Salt tolerance in mungbean is associated with controlling Na and Cl transport across roots, regulating Na and Cl accumulation in chloroplasts and maintaining high K in root and leaf mesophyll cells.

Salinity tolerance requires coordinated responses encompassing salt exclusion in roots and tissue/cellular compartmentation of salt in leaves. We investigated the possible control points for salt ions transport in roots and tissue tolerance to Na+ and Cl- in leaves of two contrasting mungbean genotypes, salt-tolerant Jade AU and salt-sensitive BARI Mung-6, grown in nonsaline and saline (75 mM NaCl) soil. Cryo-SEM X-ray microanalysis was used to determine concentrations of Na, Cl, K, Ca, Mg, P, and S in various cell types in roots related to the development of apoplastic barriers, and in leaves related to photosynthetic performance. Jade AU exhibited superior salt exclusion by accumulating higher [Na] in the inner cortex, endodermis, and pericycle with reduced [Na] in xylem vessels and accumulating [Cl] in cortical cell vacuoles compared to BARI Mung-6. Jade AU maintained higher [K] in root cells than BARI Mung-6. In leaves, Jade AU maintained lower [Na] and [Cl] in chloroplasts and preferentially accumulated [K] in mesophyll cells than BARI Mung-6, resulting in higher photosynthetic efficiency. Salinity tolerance in Jade AU was associated with shoot Na and Cl exclusion, effective regulation of Na and Cl accumulation in chloroplasts, and maintenance of high K in root and leaf mesophyll cells.

Low-Temperature X-ray Microanalysis Sheds New Light on Mineral Nutrition Aspects of Insect Leaf Galling.

Manipulation of host plant physiology by leaf-galling insects is a multifaceted process. Among fundamental knowledge gaps surrounding this scientifically intriguing phenomenon is the appropriation of plant mineral nutrients and moisture for galling advantage. Small, soluble mineral ions and watery cell contents in dense gall tissues risk disruption during routine sample preparations. In this study, an X-ray microanalysis was applied to investigate gall mineral nutrition. Morphologically diverse leaf galls were sampled from three Australian rainforest tree species. Using cryo-analytical scanning electron microscopy, real-time X-ray analytical maps of cellular mineral nutrients and water were integrated with anatomical images of gall and leaf cross-sectional surfaces. A comparison of host-leaf and gall anatomies bore direct evidence of drastic changes to leaf cells through the galling process. Distinct "wet" and "dry" regions within galls were anatomically and/or chemically differentiated, suggesting specific functionality. "Wet" regions comprising hydrated cells including soft gall-cavity linings where larvae are known to feed contained soluble plant mineral nutrients, while C-rich "dry" tissues largely devoid of mineral nutrients likely contribute structural support. Mapping immobile nutrients such as Mn may provide a means of "matching" specific gall cell types to those in ungalled host-leaf tissues. The findings here provided otherwise inaccessible insights into leaf-gall mineral nutrition.

De-MA.ch: A Web Database for Electron and X-ray Microanalysis to Assist Electron Microscope Lab Managers and Users.

Lab managers and users of scanning electron microscope or electron probe microanalyzer facilities aiming for qualitative or quantitative X-ray analyses require comprehensive, yet flexible documentation structures for their daily work and available reference material, with a complete X-ray data library, a repository of energy- and wavelength-dispersive spectra, and an instrument scheduling mechanism. An online multilaboratory database system available at https://de-ma.ch is presented with the primary goals of providing information on microanalytical reference materials, analytical setups, characteristic X-ray data, and for managing reservation and training requests. This website is designed for multiuser facilities, where experience ranges from beginners to expert users. Registered users will find these tools useful in developing and maintaining high-quality, reproducible, and efficient analyses, whereas lab managers will keep records of their microanalytical reference material database and analytical protocols. The database also serves an educational purpose by (a) providing information on reference materials, (b) encouraging students to select appropriate X-ray lines to analyze, (c) providing analytical setups for point analysis or mapping, (d) identifying unknown X-ray lines, (e) displaying energy- or wavelength-dispersive spectra, and (f) recalculating mineral formula from quantitative wt% analysis results, based on a number of oxygen atoms or cations.

