Analysis of Ionomic Profiles of Spinal Cords in a Rat Model with Bone Cancer Pain.

Background: Ionomics is used to study levels of ionome in different states of organisms and their correlations. Bone cancer pain (BCP) severely reduces quality of life of patients or their lifespan. However, the relationship between BCP and ionome remains unclear. Methods: The BCP rat model was constructed through inoculation of Walker 256 cells into the left tibia. Von Frey test, whole-cell patch-clamp recording and inductively coupled plasma mass spectrometry (ICP-MS) technologies were conducted for measuring tactile hypersensitivity, the frequency and amplitude of miniature excitatory postsynaptic currents (mEPSCs) of neurons of spinal slices, and ionome of spinal cord samples, respectively. Principal component analysis (PCA) was used to explore ionomic patterns of the spinal cord. Results: The BCP rat model was successfully constructed through implantation of Walker 256 cells into the left tibia. The frequency and amplitude of mEPSCs of neurons in the spinal cord slices from the BCP model rats were notably greater than those in the sham control. In terms of ionomics, the spinal cord levels of two macroelements (Ca and S), four microelements (Fe, Mn, Li and Sr) and the toxic element Ti in the BCP group of rats were significantly increased by inoculation of Walker 256 cancer cells, compared to the sham control. In addition, the correlation patterns between the elements were greatly changed between the sham control and BCP groups. PCA showed that inoculation of Walker 256 cells into the tibia altered the overall ionomic profiles of the spinal cord. There was a significant separation trend between the two groups. Conclusion: Taken together, inoculation of Walker 256 cells into the left tibia contributes to BCP, which could be closely correlated by some elements. The findings provided novel information on the relationship between the ionome and BCP.

Adsorption of Zn(II) from aqueous solution and separation of zinc isotopes by displacement chromatography using chelating adsorbent.

The removal of zinc ions (Zn(II)) in water and the separation of zinc isotopes were fully investigated in this study. Imidodiacetic acid (IDA) type adsorbent (named PSGI) based on polystyrene spheres (PS) was synthesized by simultaneous irradiation grafting. By adsorption method, the removal of Zn(II) from water by the chelating adsorbent was studied in batch experiments. Under optimized condition, PSGI showed the removal efficiency of more than 98 % for Zn(II) and the adsorption capacity of 70.1 mg/g. Langmuir isothermal and pseudo-second-order kinetic model fitted the experimental results better, indicating that the adsorption is dominated by chemical adsorption. The spent adsorbent (PSGI-Zn) was used for further zinc isotope separation by displacement chromatography using EDTA-NH4 solution as eluent. Due to the mass effect of isotopes, 70Zn was found to preferentially fractionated into the front-end effluents with the highest front enrichment values of 70Zn/64Zn. By extending the migration distance to 20 m, we obtained the best isotope enrichment with the front maximum enrichment values as 1.0949, 1.0739 and separation coefficient values as 1.977 × 10-3, 8.33 × 10-3 corresponding to the isotope pairs 66Zn/64Zn, 68Zn/64Zn.

Interactions of molybdenum disulfide nanosheets with wheat plants under changing environments: More than meets the eye?

