Salicylic Acid's impact on Sedum alfredii growth and cadmium tolerance: Comparative physiological, transcriptomic, and metabolomic study.

With the acceleration of industrialization, Cd pollution has emerged as a major threat to soil ecosystem health and food safety. Hyperaccumulating plants like Sedum alfredii Hance are considered to be used as part of an effective strategy for the ecological remediation of Cd polluted soils. This study delved deeply into the physiological, transcriptomic, and metabolomic responses of S. alfredii under cadmium (Cd) stress when treated with exogenous salicylic acid (SA). We found that SA notably enhanced the growth of S. alfredii and thereby increased absorption and accumulation of Cd, effectively alleviating the oxidative stress caused by Cd through upregulation of the antioxidant system. Transcriptomic and metabolomic data further unveiled the influence of SA on photosynthesis, antioxidant defensive mechanisms, and metal absorption enrichment pathways. Notably, the interactions between SA and other plant hormones, especially IAA and JA, played a central role in these processes. These findings offer us a comprehensive perspective on understanding how to enhance the growth and heavy metal absorption capabilities of hyperaccumulator plants by regulating plant hormones, providing invaluable strategies for future environmental remediation efforts.

Appropriate sulfur fertilization in contaminated soil enhanced the cadmium uptake by hyperaccumulator Sedum alfredii Hance.

The biogeochemical processes of sulfur and heavy metals in the environment are closely related to each other. We investigated the influence of sulfur addition on hyperaccumulator Sedum alfredii Hance growth, cadmium (Cd) accumulation, soil Cd bioavailability, soil bacterial communities and plant transcriptome responses. The results showed that an appropriate rate of sulfur addition (1.0 or 2.5 g/kg) enhanced the growth of Sedum alfredii Hance plants as well as their accumulation of Cd. A high rate of sulfur addition (5.0 or 10.0 g/kg) causes toxicity to Sedum alfredii Hance plants. The application of an appropriate amount of sulfur to the soil increased the abundance of sulfur-oxidizing bacteria such as Sulfuriferula and Thiobacillus; acid-fast bacillus such as Alicyclobacillus; and cadmium-tolerant bacteria such as Bacillus and Rhodanobacter. This led to a decrease in pH and an increase in bioavailable Cd in the soil. RNA sequencing revealed that the addition of sulfur to soils led to the up regulation of most of the differentially expressed genes (DEGs) involved in "photosynthesis" and "photosynthesis, light reaction" in Sedum alfredii Hance leaves. Moreover, the "plant hormone signal transduction" pathway was significantly enriched with sulfur addition. Sulfur assimilation in Sedum alfredii Hance plants may promote photosynthesis and hormone synthesis, leading to Cd tolerance in these plants. Our study revealed that sulfur fertilization enhanced the efficiency of Cd phytoremediation in Sedum alfredii Hance plants.

Rapid assessment of heavy metal accumulation capability of Sedum alfredii using hyperspectral imaging and deep learning.

Hyperaccumulators are the material basis and key to the phytoremediation of heavy metal contaminated soils. Conventional methods for screening hyperaccumulators are highly dependent on the time- and labor-consuming sampling and chemical analysis. In this study, a novel spectral approach assisted with multi-task deep learning was proposed to streamline accumulating ecotype screening, heavy metal stress discrimination, and heavy metals quantification in plants. The significant Cd/Zn co-hyperaccumulator Sedum alfredii and its non-accumulating ecotype were stressed by Cd, Zn, and Pb. Spectral images of leaves were rapidly acquired by hyperspectral imaging. The self-designed deep learning architecture was composed of a shallow network (ENet) for accumulating ecotype identification, and a multi-task network (HMNet) for heavy metal stress type and accumulation prediction simultaneously. To further assess the robustness of the networks, they were compared with conventional machine learning models (i.e., partial least squares (PLS) and support vector machine (SVM)) on a series of evaluation metrics of classification, multi-label classification, and regression. S. alfredii with heavy metals accumulation capability was identified by ENet with 100 % accuracy. HMNet reduced overfitting and outperformed machine learning models with the average exact match ratio (EMR) of heavy metal stress discrimination increased by 7.46 %, and residual prediction deviations (RPD) of heavy metal concentrations prediction increased by 53.59 %. The method succeeded in rapidly and accurately discriminating heavy metal stress with EMRs over 91 % and accuracies over 96 %, and in predicting heavy metals accumulation with an average RPD of 3.29 for Zn, 2.57 for Cd, and 2.53 for Pb, indicating the satisfactory practicability and potential for sensing heavy metals accumulation. This study provides a relatively novel spectral method to facilitate hyperaccumulator screening and heavy metals accumulation prediction in the phytoremediation process.