Enhanced Na+ and Cl- sequestration and secretion selectivity contribute to high salt tolerance in the tetraploid recretohalophyte Plumbago auriculata Lam.

MAIN CONCLUSION: Enhanced secretion of Na+ and Cl- in leaf glands and leaf vacuolar sequestration of Na+ or root retention of Cl-, combined with K+ retention, contribute to the improved salt tolerance of tetraploid recretohalophyte P. auriculata. Salt stress is one of the major abiotic factors threatening plant growth and development, and polyploids generally exhibit higher salt stress resistance than diploids. In recretohalophytes, which secrete ions from the salt gland in leaf epidermal cells, the effects of polyploidization on ion homeostasis and secretion remain unknown. In this study, we compared the morphology, physiology, and ion homeostasis regulation of diploid and autotetraploid accessions of the recretohalophyte Plumbago auriculata Lam. after treatment with 300 mM NaCl for 0, 2, 4, 6, and 8 days. The results showed that salt stress altered the morphology, photosynthetic efficiency, and chloroplast structure of diploid P. auriculata to a greater extent than those of its tetraploid counterpart. Moreover, the contents of organic osmoregulatory substances (proline and soluble sugars) were significantly higher in the tetraploid than in the diploid, while those of H2O2 and malondialdehyde (MDA) were significantly lower. Analysis of ion homeostasis revealed that the tetraploid cytotype accumulated more Na+ in stems and leaves and more Cl- in roots but less K+ loss in roots compared with diploid P. auriculata. Additionally, the rate of Na+ and Cl- secretion from the leaf surface was higher, while that of K+, Mg2+, and Ca2+ secretion was lower in tetraploid plants. X-ray microanalysis of mesophyll cells revealed that Na+ mainly accumulated in different cellular compartments in the tetraploid (vacuole) and diploid (cytoplasm) plants. Our results suggest that polyploid recretohalophytes require the ability to sequester Na+ and Cl-(via accumulation in leaf cell vacuoles or unloading by roots) and selectively secrete these ions (through salt glands) together with the ability to prevent K+ loss (by roots). This mechanism required to maintain K+/Na+ homeostasis in polyploid recretohalophytes under high salinity provides new insights in the improved maintenance of ion homeostasis in polyploids under salt stress.

Differences in foliar phosphorus fractions, rather than in cell-specific phosphorus allocation, underlie contrasting photosynthetic phosphorus use efficiency among chickpea genotypes.

Although significant intraspecific variation in photosynthetic phosphorus (P) use efficiency (PPUE) has been shown in numerous species, we still know little about the biochemical basis for differences in PPUE among genotypes within a species. Here, we grew two high PPUE and two low PPUE chickpea (Cicer arietinum) genotypes with low P supply in a glasshouse to compare their photosynthesis-related traits, total foliar P concentration ([P]) and chemical P fractions (i.e. inorganic P (Pi), metabolite P, lipid P, nucleic acid P, and residual P). Foliar cell-specific nutrient concentrations including P were characterized using elemental X-ray microanalysis. Genotypes with high PPUE showed lower total foliar [P] without slower photosynthetic rates. No consistent differences in cellular [P] between the epidermis and mesophyll cells occurred across the four genotypes. In contrast, high PPUE was associated with lower allocation to Pi and metabolite P, with PPUE being negatively correlated with the percentage of these two fractions. Furthermore, a lower allocation to Pi and metabolite P was correlated with a greater allocation to nucleic acid P, but not to lipid P. Collectively, our results suggest that a different allocation to foliar P fractions, rather than preferential P allocation to specific leaf tissues, underlies the contrasting PPUE among chickpea genotypes.