Molybdenum disulfide (MoS2) nanosheets are being increasingly employed in various applications. It is therefore imperative to assess their potential environmental implications in a changing world, particularly in the context of global warming. Here, we assessed the effects of MoS2 nanosheets on wheat Triticum aestivum L. under today's typical climatic conditions (22 °C) and future climatic conditions (30 °C), respectively. The results showed that MoS2 nanosheets (10 and 100 Mo mg/L) did not significantly affect wheat plant growth, root morphological traits, and chlorophyll fluorescence, regardless of dose and temperature. However, the metabolic processes were significantly altered in T. aestivum upon exposure to individual MoS2 nanosheets and to a combination of MoS2 nanosheets and future global warming. As a non-specific protective strategy, the wheat plants that were under stress conditions maintained the stability of cell membranes and thus relieved cell injury by accumulating more glycerophospholipids. Warming additionally influenced the nitrogen and carbon pool reallocation in wheat root. MoS2 nanosheets mainly depleted a range of antioxidant metabolites involved in phenylpropanoid biosynthesis and taurine and hypotaurine metabolism, while warming activated vitamin B6 cofactors related to vitamin B6 metabolism. Metabolites involved in glutathione metabolism were uniquely upregulated while most metabolites associated with nucleotide metabolisms were uniquely downregulated in combination-treated wheat. Overall, wheat plants regulated a wide range of growth-related processes, including carbohydrate, amino acids, lipid, vitamins, and nucleotide metabolism, to maintain optimal metabolite pool sizes and eventually global metabolic homeostasis upon different stress conditions. Our findings provide novel insights into MoS2 nanosheets-mediated crop responses under global warming.

The ABC transporter ABCG36 is required for cadmium tolerance in rice.

Cadmium (Cd) is a highly toxic heavy metal in nature, which causes severe damage to plant growth. The molecular mechanisms for Cd detoxification are poorly understood. Here, we report that a G-type ATP-binding cassette transporter, OsABCG36, is involved in Cd tolerance in rice. OsABCG36 was expressed in both roots and shoots at a low level, but expression in the roots rather than the shoots was greatly up-regulated by a short exposure to Cd. A spatial expression analysis showed that Cd-induced expression of OsABCG36 was found in both the root tip and the mature root region. Transient expression of OsABCG36 in rice protoplast cells showed that it was localized to the plasma membrane. Immunostaining showed that OsABCG36 was localized in all root cells except the epidermal cells. Knockout of OsABCG36 resulted in increased Cd accumulation in root cell sap and enhanced Cd sensitivity, but did not affect tolerance to other metals including Al, Zn, Cu, and Pb. The concentration of Cd in the shoots was similar between the knockout lines and wild-type rice. Heterologous expression of OsABCG36 in yeast showed an efflux activity for Cd, but not for Zn. Taken together, our results indicate that OsABCG36 is not involved in Cd accumulation in the shoots, but is required for Cd tolerance by exporting Cd or Cd conjugates from the root cells in rice.

TSC1 enables plastid development under dark conditions, contributing to rice adaptation to transplantation shock.

Since its domestication from wild rice thousands of years ago, rice has been cultivated largely through transplantation. During transplantation from the nursery to the paddy field, rice seedlings experience transplantation shock which affects their physiology and production. However, the mechanisms underlying transplantation shock and rice adaptation to this shock are largely unknown. Here, we isolated a transplant-sensitive chloroplast-deficient (tsc1) rice mutant that produces albino leaves after transplantation. Blocking light from reaching the juvenile leaves and leaf primordia caused chloroplast deficiencies in transplanted tsc1 seedlings. TSC1 encodes a noncanonical adenosine triphosphate-binding cassette (ABC) transporter homologous to AtNAP14 and is of cyanobacterial origin. We demonstrate that TSC1 controls plastid development in rice under dark conditions, and functions independently of light signaling. However, light rescued the tsc1 mutant phenotype in a spectrum-independent manner. TSC1 was upregulated following transplantation, and modulated the iron and copper levels, thereby regulating prolamellar body formation during the early P4 stage of leaf development. Therefore, TSC1 is indispensable for plastid development in the absence of light, and contributes to adaptation to transplantation shock. Our study provides insight into the regulation of plastid development and establishes a framework for improving recovery from transplantation shock in rice.

A new vesicle trafficking regulator CTL1 plays a crucial role in ion homeostasis.