Effect of plant extracts and citric acid on phytoremediation of metal-contaminated soil.

Application of activating agents can significantly improve efficiency of plants for remediation of soils contaminated by heavy metals, however, damage to soil and plants limits application of traditional activating agents. The aim of our experiments is to select an efficient,green and low-cost activating agent to improve efficiency of plant extraction technology. In this study, contaminated soil was remediated by Sedum alfredii. The effects of two plant extracts (i.e., Oxalis corniculata,OX and Medicago sativaextract, ME) in addition to citric acid (CA) were studied in oscillatory activation experiment and pot experiment. The oscillation activation experiment revealed that extraction quantity of heavy metals in the soil was enhanced significantly with concentration of plant extract. The extraction quantity of Zn from 100% OX extract and ME extract were significantly higher than 10 mmol/L CA (54.04% and 33.09%, respectively). The 10 mmol/L CA has best extraction efficiency for Cd, up to 41.36 µg/kg, which is significantly higher than CK (control) (p < 0.05). The pot experiment exhibited that application of CA has significantly reduced soil pH and organic matter content by 8.63% and 28.21%, respectively, however the two extracts have no significant effect on soil properties. The study indicated that application of CA has negative effects on root morphological parameters and chlorophyll fluorescence parameters of Sedum alfredii.The addition of extracts of two plants have not caused any harm to Sedum alfredii. The application of three activating agents was beneficial for purification of Cd and Zn in soils, and its repairing efficiency was improved by 3.92, 3.37, 3.33 times and 0.44, 0.20, 0.86 times, respectively. The combination of plant extracts and hyperaccumulators can effectively remove heavy metals from contaminated soils, which provided a theoretical basis for mitigation of pollution in soils.

Auxin-Induced SaARF4 Downregulates SaACO4 to Inhibit Lateral Root Formation in Sedum alfredii Hance.

Lateral root (LR) formation promotes plant resistance, whereas high-level ethylene induced by abiotic stress will inhibit LR emergence. Considering that local auxin accumulation is a precondition for LR generation, auxin-induced genes inhibiting ethylene synthesis may thus be important for LR development. Here, we found that auxin response factor 4 (SaARF4) in Sedum alfredii Hance could be induced by auxin. The overexpression of SaARF4 decreased the LR number and reduced the vessel diameters. Meanwhile, the auxin distribution mode was altered in the root tips and PIN expression was also decreased in the overexpressed lines compared with the wild-type (WT) plants. The overexpression of SaARF4 could reduce ethylene synthesis, and thus, the repression of ethylene production decreased the LR number of WT and reduced PIN expression in the roots. Furthermore, the quantitative real-time PCR, chromatin immunoprecipitation sequencing, yeast one-hybrid, and dual-luciferase assay results showed that SaARF4 could bind the promoter of 1-aminocyclopropane-1-carboxylate oxidase 4 (SaACO4), associated with ethylene biosynthesis, and could downregulate its expression. Therefore, we concluded that SaARF4 induced by auxin can inhibit ethylene biosynthesis by repressing SaACO4 expression, and this process may affect auxin transport to delay LR development.