Detection and Quantification of Extraterrestrial Platinum Group Element Alloy Micronuggets from Archean Impactite Deposits by Low-Voltage Scanning Electron Microscopy/Energy-Dispersive X-ray Spectrometry.

Rare, heterogeneously composed platinum group element alloy micronuggets (PGNs) occur in primitive meteorites, micrometeorites, and terrestrial impactite deposits. To gain insight into the nature of these phases, we developed a workflow for the characterization of PGNs using modern scanning electron microscopy (SEM) and energy-dispersive X-ray spectrometry at a low accelerating voltage of 6 kV. Automated feature analysis-a combination of morphological image analysis and elemental analysis with stage control-allowed us to detect PGNs down to 200 nm over a relatively large analysis area of 53 mm2 with a conventional silicon drift detector (SDD). Hyperspectral imaging with a high-sensitivity, annular SDD can be performed at low beam current (∼100 pA) which improves the SEM image resolution and minimizes hydrocarbon contamination. The severe overlapping peaks of the platinum group element L and M line families at 2-3 keV and the Fe and Ni L line families at <1 keV can be resolved by peak deconvolution. Quantitative elemental analysis can be performed at a spatial resolution of <80 nm; however, the results are affected by background subtraction errors for the Fe L line family. Furthermore, the inaccuracy of the matrix correction coefficients may influence standards-based quantification with pure element reference samples.

Quantification of Unsupported Thin-Film X-ray Spectra Using Bulk Standards.

A quantification model which uses standard X-ray spectra collected from bulk materials to determine the composition and mass thickness of single-layer and multilayer unsupported thin films is presented. The multivariate model can be iteratively solved for single layers in which each element produces at least one visible characteristic X-ray line. The model can be extended to multilayer thin films in which each element is associated with only one layer. The model may sometimes be solved when an element is present in multiple layers if additional information is added in the form of independent k-ratios or model assumptions. While the algorithm is suitable for any measured k-ratios, it is particularly well suited to energy-dispersive X-ray spectrometry where the bulk standard spectra can be used to deconvolve peak interferences in the thin-film spectra. The algorithm has been implemented and made available in the Open Source application National Institute of Standards and Technology DTSA-II. We present experimental data and Monte Carlo simulations supporting the quantification model.

Salt tolerance in relation to elemental concentrations in leaf cell vacuoles and chloroplasts of a C4 monocotyledonous halophyte.

Halophytes accumulate and sequester high concentrations of salt in vacuoles while maintaining lower levels of salt in the cytoplasm. The current data on cellular and subcellular partitioning of salt in halophytes are, however, limited to only a few dicotyledonous C3 species. Using cryo-scanning electron microscopy X-ray microanalysis, we assessed the concentrations of Na, Cl, K, Ca, Mg, P and S in various cell types within the leaf-blades of a monocotyledonous C4 halophyte, Rhodes grass (Chloris gayana). We also linked, for the first time, elemental concentrations in chloroplasts of mesophyll and bundle sheath cells to their ultrastructure and photosynthetic performance of plants grown in nonsaline and saline (200 mM NaCl) conditions. Na and Cl accumulated to the highest levels in xylem parenchyma and epidermal cells, but were maintained at lower concentrations in photosynthetically active mesophyll and bundle sheath cells. Concentrations of Na and Cl in chloroplasts of mesophyll and bundle sheath cells were lower than in their respective vacuoles. No ultrastructural changes were observed in either mesophyll or bundle sheath chloroplasts, and photosynthetic activity was maintained in saline conditions. Salinity tolerance in Rhodes grass is related to specific cellular Na and Cl distributions in leaf tissues, and the ability to regulate Na and Cl concentrations in chloroplasts.

Assessing the Impact of Secondary Fluorescence on X-Ray Microanalysis Results from Semiconductor Thin Films.