Ion homeostasis is essential for plant growth and environmental adaptation, and maintaining ion homeostasis requires the precise regulation of various ion transporters, as well as correct root patterning. However, the mechanisms underlying these processes remain largely elusive. Here, we reported that a choline transporter gene, CTL1, controls ionome homeostasis by regulating the secretory trafficking of proteins required for plasmodesmata (PD) development, as well as the transport of some ion transporters. Map-based cloning studies revealed that CTL1 mutations alter the ion profile of Arabidopsis thaliana. We found that the phenotypes associated with these mutations are caused by a combination of PD defects and ion transporter misregulation. We also established that CTL1 is involved in regulating vesicle trafficking and is thus required for the trafficking of proteins essential for ion transport and PD development. Characterizing choline transporter-like 1 (CTL1) as a new regulator of protein sorting may enable researchers to understand not only ion homeostasis in plants but also vesicle trafficking in general.

AtHKT1 drives adaptation of Arabidopsis thaliana to salinity by reducing floral sodium content.

Arabidopsis thaliana high-affinity potassium transporter 1 (AtHKT1) limits the root-to-shoot sodium transportation and is believed to be essential for salt tolerance in A. thaliana. Nevertheless, natural accessions with 'weak allele' of AtHKT1, e.g. Tsu-1, are mainly distributed in saline areas and are more tolerant to salinity. These findings challenge the role of AtHKT1 in salt tolerance and call into question the involvement of AtHKT1 in salinity adaptation in A. thaliana. Here, we report that AtHKT1 indeed drives natural variation in the salt tolerance of A. thaliana and the coastal AtHKT1, so-called weak allele, is actually hyper-functional in reducing flowers sodium content upon salt stress. Our data showed that AtHKT1 positively contributes to saline adaptation in a linear manner. Forward and reverse genetics analysis established that the single AtHKT1 locus is responsible for the variation in the salinity adaptation between Col-0 and Tsu-1. Reciprocal grafting experiments revealed that shoot AtHKT1 determines the salt tolerance of Tsu-1, whereas root AtHKT1 primarily drives the salt tolerance of Col-0. Furthermore, evidence indicated that Tsu-1 AtHKT1 is highly expressed in stems and is more effective compared to Col-0 AtHKT1 at limiting sodium flow to the flowers. Such efficient retrieval of sodium to the reproductive organ endows Tsu-1 with stronger fertility compared to Col-0 upon salt stress, thus improving Tsu-1 adaptation to a coastal environment. To conclude, our data not only confirm the role of AtHKT1 in saline adaptation, but also sheds light on our understanding of the salt tolerance mechanisms in plants.

Structure and mechanism of a group-I cobalt energy coupling factor transporter.

Energy-coupling factor (ECF) transporters are a large family of ATP-binding cassette transporters recently identified in microorganisms. Responsible for micronutrient uptake from the environment, ECF transporters are modular transporters composed of a membrane substrate-binding component EcfS and an ECF module consisting of an integral membrane scaffold component EcfT and two cytoplasmic ATP binding/hydrolysis components EcfA/A'. ECF transporters are classified into groups I and II. Currently, the molecular understanding of group-I ECF transporters is very limited, partly due to a lack of transporter complex structural information. Here, we present structures and structure-based analyses of the group-I cobalt ECF transporter CbiMNQO, whose constituting subunits CbiM/CbiN, CbiQ, and CbiO correspond to the EcfS, EcfT, and EcfA components of group-II ECF transporters, respectively. Through reconstitution of different CbiMNQO subunits and determination of related ATPase and transporter activities, the substrate-binding subunit CbiM was found to stimulate CbiQO's basal ATPase activity. The structure of CbiMQO complex was determined in its inward-open conformation and that of CbiO in ß, γ-methyleneadenosine 5'-triphosphate-bound closed conformation. Structure-based analyses revealed interactions between different components, substrate-gating function of the L1 loop of CbiM, and conformational changes of CbiO induced by ATP binding and product release within the CbiMNQO transporter complex. These findings enabled us to propose a working model of the CbiMNQO transporter, in which the transport process requires the rotation or toppling of both CbiQ and CbiM, and CbiN might function in coupling conformational changes between CbiQ and CbiM.