Analysis of the effect of cadmium stress on root exudates of Sedum plumbizincicola based on metabolomics.

Root exudates are the most direct manifestation of the response of plants changes in the external environment. Therefore, based on non-targeted gas chromatography-time-of-flight mass spectrometry and metabolomics, the response of Sedum plumbizincicola root exudates to Cd stress was used to reveal the possible mechanism of resistance to or accumulation of Cd. The results showed that Cd significantly changed the composition and contents of S. plumbizincicola root exudates. A total of 155 metabolites were identified in S. plumbizincicola root exudates, among which 33 showed significant differences under Cd stress, including organic acids, amino acids, lipids, and polyols. Cd stress suppressed organic acid metabolism and lipid metabolism in S. plumbizincicola and significantly affected amino acid metabolism. There were 16 metabolic pathways related to Cd stress, among which arginine and proline metabolism, valine, leucine, and isoleucine biosynthesis, glycine, serine, and threonine metabolism, glutathione metabolism, and purine metabolism were the key pathways with the highest correlation, and were closely related to the stress resistance of S. plumbizincicola.

Site-specific regulation of transcriptional responses to cadmium stress in the hyperaccumulator, Sedum alfredii: based on stem parenchymal and vascular cells.

KEY MESSAGE: We compared the transcriptomes of parenchymal and vascular cells of Sedum alfredii stem under Cd stress to reveal gene regulatory networks underlying Cd hyperaccumulation. Cadmium (Cd) hyperaccumulation in plants is a complex biological process controlled by gene regulatory networks. Efficient transport through vascular systems and storage by parenchymal cells are vital for Cd hyperaccumulation in the Cd hyperaccumulator Sedum alfredii, but the genes involved are poorly understood. We investigated the spatial gene expression profiles of transport and storage sites in S. alfredii stem using laser-capture microdissection coupled with RNA sequencing. Gene expression patterns in response to Cd were distinct in vascular and parenchymal cells, indicating functional divisions that corresponded to Cd transportation and storage, respectively. In vascular cells, plasma membrane-related terms enriched a large number of differentially-expressed genes (DEGs) for foundational roles in Cd transportation. Parenchymal cells contained considerable DEGs specifically concentrated on vacuole-related terms associated with Cd sequestration and detoxification. In both cell types, DEGs were classified into different metabolic pathways in a similar way, indicating the role of Cd in activating a systemic stress signalling network where ATP-binding cassette transporters and Ca2+ signal pathways were probably involved. This study identified site-specific regulation of transcriptional responses to Cd stress in S. alfredii and analysed a collection of genes that possibly function in Cd transportation and detoxification, thus providing systemic information and direction for further investigation of Cd hyperaccumulation molecular mechanisms.

Effects of CO2 application coupled with endophyte inoculation on rhizosphere characteristics and cadmium uptake by Sedum alfredii Hance in response to cadmium stress.

Comparative impact of CO2 application and endophyte inoculation was investigated on the growth, rhizosphere characteristics, and cadmium (Cd) absorption of two ecotypes of Sedum alfredii Hance in response to Cd stress under hydroponic or rhizo-box culture conditions. The results showed that both CO2 application and endophyte inoculation significantly (P < 0.05) promoted plant growth (fresh weight and dry weight), improved root morphological properties (SRL, SRA, SRV, ARD and RTN) and exudation (pH, TOC, TN, soluble sugar and organic acids), changed Cd uptake and distribution of both ecotypes of S. alfredii. Meanwhile soil total and DTPA extractable Cd in rhizo-box decreased by biofortification treatments. Superposition biofortification exhibits utmost improvement for the above mentioned parameters, and has potential for enhancing phytoremediation efficiency of hyperaccumulator and sustaining regular growth of non-hyperaccumulator in Cd contaminated soils.

Heavy metal ATPase 3 (HMA3) confers cadmium hypertolerance on the cadmium/zinc hyperaccumulator Sedum plumbizincicola.