The impact of secondary fluorescence on the material compositions measured by X-ray analysis for layered semiconductor thin films is assessed using simulations performed by the DTSA-II and CalcZAF software tools. Three technologically important examples are investigated: AlxGa1−xN layers on either GaN or AlN substrates, InxAl1−xN on GaN, and Si-doped (SnxGa1−x)2O3 on Si. Trends in the differences caused by secondary fluorescence are explained in terms of the propensity of different elements to reabsorb either characteristic or bremsstrahlung X-rays and then to re-emit the characteristic X-rays used to determine composition of the layer under investigation. Under typical beam conditions (7­12 keV), the quantification of dopants/trace elements is found to be susceptible to secondary fluorescence and care must be taken to prevent erroneous results. The overall impact on major constituents is shown to be very small with a change of approximately 0.07 molar cation percent for Al0.3Ga0.7N/AlN layers and a maximum change of 0.08 at% in the Si content of (SnxGa1−x)2O3/Si layers. This provides confidence that previously reported wavelength-dispersive X-ray compositions are not compromised by secondary fluorescence.

Energy-Dispersive X-Ray Spectrum Simulation with NIST DTSA-II: Comparing Simulated and Measured Electron-Excited Spectra.

Electron-excited X-ray microanalysis with energy-dispersive spectrometry (EDS) proceeds through the application of the software that extracts characteristic X-ray intensities and performs corrections for the physics of electron and X-ray interactions with matter to achieve quantitative elemental analysis. NIST DTSA-II is an open-access, fully documented, and freely available comprehensive software platform for EDS quantification, measurement optimization, and spectrum simulation. Spectrum simulation with DTSA-II enables the prediction of the EDS spectrum from any target composition for a specified electron dose and for the solid angle and window parameters of the EDS spectrometer. Comparing the absolute intensities for measured and simulated spectra reveals correspondence within ±25% relative to K-shell and L-shell characteristic X-ray peaks in the range of 1­11 keV. The predicted M-shell intensity exceeds the measured value by a factor of 1.4­2.2 in the range 1­3 keV. The X-ray continuum (bremsstrahlung) generally agrees within ±10% over the range of 1­10 keV. Simulated EDS spectra are useful for developing an analytical strategy for challenging problems such as estimating trace detection levels.

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.

Role of roots in adaptation of soil-indifferent Proteaceae to calcareous soils in south-western Australia.

Very few of the >650 Proteaceae species in south-western Australia cope with the high calcium (Ca) levels in young, calcareous soils (soil indifferent); most are Ca sensitive and occur on nutrient-impoverished, acidic soils (calcifuge). We assessed possible control points for Ca transport across roots of two soil-indifferent (Hakea prostrata and Banksia prionotes) and two calcifuge (H. incrassata and B. menziesii) Proteaceae. Using quantitative X-ray microanalysis, we investigated cell-specific elemental Ca concentrations at two positions behind the apex in relation to development of apoplastic barriers in roots of plants grown in nutrient solution with low or high Ca supply. In H. prostrata, Ca accumulated in outer cortical cells at 20 mm behind the apex, but [Ca] was low in other cell types. In H. incrassata, [Ca] was low in all cells. Accumulation of Ca in roots of H. prostrata corresponded to development of apoplastic barriers in the endodermis. We found similar [Ca] profiles in roots and similar [Ca] in leaves of two contrasting Banksia species. Soil-indifferent Hakea and Banksia species show different strategies to inhabit calcareous soils: H. prostrata intercepts Ca in roots, reducing transport to shoots, whereas B. prionotes allocates Ca to specific leaf cells.

An x-ray microanalytical method for measuring in vivo element and water concentrations, relating to osmoregulation, in cells and tissues of the posterior eye.