Arabidopsis AMINO ACID PERMEASE1 Contributes to Salt Stress-Induced Proline Uptake from Exogenous Sources.

Stress-induced proline accumulation in plants is thought to result primarily from enhanced proline biosynthesis and decreased proline degradation. To identify regulatory components involved in proline transport, we screened for Arabidopsis thaliana T-DNA mutants with enhanced tolerance to toxic levels of exogenous proline (45 mM). We isolated the proline resistant 1-1 (pre1-1) mutant and map-based cloning identified PRE1 as AMINO ACID PERMEASE1 (AAP1, At1g58360), which encodes a plasma membrane-localized amino acid permease. AAP1 expression is induced by salt stress and abscisic acid, but not by proline. In pre1-1 mutants, a 19-nucleotide deletion in the AAP1 coding region produced a premature stop codon. When grown on proline-containing medium, pre1-1 mutants accumulated significantly less proline than did the wild type. Under salt stress, proline uptake decreased significantly in pre1-1 mutants. By contrast, proline uptake increased significantly in the wild type. These results suggest that AAP1 functions in the increase of proline uptake during salt stress. In addition, proline uptake promotes salt tolerance in Arabidopsis seedlings. We conclude that plants can increase proline accumulation by AtAAP1-mediated proline uptake from exogenous source, which help to improve the salt tolerance of seedlings.

Genome-wide association mapping identifies a new arsenate reductase enzyme critical for limiting arsenic accumulation in plants.

Inorganic arsenic is a carcinogen, and its ingestion through foods such as rice presents a significant risk to human health. Plants chemically reduce arsenate to arsenite. Using genome-wide association (GWA) mapping of loci controlling natural variation in arsenic accumulation in Arabidopsis thaliana allowed us to identify the arsenate reductase required for this reduction, which we named High Arsenic Content 1 (HAC1). Complementation verified the identity of HAC1, and expression in Escherichia coli lacking a functional arsenate reductase confirmed the arsenate reductase activity of HAC1. The HAC1 protein accumulates in the epidermis, the outer cell layer of the root, and also in the pericycle cells surrounding the central vascular tissue. Plants lacking HAC1 lose their ability to efflux arsenite from roots, leading to both increased transport of arsenic into the central vascular tissue and on into the shoot. HAC1 therefore functions to reduce arsenate to arsenite in the outer cell layer of the root, facilitating efflux of arsenic as arsenite back into the soil to limit both its accumulation in the root and transport to the shoot. Arsenate reduction by HAC1 in the pericycle may play a role in limiting arsenic loading into the xylem. Loss of HAC1-encoded arsenic reduction leads to a significant increase in arsenic accumulation in shoots, causing an increased sensitivity to arsenate toxicity. We also confirmed the previous observation that the ACR2 arsenate reductase in A. thaliana plays no detectable role in arsenic metabolism. Furthermore, ACR2 does not interact epistatically with HAC1, since arsenic metabolism in the acr2 hac1 double mutant is disrupted in an identical manner to that described for the hac1 single mutant. Our identification of HAC1 and its associated natural variation provides an important new resource for the development of low arsenic-containing food such as rice.

An update on magnesium homeostasis mechanisms in plants.

Worldwide, nearly two-thirds of the population do not consume the recommended amount of magnesium (Mg) in their diet. Furthermore, low Mg status (hypomagnesaemia) is known to contribute to a number of human chronic disease conditions. Because the principal dietary Mg source is of plant origin, agronomic and genetic biofortification strategies are aimed at improving nutritional Mg content in food crops to overcome this mineral deficiency in humans. This update incorporates the contributions of annotated permeases involved in Mg uptake, storage and recycling with a schematic model of Mg movement at the organ and cellular levels in the model species Arabidopsis thaliana. Furthermore, approaches using mutagenesis or natural ionomic variation to identify loci involved in Mg homeostasis in roots, leaves and seeds will be summarised. A brief overview will be presented on how Arabidopsis research can help to develop strategies for biofortification of crops.