Cadmium (Cd) is highly toxic to most organisms, but some rare plant species can hyperaccumulate Cd in aboveground tissues without suffering from toxicity. The mechanism underlying Cd detoxification by hyperaccumulators is interesting but unclear. Here, the heavy metal ATPase 3 (SpHMA3) gene responsible for Cd detoxification was isolated from the Cd/zinc (Zn) hyperaccumulator Sedum plumbizincicola. RNA interference (RNAi)-mediated silencing and overexpression of SpHMA3 were induced to investigate its physiological functions in S. plumbizincicola and a nonhyperaccumulating ecotype of Sedum alfredii. Heterologous expression of SpHMA3 in Saccharomyces cerevisiae showed Cd-specific transport activity. SpHMA3 was highly expressed in the shoots and the protein was localized to the tonoplast. The SpHMA3-RNAi lines were hypersensitive to Cd but not to Zn, with the growth of shoots and young leaves being severely inhibited by Cd. Overexpressing SpHMA3 in the nonhyperaccumulating ecotype of S. alfredii greatly increased its tolerance to and accumulation of Cd, but not Zn. These results indicate that elevated expression of the tonoplast-localized SpHMA3 in the shoots plays an essential role in Cd detoxification, which contributes to the maintenance of the normal growth of young leaves of S. plumbizincicola in Cd-contaminated soils.

Enhanced expression of SaHMA3 plays critical roles in Cd hyperaccumulation and hypertolerance in Cd hyperaccumulator Sedum alfredii Hance.

MAIN CONCLUSION: The enhanced expression of a P 1B -type ATPase gene ( SaHMA3 ) is essential for Cd hyperaccumulation and hypertolerance in Sedum alfredii Hance. A functional understanding of the mechanism through which hyperaccumulator plants accumulate and tolerate extremely toxic metals is a prerequisite for the development of novel strategies for improving phytoremediation using engineered plants or natural hyperaccumulators as well as biofortification and food crop safety. Most hyperaccumulator species, however, are small and slow-growing, and their potential for large-scale decontamination of polluted soils is limited. Sedum alfredii Hance, the only one metal hyperaccumulator from the Crassulaceae family, is an ideal candidate for gaining a functional understanding of the intra-family hyperaccumulation mechanisms as well as their potential applications. In the present study, we isolated and functionally characterized a P1B-type ATPase gene (SaHMA3) from S. alfredii Hance. SaHMA3 alleles from a hyperaccumulating ecotype (HE) and non-hyperaccumulating ecotype (NHE) were constitutively expressed in both shoot and root and encoded tonoplast-localized proteins, but showed differences in transport substrate specificity and expression level. SaHMA3 h from the HE plant was a Cd transporter. In contrast, SaHMA3n from NHE plants was able to transport both Zn and Cd. SaHMA3 showed a significantly higher constitutive expression level in HE plants than in NHE plants. Furthermore, the expression level of SaHMA3 in the shoots of HE plants was considerably higher than in the roots. Overexpression of SaHMA3h in tobacco plants significantly enhanced Cd tolerance and accumulation and greatly increased the root sequestration of Cd. In summary, our data suggested that SaHMA3 plays critical roles in Cd hyperaccumulation and hypertolerance in Cd hyperaccumulator S. alfredii Hance.

Integration of small RNAs, degradome and transcriptome sequencing in hyperaccumulator Sedum alfredii uncovers a complex regulatory network and provides insights into cadmium phytoremediation.