Osmoregulation is critical for cell and tissue survival yet there are relatively few methods available to determine osmotic gradients from water and elemental concentration either in single cells or across multiple cellular layers of tissue. X-ray microanalysis of frozen-hydrated preparations in a scanning electron microscope is one such powerful, sensitive, nondestructive technique. Here we use x-ray microanalysis to quantitatively analyse intracellular element concentrations and oxygen concentrations, as a proxy for water concentrations, in selected individual cells of the posterior eye. Using frozen-hydrated preparations of the retinal complex of chicken eyes, it is shown that structural preservation is sufficient to identify cell layers and individual cells. The quantitative analysis of selected areas in the photoreceptor layer, inner nuclear layer and ganglion cell layer, where specific cell types were known to be present, provided measurements of intracellular element concentrations comparable with the analysis of individual cells. It is also shown that in the cells of the retinal pigment epithelium and outer photoreceptor segments elemental analyses were reasonably consistent at the cellular level in different depth levels of the same sample. Comparison of oxygen concentrations, as a proxy for water concentration, at two accelerating voltages (15 and 5 kV) indicated that at 15 kV oxygen concentration was largely derived from intracellular water. Water concentrations could be calculated and concentrations of diffusible elements (Na, K) could be defined in mmol/L. From the latter it is possible to calculate osmotic concentrations of individual cells and osmotic gradients across the tissue. LAY DESCRIPTION: Understanding many cellular processes, in both healthy and diseased states, depends on knowing how the water content of cells and their surrounding fluids is controlled. The transport of water is generally down its concentration gradient or against the osmotic concentration gradient defined by solutes such as sodium, potassium and chloride dissolved in the water. We have refined a microanalytical method, that detects the x-rays emitted from specific elements when they are bombarded by electrons in a scanning electron microscope, to apply it to the analysis of the retina of the eye. The method facilitates the measurement of the elemental composition, water and osmotic concentration gradients of cells and tissues in the eye, that may be involved in the development of myopia, or short sightedness, a condition that afflicts many people including some 80 - 90% of children in Asia.

Quantitative Electron-Excited X-ray Microanalysis With Low-Energy L-shell X-ray Peaks Measured With Energy-Dispersive Spectrometry.

Quantification of electron-exited X-ray spectra following the standards-based "k-ratio" (unknown/standard intensity) protocol with corrections for "matrix effects" (electron energy loss and backscattering, X-ray absorption, and secondary X-ray fluorescence) is a well-established method with a record of rigorous testing and extensive experience. Two recent studies by Gopon et al. working in the Fe­Si system and Llovet et al. working in the Ni­Si system have renewed interest in studying the accuracy of measurements made using L-shell X-ray peaks. Both have reported unexpectedly large deviations in analytical accuracy when analyzing intermetallic compounds when using the low photon energy Fe or Ni L-shell X-ray peaks with pure element standards and wavelength-dispersive X-ray spectrometry. This study confirms those observations on the Ni-based intermetallic compounds using energy-dispersive X-ray spectrometry and extends the study of analysis with low photon energy L-shell peaks to a wide range of elements, Ti to Se. Within this range of elements, anomalies in analytical accuracy have been found for Fe, Co, and Ge in addition to Ni. For these elements, the use of compound standards instead of pure elements usually resulted in significantly improved analytical accuracy. However, compound standards do not always provide satisfactory accuracy as is demonstrated for L-shell peak analysis in the Fe­S system: FeS and FeS2 unexpectedly do not provide good accuracy when used as mutual standards.

Aluminium-silicon interactions in higher plants: an update.

Aluminium (Al) and silicon (Si) are abundant in soils, but their availability for plant uptake is limited by low solubility. However, Al toxicity is a major problem in naturally occurring acid soils and in soils affected by acidic precipitation. When, in 1995, we reviewed this topic for the Journal of Experimental Botany, it was clear that under certain circumstances soluble Si could ameliorate the toxic effects of Al, an effect mirrored in organisms beyond the plant kingdom. In the 25 years since our review, it has become evident that the amelioration phenomenon occurs in the root apoplast, with the formation of hydroxyaluminosilicates being part of the mechanism. A much better knowledge of the molecular basis for Si and Al uptake by plants and of Al toxicity mechanisms has been developed. However, relating this work to amelioration by Si is at an early stage. It is now clear that co-deposition of Al and Si in phytoliths is a fairly common phenomenon in the plant kingdom, and this may be important in detoxification of Al. Relatively little work on Al-Si interactions in field situations has been done in the last 25 years, and this is a key area for future development.

