenced by genotype and submergence duration. Four contrasting genotypes (FR13A, IR42, Swarna, and Swarna-Sub1) were submerged for 13 days in the field and leaf gas films, chlorophyll, and the capacity for underwater PN at near ambient and high CO2 were assessed with time of submergence. At high CO2 during the PN assay, all genotypes initially showed high rates of underwater PN, and this rate was not affected by time of submergence in FR13A. This superior photosynthetic performance of FR13A was not evident in Swarna-Sub1 (carrying the SUB1 QTL) and the declines in underwater PN in both Swarna-Sub1 and Swarna were equal to that in IR42. At near ambient CO2 concentration, underwater PN declined in all four genotypes and this corresponded with loss of leaf gas films with time of submergence. FR13A retained leaf gas films moderately longer than the other genotypes, but gas film retention was not linked to SUB1. Diverse rice germplasm should be screened for gas film persistence during submergence, as this trait could potentially increase carbohydrate status and internal aeration owing to increased underwater PN, which contributes to submergence tolerance in rice.

The mechanism of improved aeration due to gas films on leaves of submerged rice.

Some terrestrial wetland plants, such as rice, have super-hydrophobic leaf surfaces which retain a gas film when submerged. O2 movement through the diffusive boundary layer (DBL) of floodwater, gas film and stomata into leaf mesophyll was explored by means of a reaction-diffusion model that was solved in a three-dimensional leaf anatomy model. The anatomy and dark respiration of leaves of rice (Oryza sativa L.) were measured and used to compute O2 fluxes and partial pressure of O2 (pO2 ) in the DBL, gas film and leaf when submerged. The effects of floodwater pO2 , DBL thickness, cuticle permeability, presence of gas film and stomatal opening were explored. Under O2 -limiting conditions of the bulk water (pO2 < 10 kPa), the gas film significantly increases the O2 flux into submerged leaves regardless of whether stomata are fully or partly open. With a gas film, tissue pO2 substantially increases, even for the slightest stomatal opening, but not when stomata are completely closed. The effect of gas films increases with decreasing cuticle permeability. O2 flux and tissue pO2 decrease with increasing DBL thickness. The present modelling analysis provides a mechanistic understanding of how leaf gas films facilitate O2 entry into submerged plants.

Alternative oxidase, a determinant of plant gametophyte fitness and fecundity.

The alternative oxidase (AOX) is the terminal oxidase that comprises the cyanide-resistant respiratory pathway in plant mitochondria. While the role of AOX in plant thermogenesis is well established, its role in the reproductive development of non-thermogenic species is not well understood. AOX genes can be separated into two groups based on sequence homology, AOX1 and AOX2. Reverse genetic experiments carried out primarily in Arabidopsis and tobacco have largely focussed on examining the role of AOX1-type genes in stress responses. We recently reported a systematic characterisation of the reproductive phenotypes of three AOX2 antisense lines of soybean. This addendum summarises the key evidence in our recent paper that points to a role for AOX in the development and function of both male and female gametophytes. Furthermore, we discuss the relative importance of AOX in the reproductive biology of plant species examined to date and highlight practical implications of our findings to crop improvement research.

Does anoxia tolerance involve altering the energy currency towards PPi?

Oxygen deficit is an important abiotic stress influencing plants, because this condition results in an 'energy crisis'. Most species only survive short periods of anoxia, but several wetland species tolerate prolonged anoxia. Transcriptomic and proteomic studies, using anoxia-tolerant rice and anoxia-intolerant Arabidopsis, have provided evidence for the selective adoption of pyrophosphate (PPi) over ATP as high-energy donor molecules, which may contribute to anoxia tolerance. The use of PPi in some tolerant plant species is similar to that observed in many anaerobic prokaryotes. Investigations are being performed to better understand the origin and regulation of reversible PPi-dependent glycolytic enzymes such as cytosolic pyruvate phosphate dikinase, as well as PPi-consuming enzymes, which are engaged during the anoxic energy crisis. This will be crucial in unraveling this currency switch and its contribution to anoxia tolerance.