The hyperaccumulating ecotype of Sedum alfredii Hance is a cadmium (Cd)/zinc/lead co-hyperaccumulating species of Crassulaceae. It is a promising phytoremediation candidate accumulating substantial heavy metal ions without obvious signs of poisoning. However, few studies have focused on the regulatory roles of miRNAs and their targets in the hyperaccumulating ecotype of S. alfredii. Here, we combined analyses of the transcriptomics, sRNAs and the degradome to generate a comprehensive resource focused on identifying key regulatory miRNA-target circuits under Cd stress. A total of 87 721 unigenes and 356 miRNAs were identified by deep sequencing, and 79 miRNAs were differentially expressed under Cd stress. Furthermore, 754 target genes of 194 miRNAs were validated by degradome sequencing. A gene ontology (GO) enrichment analysis of differential miRNA targets revealed that auxin, redox-related secondary metabolism and metal transport pathways responded to Cd stress. An integrated analysis uncovered 39 pairs of miRNA targets that displayed negatively correlated expression profiles. Ten miRNA-target pairs also exhibited negative correlations according to a real-time quantitative PCR analysis. Moreover, a coexpression regulatory network was constructed based on profiles of differentially expressed genes. Two hub genes, ARF4 (auxin response factor 4) and AAP3 (amino acid permease 3), which might play central roles in the regulation of Cd-responsive genes, were uncovered. These results suggest that comprehensive analyses of the transcriptomics, sRNAs and the degradome provided a useful platform for investigating Cd hyperaccumulation in S. alfredii, and may provide new insights into the genetic engineering of phytoremediation.

The hyperaccumulator Sedum plumbizincicola harbors metal-resistant endophytic bacteria that improve its phytoextraction capacity in multi-metal contaminated soil.

Endophyte-assisted phytoremediation has recently been suggested as a successful approach for ecological restoration of metal contaminated soils, however little information is available on the influence of endophytic bacteria on the phytoextraction capacity of metal hyperaccumulating plants in multi-metal polluted soils. The aims of our study were to isolate and characterize metal-resistant and 1-aminocyclopropane-1-carboxylate (ACC) utilizing endophytic bacteria from tissues of the newly discovered Zn/Cd hyperaccumulator Sedum plumbizincicola and to examine if these endophytic bacterial strains could improve the efficiency of phytoextraction of multi-metal contaminated soils. Among a collection of 42 metal resistant bacterial strains isolated from the tissues of S. plumbizincicola grown on Pb/Zn mine tailings, five plant growth promoting endophytic bacterial strains (PGPE) were selected due to their ability to promote plant growth and to utilize ACC as the sole nitrogen source. The five isolates were identified as Bacillus pumilus E2S2, Bacillus sp. E1S2, Bacillus sp. E4S1, Achromobacter sp. E4L5 and Stenotrophomonas sp. E1L and subsequent testing revealed that they all exhibited traits associated with plant growth promotion, such as production of indole-3-acetic acid and siderophores and solubilization of phosphorus. These five strains showed high resistance to heavy metals (Cd, Zn and Pb) and various antibiotics. Further, inoculation of these ACC utilizing strains significantly increased the concentrations of water extractable Cd and Zn in soil. Moreover, a pot experiment was conducted to elucidate the effects of inoculating metal-resistant ACC utilizing strains on the growth of S. plumbizincicola and its uptake of Cd, Zn and Pb in multi-metal contaminated soils. Out of the five strains, B. pumilus E2S2 significantly increased root (146%) and shoot (17%) length, fresh (37%) and dry biomass (32%) of S. plumbizincicola as well as plant Cd uptake (43%), whereas Bacillus sp. E1S2 significantly enhanced the accumulation of Zn (18%) in plants compared with non-inoculated controls. The inoculated strains also showed high levels of colonization in rhizosphere and plant tissues. Results demonstrate the potential to improve phytoextraction of soils contaminated with multiple heavy metals by inoculating metal hyperaccumulating plants with their own selected functional endophytic bacterial strains.

Effect of elevated CO2 concentration on photosynthetic characteristics of hyperaccumulator Sedum alfredii under cadmium stress.