Manganese hyperaccumulation capacity of Ilex paraguariensis A. St. Hil. and occurrence of interveinal chlorosis induced by transient toxicity.

Manganese (Mn) toxicity is common in plants grown on very acid soils. However, some plants species that grow in this condition can take up high amounts of Mn and are referred to as hyperaccumulating species. In this study, we evaluated the capacity of Ilex paraguariensis to accumulate Mn and the effect of excessive concentrations on plant growth and nutrition. For this, a container experiment was conducted using soils from different parent materials (basalt and sandstone), with and without liming, and at six doses of applied Mn (0, 30, 90, 270, 540 and 1,080 mg kg-1). Clonal plants grown for 203 days were harvested to evaluate yield, and leaf tissue samples were evaluated for Mn and other elements. Without liming and with high Mn doses, leaf Mn concentrations reached 13,452 and 12,127 mg kg-1 in sandstone and basalt soils, respectively; concentrations in excess of 10,000 mg kg-1 are characteristic of hyperaccumulating plants. Liming reduced these values to 7203 and 8030 mg kg-1. More plant growth accompanied increased Mn leaf concentrations, with a growth reduction noted at the highest dose in unlimed soils. Elemental distribution showed Mn presence in the mesophyll, primarily in vascular bundles, without high Mn precipitates. Interveinal chlorosis of young leaves associated with high Mn concentration and lower Fe concentrations was observed, especially in sandstone soil without liming. However, the occurrence of this symptom was not associated with decreased plant growth.

Grafting alleviates potassium stress and improves growth in tobacco.

BACKGROUND: Potassium is a nutrient element necessary for tobacco growth. Tobacco leaves with high potassium content are elastic and tough, rich in oil. And the same time, potassium can also improve the scent and aromatic value of flue-cured tobacco by regulating the synthesis of aromatic hydrocarbons in leaves.. It is an important quality indicator for flue-cured tobacco. However, the potassium concentration in tobacco leaves in most areas of China is generally lower than the global standard for high quality tobacco. Two tobacco genotypes were grafted to each other under different potassium levels to test whether potassium content and plant growth can be improved by grafting in tobacco. RESULTS: The growth of tobacco in all treatments was inhibited under potassium starvation, and grafting significantly alleviated this potassium stress in 'Yunyan 87'. The trends in whole plant K+ uptake and K+ transfer efficiency to the leaves corresponded to the growth results of the different grafts. The nutrient depletion test results showed that the roots of 'Wufeng No.2' had higher K+ absorption potential, K+ affinity, and K+ inward flow rate. K+ enrichment circles appeared at the endoderm of the root section in the energy dispersive X-ray figure, indicating that the formation of Casparian strips may be partly responsible for the lower rate of lateral movement of K+ in the roots of 'Yunyan 87'. Gene expression analysis suggested that energy redistribution at the whole plant level might constitute one strategy for coping with potassium starvation. The feedback regulation effects between scion 'Wufeng No.2' and rootstock 'Yunyan 87' indicated that the transmission of certain signaling substances had occurred during grafting. CONCLUSIONS: 'Wufeng No.2' tobacco rootstock grafting can increase the K+ uptake and transport efficiency of 'Yunyan 87' and enhance plant growth under potassium stress. The physiological mechanism of the improved performance of grafted tobacco is related to higher K+ uptake and utilization ability, improved xylem K+ loading capacity, and up-regulated expression of genes related to energy supply systems.

Globular structures in roots accumulate phosphorus to extremely high concentrations following phosphorus addition.