Use of wild relatives to improve salt tolerance in wheat.

There is considerable variability in salt tolerance amongst members of the Triticeae, with the tribe even containing a number of halophytes. This is a review of what is known of the differences in salt tolerance of selected species in this tribe of grasses, and the potential to use wild species to improve salt tolerance in wheat. Most investigators have concentrated on differences in ion accumulation in leaves, describing a desirable phenotype with low leaf Na+ concentration and a high K+/Na+ ratio. Little information is available on other traits (such as "tissue tolerance" of accumulated Na+ and Cl-) that might also contribute to salt tolerance. The sources of Na+ "exclusion" amongst the various genomes that make up tetraploid (AABB) durum wheat (Triticum turgidum L. ssp. durum), hexaploid (AABBDD) bread wheat (Triticum aestivum L. ssp. aestivum), and wild relatives (e.g. Aegilops spp., Thinopyrum spp., Elytrigia elongata syn. Lophopyrum elongatum, Hordeum spp.) are described. The halophytes display a capacity for Na+ "exclusion", and in some cases Cl- "exclusion", even at relatively high salinity. Significantly, it is possible to hybridize several wild species in the Triticeae with durum and bread wheat. Progenitors have been used to make synthetic hexaploids. Halophytic relatives, such as tall wheatgrass spp., have been used to produce amphiploids, disomic chromosome addition and substitution lines, and recombinant lines in wheat. Examples of improved Na+ "exclusion" and enhanced salt tolerance in various derivatives from these various hybridization programmes are given. As several sources of improved Na+ "exclusion" are now known to reside on different chromosomes in various genomes of species in the Triticeae, further work to identify the underlying mechanisms and then to pyramid the controlling genes for the various traits, that could act additively or even synergistically, might enable substantial gains in salt tolerance to be achieved.

Ethylene regulates fast apoplastic acidification and expansin A transcription during submergence-induced petiole elongation in Rumex palustris.

The semi-aquatic dicot Rumex palustris responds to complete submergence by enhanced elongation of young petioles. This elongation of petiole cells brings leaf blades above the water surface, thus reinstating gas exchange with the atmosphere and increasing survival in flood-prone environments. We already know that an enhanced internal level of the gaseous hormone ethylene is the primary signal for underwater escape in R. palustris. Further downstream, concentration changes in abscisic acid (ABA), gibberellin (GA) and auxin are required to gain fast cell elongation under water. A prerequisite for cell elongation in general is cell wall loosening mediated by proteins such as expansins. Expansin genes might, therefore, be important target genes in submergence-induced and plant hormone-mediated petiole elongation. To test this hypothesis we have studied the identity, kinetics and regulation of expansin A mRNA abundance and protein activity, as well as examined pH changes in cell walls associated with this adaptive growth. We found a novel role of ethylene in triggering two processes affecting cell wall loosening during submergence-induced petiole elongation. First, ethylene was shown to promote fast net H(+) extrusion, leading to apoplastic acidification. Secondly, ethylene upregulates one expansin A gene (RpEXPA1), as measured with real-time RT-PCR, out of a group of 13 R. palustris expansin A genes tested. Furthermore, a significant accumulation of expansin proteins belonging to the same size class as RpEXPA1, as well as a strong increase in expansin activity, were apparent within 4-6 h of submergence. Regulation of RpEXPA1 transcript levels depends on ethylene action and not on GA and ABA, demonstrating that ethylene evokes at least three, parallel operating pathways that, when integrated at the whole petiole level, lead to coordinated underwater elongation. The first pathway involves ethylene-modulated changes in ABA and GA, these acting on as yet unknown downstream components, whereas the second and third routes encompass ethylene-induced apoplastic acidification and ethylene-induced RpEXPA1 upregulation.