The combined effects of elevated CO2 and cadmium (Cd) on photosynthetic rate, chlorophyll fluorescence and Cd accumulation in hyperaccumulator Sedum alfredii Hance were investigated to predict plant growth under Cd stress with rising atmospheric CO2 concentration. Both pot and hydroponic experiments were conducted and the plants were grown under ambient (350 µL L(-1)) or elevated (800 µL L(-1)) CO2 . Elevated CO2 significantly (P < 0.05) increased Pn (105%-149%), Pnmax (38.8%-63.0%) and AQY (20.0%-34.8%) of S. alfredii in all the Cd treatments, but reduced chlorophyll concentration, dark respiration and photorespiration. After 10 days growth in medium with 50 µM Cd under elevated CO2 , PSII activities were significantly enhanced (P < 0.05) with Pm, Fv/Fm, Φ(II) and qP increased by 66.1%, 7.5%, 19.5% and 16.4%, respectively, as compared with ambient-grown plants. Total Cd uptake in shoot of S. alfredii grown under elevated CO2 was increased by 44.1%-48.5%, which was positively correlated with the increase in Pn. These results indicate that elevated CO2 promoted the growth of S. alfredii due to increased photosynthetic carbon uptake rate and photosynthetic light-use efficiency, and showed great potential to improve the phytoextraction of Cd by S. alfredii.

Role of sulfur assimilation pathway in cadmium hyperaccumulation by Sedum alfredii Hance.

Sedum alfredii Hance is a promising cadmium (Cd) hyperaccumulating plant recently identified in China. However, the physiological and molecular mechanisms underlying Cd accumulation, which differentiate hyperaccumulating ecotype (HE) from non-hyperaccumulating ecotype (NHE) has not been elucidated yet. A hydroponic experiment was conducted to investigate the role of sulfur assimilation pathway in Cd hyperaccumulation by the S. alfredii Hance, by analyzing gene expression pattern in sulfur assimilation pathway and the concentration of some sulfur containing compounds. The results show that, sulfur assimilation pathway was affected by Cd differently in HE and NHE S. alfredii Hance. The gene expression pattern of sulfur assimilation pathway was regulated differently in HE and NHE plants, especially the nicotianamine synthase (NAS). NAS transcript levels in root of HE was 141-fold higher than NHE, while in shoots of HE only 0.31-fold higher than NHE. In HE roots, NAS expression level was maximum 3171-fold higher than shoots, while in NHE plants roots NAS expression level was maximum 45.3-fold higher than shoots. In HE plant roots, sulfur, cysteine and methionine concentrations increased 30%, 46% and 835% respectively, by Cd treatment, but in NHE plants roots, sulfur concentration increased less than 1%, cysteine and methionine concentrations decreased 78.5% and 13.3% respectively, by Cd. Cd exposure increased glutathione levels by 142% in HE but less than 10% in NHE plant roots.

SpCTP3 from the hyperaccumulator Sedum plumbizincicola positively regulates cadmium tolerance by interacting with SpMDH1.

Cadmium (Cd) is a heavy metal pollutant mainly originating from the discharge of industrial sewage, irrigation with contaminated water, and the use of fertilizers. The phytoremediation of Cd polluted soil depends on the identification of the associated genes in hyperaccumulators. Here, a novel Cd tolerance gene (SpCTP3) was identified in hyperaccumulator Sedum plumbizincicola. The results of Cd2+ binding and thermodynamic analyses, revealed the CXXC motif in SpCTP3 functions is a Cd2+ binding site. A mutated CXXC motif decreased binding to Cd by 59.93%. The subcellular localization analysis suggested that SpCTP3 is primarily a cytoplasmic protein. Additionally, the SpCTP3-overexpressing (OE) plants were more tolerant to Cd and accumulated more Cd than wild-type Sedum alfredii (NHE-WT). The Cd concentrations in the cytoplasm of root and leaf cells were significantly higher (53.75% and 71.87%, respectively) in SpCTP3-OE plants than in NHE-WT. Furthermore, malic acid levels increased and decreased in SpCTP3-OE and SpCTP3-RNAi plants, respectively. Moreover, SpCTP3 interacted with malate dehydrogenase 1 (MDH1). Thus, SpCTP3 helps regulate the subcellular distribution of Cd and increases Cd accumulation when it is overexpressed in plants, ultimately Cd tolerance through its interaction with SpMDH1. This study provides new insights relevant to improving the Cd uptake by Sedum plumbizincicola.