Crops with improved uptake of fertilizer phosphorus (P) would reduce P losses and confer environmental benefits. We examined how P-sufficient 6-week-old soil-grown Trifolium subterraneum plants, and 2-week-old seedlings in solution culture, accumulated P in roots after inorganic P (Pi) addition. In contrast to our expectation that vacuoles would accumulate excess P, after 7 days, X-ray microanalysis showed that vacuolar [P] remained low (<12 mmol kg-1 ). However, in the plants after P addition, some cortex cells contained globular structures extraordinarily rich in P (often >3,000 mmol kg-1 ), potassium, magnesium, and sodium. Similar structures were evident in seedlings, both before and after P addition, with their [P] increasing threefold after P addition. Nuclear magnetic resonance (NMR) spectroscopy showed seedling roots accumulated Pi following P addition, and transmission electron microscopy (TEM) revealed large plastids. For seedlings, we demonstrated that roots differentially expressed genes after P addition using RNAseq mapped to the T. subterraneum reference genome assembly and transcriptome profiles. Among the most up-regulated genes after 4 hr was TSub_g9430.t1, which is similar to plastid envelope Pi transporters (PHT4;1, PHT4;4): expression of vacuolar Pi-transporter homologs did not change. We suggest that subcellular P accumulation in globular structures, which may include plastids, aids cytosolic Pi homeostasis under high-P availability.

Salinity tolerance in chickpea is associated with the ability to 'exclude' Na from leaf mesophyll cells.

Salinity tolerance is associated with Na 'exclusion' from, or 'tissue tolerance' in, leaves. We investigated whether two contrasting chickpea genotypes, salt-tolerant Genesis836 and salt-sensitive Rupali, differ in leaf tissue tolerance to NaCl. We used X-ray microanalysis to evaluate cellular Na, Cl, and K concentrations in various cell types within leaflets and also in secretory trichomes of the two chickpea genotypes in relation to photosynthesis in control and saline conditions. TEM was used to assess the effects of salinity on the ultrastructure of chloroplasts. Genesis836 maintained net photosynthetic rates (A) for the 21 d of salinity treatment (60 mM NaCl), whereas A in Rupali substantially decreased after 11 d. Leaflet tissue [Na] was low in Genesis836 but had increased markedly in Rupali. In Genesis836, Na was accumulated in epidermal cells but was low in mesophyll cells, whereas in Rupali cellular [Na] was high in both cell types. The excessive accumulation of Na in mesophyll cells of Rupali corresponded to structural damage to the chloroplasts. Maintenance of photosynthesis and thus salinity tolerance in Genesis836 was associated with an ability to 'exclude' Na from leaflets and in particular from the photosynthetically active mesophyll cells, and to compartmentalize Na in epidermal cells.

Calcium modulates leaf cell-specific phosphorus allocation in Proteaceae from south-western Australia.

Over 650 Proteaceae occur in south-western Australia, contributing to the region's exceptionally high biodiversity. Most Proteaceae occur exclusively on severely nutrient-impoverished, acidic soils (calcifuge), whilst only few also occur on young, calcareous soils (soil-indifferent), higher in calcium (Ca) and phosphorus (P). The calcifuge habit of Proteaceae is explained by Ca-enhanced P toxicity, putatively linked to the leaf cell-specific allocation of Ca and P. Separation of these elements is essential to avoid the deleterious precipitation of Ca-phosphate. We used quantitative X-ray microanalysis to determine leaf cell-specific nutrient concentrations of two calcifuge and two soil-indifferent Proteaceae grown in hydroponics at a range of Ca and P concentrations. Calcium enhanced the preferential allocation of P to palisade mesophyll (PM) cells under high P conditions, without a significant change in whole leaf [P]. Calcifuges showed a greater PM [P] compared with soil-indifferent species, corresponding to their greater sensitivity. This study advances our mechanistic understanding of Ca-enhanced P toxicity, supporting the proposed model, and demonstrating its role in the calcifuge distribution of Proteaceae. This furthers our understanding of nutrient interactions at the cellular level and highlights its importance to plant functioning.