Variation in HMA4 gene copy number and expression among Noccaea caerulescens populations presenting different levels of Cd tolerance and accumulation.

There is huge variability among populations of the hyperaccumulator Noccaea caerulescens (formerly Thlaspi caerulescens) in their capacity to tolerate and accumulate cadmium. To gain new insights into the mechanisms underlying this variability, we estimated cadmium fluxes and further characterized the N. caerulescens heavy metal ATPase 4 (NcHMA4) gene in three populations (two calamine, Saint-Félix-de-Pallières, France and Prayon, Belgium; one serpentine, Puente Basadre, Spain) presenting contrasting levels of tolerance and accumulation. Cadmium uptake and translocation varied among populations in the same way as accumulation; the population with the highest cadmium concentration in shoots (Saint Félix-de-Pallières) presented the highest capacity for uptake and translocation. We demonstrated that the four NcHMA4 copies identified in a previous study are not fixed at the species level, and that the copy truncated in the C-terminal part encodes a functional protein. NcHMA4 expression and gene copy number was lower in the serpentine population, which was the least efficient in cadmium translocation compared to the calamine populations. NcHMA4 expression was associated with the vascular tissue in all organs, with a maximum at the crown. Overall, our results indicate that differences in cadmium translocation ability of the studied populations appear to be controlled, at least partially, by NcHMA4, while the overexpression of NcHMA4 in the two calamine populations may result from convergent evolution.

Low magnesium status in plants enhances tolerance to cadmium exposure.

In a transcriptomic study of magnesium (Mg) starvation in Arabidopsis, we identified several genes that were differentially regulated which are involved in the detoxification process of nonessential heavy metals such as cadmium (Cd). We further tested the impact of low Mg status on Cd sensitivity in plants. Interestingly, a -Mg pretreatment of 7 d alleviated the bleaching of young leaves caused by Cd. No or little difference in Cd tissue concentration between the +Mg and -Mg plants was observed, suggesting that lower Cd toxicity was probably not attributable to modified root to shoot translocation. Mg deficiency also promoted an increase in the iron (Fe) concentration (up to one-fourth) in Cd-treated leaves. Because high Fe concentrations have previously been reported to prevent the harmful effects of Cd, we explored whether Fe homeostasis plays a role in the Mg-Cd interaction. A protective effect of -Mg pretreatment was also observed on Fe starvation. However, Fe foliar spray partially alleviated Cd-induced chloroses, while it almost completely restored chlorophyll content in Fe-deficient leaves. In conclusion, the protective effect of Mg against Cd toxicity could be attributable partly to the maintenance of Fe status but also to the increase in antioxidative capacity, detoxification and/or protection of the photosynthetic apparatus.

Proline induces calcium-mediated oxidative burst and salicylic acid signaling.

Although free proline accumulation is a well-documented phenomenon in many plants in response to a variety of environmental stresses, and is proposed to play protective roles, high intracellular proline content, by either exogenous application or endogenous over-production, in the absence of stresses, is found to be inhibitory to plant growth. We have shown here that exogenous application of proline significantly induced intracellular Ca(2+) accumulation in tobacco and calcium-dependent ROS production in Arabidopsis seedlings, which subsequently enhanced salicylic acid (SA) synthesis and PR genes expression. This suggested that proline can promote a reaction similar to hypersensitive response during pathogen infection. Other amino acids, such as glutamate, but not arginine and phenylalanine, were also found to be capable of inducing PR gene expression. In addition, proline at concentration as low as 0.5 mM could induce PR gene expression. However, proline could not induce the expression of PDF1.2 gene, the marker gene for jasmonic acid signaling pathway. Furthermore, proline-induced SA production is mediated by NDR1-dependent signaling pathway, but not that mediated by PAD4. Our data provide evidences that exogenous proline, and probably some other amino acids can specifically induce SA signaling and defense response.