Physiology and transcriptomic analysis revealed the mechanism of silicon promoting cadmium accumulation in Sedum alfredii Hance.

Silicon (Si) effectively promote the yield of many crops, mainly due to its ability to enhance plants resistance to stress. However, how Si helps hyperaccumulators to extract Cadmium (Cd) from soil has remained unclear. In this study, Sedum alfredii Hance (S. alfredii) was used as material to study how exogenous Si affected biomass, Cd accumulation, antioxidation, cell ultrastructure, subcellular distribution and changes in gene expression after Cd exposure. The study has shown that as Si concentration increases (1, 2 mM), the shoot biomass of plants increased by 33.1%-63.6%, the Cd accumulation increased by 31.9%-96.6%, and the chlorophyll, carotenoid content, photosynthetic gas exchange parameters significantly increased. Si reduced Pro and MDA, promoted the concentrations of SOD, CAT and POD to reduce antioxidant stress damage. In addition, Si promoted GSH and PC to chelate Cd in vacuoles, repaired damaged cell ultrastructure, improved the fixation of Cd and cell wall (especially in pectin), and reduced the toxic effects of Cd. Transcriptome analysis found that genes encoding Cd detoxification, Cd absorption and transport were up-regulated by Si supplying, including photosynthetic pathways (PSB, LHCB, PSA), antioxidant defense systems (CAT, APX, CSD, RBOH), cell wall biosynthesis such as pectinesterase (PME), chelation (GST, MT, NAS, GR), Cd absorption (Nramp3, Nramp5, ZNT) and Cd transport (HMA, PCR). Our result revealed the tentative mechanism of Si promotes Cd accumulation and enhances Cd tolerance in S. alfredii, and thereby provides a solid theoretical support for the practical use of Si fertilizer in phytoextraction.

Sedoheptulose accumulation under CO2 enrichment in leaves of Kalanchoë pinnata: a novel mechanism to enhance C and P homeostasis?

In contrast to the well-documented roles of its mono- and bisphosphate esters, the occurrence of free sedoheptulose in plant tissues remains a matter of conjecture. The present work sought to determine the origin of sedoheptulose formation in planta, as well as its physiological importance. Elevated CO2 and sucrose induction experiments were used to study sedoheptulose metabolism in the Crassulacean acid metabolism (CAM) plants Kalanchoë pinnata and Sedum spectabile. Experimental evidence suggested that sedoheptulose is produced from the oxidative pentose phosphate pathway intermediate sedoheptulose-7-phosphate, by a sedoheptulose-7-phosphate phosphatase. Carbon flux through this pathway was stimulated by increased triose-phosphate levels (elevated CO2, compromised sink availability, and sucrose incubation of source leaves) and attenuated by ADP and inorganic phosphate (Pi). The accumulation of free sedoheptulose is proposed to act as a mechanism contributing to both C and P homeostasis by serving as an alternative carbon store under elevated CO2 or a compromised sink capacity to avoid sucrose accumulation, depletion of inorganic phosphate, and suppression of photosynthesis. It remains to be established whether this acclimation-avoiding mechanism is confined to CAM plants, which might be especially vulnerable to Pi imbalances, or whether some C3 and C4 plants also dispose of the genetic capacity to induce and accelerate sedoheptulose synthesis upon CO2 elevation.

Cellular sequestration of cadmium in the hyperaccumulator plant species Sedum alfredii.

Spatial imaging of cadmium (Cd) in the hyperaccumulator Sedum alfredii was investigated in vivo by laser ablation inductively coupled plasma mass spectrometry and x-ray microfluorescence imaging. Preferential Cd accumulation in the pith and cortex was observed in stems of the Cd hyperaccumulating ecotype (HE), whereas Cd was restricted to the vascular bundles in its contrasting nonhyperaccumulating ecotype. Cd concentrations of up to 15,000 µg g(-1) were measured in the pith cells, which was many fold higher than the concentrations in the stem epidermis and vascular bundles in the HE plants. In the leaves of the HE, Cd was mainly localized to the mesophyll and vascular cells rather than the epidermis. The distribution pattern of Cd in both stems and leaves of the HE was very similar to calcium but not zinc, irrespective of Cd exposure levels. Extended x-ray absorption fine structure spectroscopy analysis showed that Cd in the stems and leaves of the HE was mainly associated with oxygen ligands, and a larger proportion (about 70% in leaves and 47% in stems) of Cd was bound with malic acid, which was the major organic acid in the shoots of the plants. These results indicate that a majority of Cd in HE accumulates in the parenchyma cells, especially in stems, and is likely associated with calcium pathways and bound with organic acid (malate), which is indicative of a critical role of vacuolar sequestration of Cd in the HE S. alfredii.

Heavy metal phytoextraction by Sedum alfredii is affected by continual clipping and phosphorus fertilization amendment.

Improving the efficacy of phytoextraction is critical for its successful application in metal contaminated soils. Mineral nutrition affects plant growth and metal absorption and subsequently the accumulation of heavy metal through hyper-accumulator plants. This study assessed the effects of di-hydrogen phosphates (KH2PO4, Ca(H2PO4)2, NaH2PO4 and NH4H2PO4) application at three levels (22, 88 and 352 mg P/kg soil) on Sedum alfredii growth and metal uptake by three consecutive harvests on aged and Zn/Cd combined contaminated paddy soil. The addition of phosphates (P) significantly increased the amount of Zn taken up by S. alfredii due to increased shoot Zn concentration and dry matter yield (DMY) (P < 0.05). The highest phytoextraction of Zn and Cd was observed in KH2PO4 and NH4H2PO4 treatment at 352 mg P/kg soil. The amount of Zn removed by phytoextraction increased in the order of 1st clipping < 2nd clipping < 3rd clipping, and for Cd extraction the order was 2nd clipping < 1st clipping < 3rd clipping. These results indicate that the application of P fertilizers coupled with multiple cuttings can enhance the removal of Zn and Cd from contaminated soils by S. alfredii, thus shortening the time needed for accomplishing remediation goals.

Genome-wide characterization of the hyperaccumulator Sedum alfredii F-box family under cadmium stress.

The F-box genes, which form one of the largest gene families in plants, are vital for plant growth, development and stress response. However, F-box gene family in Sedum alfredii remains unknown. Comprehensive studies addressing their function responding to cadmium stress is still limited. In the present study, 193 members of the F-box gene (SaFbox) family were identified, which were classified into nine subfamilies. Most of the SaFboxs had highly conserved domain and motif. Various functionally related cis-elements involved in plant growth regulation, stress and hormone responses were located in the upstream regions of SaFbox genes. RNA-sequencing and co-expression network analysis revealed that the identified SaFbox genes would be involved in Cd stress. Expression analysis of 16 hub genes confirmed their transcription level in different tissues. Four hub genes (SaFbox40, SaFbox51, SaFbox136 and SaFbox170) were heterologously expressed in a Cd-sensitive yeast cell to assess their effects on Cd tolerance. The transgenic yeast cells carrying SaFbox40, SaFbox51, SaFbox136, or SaFbox170 were more sensitive and accumulated more cadmium under Cd stress than empty vector transformed control cells. Our results performed a comprehensive analysis of Fboxs in S. alfredii and identified their potential roles in Cd stress response.