Mechanism Of Heavy Metal Tolerance Research Articles

Genome architecture of the heavy metal tolerant and accumulator Hirschfeldia incana: Insights from genome sequencing, assembly, and comparative analysis

Hirschfeldia incana L., a member of the Brassicaceae family commonly found in Mediterranean regions, is known for its capacity to withstand and accumulate heavy metals, particularly lead (Pb) both in soil environments and hydroponic systems. This plant has been used as a model to study plant responses to heavy metals. Nonetheless, the molecular mechanisms underlying its tolerance and heavy metal accumulation are not fully understood, partly because of the limited knowledge about its genome. In this study, the genome of H. incana was sequenced, assembled, characterized, and annotated. Approximately 8.6 Gpb of data were generated using Oxford Nanopore Technology (ONT), resulting in a genome assembly of 390 Mb, comprising 5196 contigs with an N50 exceeding 131 kb. The genome had a BUSCO score of 97.2 %, with 38,454 genes and a repetition content of 38.25 %. Subsequently, the assembled genome was annotated using several databases including GO, InterPro, MetaCyc, PANTHER, Pfam, Reactome, SUPERFAMILY, and KEGG. This annotation yielded 22,661 GO terms and 143 KEGG maps. A comparative genomic analysis between H. incana and six Brassicaceae species (five hyperaccumulators of heavy metals and one non-hyperaccumulator) was also conducted. This analysis revealed that H. incana shares a substantial proportion of orthologous genes (89.7 % of orthogroups) with six Brassicaceae species. The generated phylogenetic tree suggests that H. incana is closely related to B. juncea, B. napus, and B. oleracea, indicating a common ancestry and potentially shared genetic factors contributing to hyperaccumulation in these species. Moreover, the copy number of twenty-nine genes involved in heavy metal tolerance and accumulation mechanisms in H. incana and six brassicaceae were assayed. This analysis revealed that H. incana and other hyperaccumulator species possess a higher copy number of genes related to heavy metal tolerance than the sensitive plant A. thaliana. Notably, the variation in gene copy numbers highlights their potential role in the adaptation of H. incana to heavy metal stress. This study provides a comprehensive genomic framework that enhance our understanding of H. incana adaptation to heavy metal stress, and offers valuable data for further genomic investigations of the molecular mechanisms of heavy metal tolerance and accumulation in plants.

Root endophytic Phialocephala fortinii and Talaromyces verruculosus enhance growth and affect heavy metal tolerance of Miscanthus sinensis Andersson growing naturally at a mine site

ABSTRACT Miscanthus sinensis Andersson naturally grows at mining sites and accumulates heavy metals, suggesting heavy metal tolerance. This study aimed to elucidate the heavy metal tolerance mechanisms of M. sinensis, which naturally grew at a mine site, considering root endophytes. Elemental analysis by inductively coupled plasma-optical emission spectroscopy showed that M. sinensis highly accumulated zinc and lead in adventitious roots and dead root bark, exhibiting tolerance via these elemental removals in dead root bark. Phenolic analysis by high-performance liquid chromatography revealed chlorogenic acid production in adventitious roots. Phialocephala fortinii, which showed the highest appearance, and Talaromyces verruculosus, which produced gluconic acid as a zinc detoxicant, were isolated from adventitious roots of M. sinensis. In inoculation tests, P. fortinii and T. verruculosus significantly enhanced seedling growth by increasing potassium uptake and possibly indole-3-acetic acid production. Our results provide fundamental knowledge for using M. sinensis and root endophytes for revegetation at mining sites.

Heavy metal tolerance and detoxification mechanism mediated by heavy metal resistance genes in compost habitat.

Heavy metal pollution from compost is one of the most concerned environmental problems, which poses a threat to the ecosystem and human health. This study aims to reveal the heavy metal tolerance and detoxification mechanism mediated by heavy metal resistance genes (HMRGs) in compost habitat through metagenomics combined with chemical speciation analysis of heavy metals. The results showed that there were 37 HMRGs corresponding to 7 common heavy metal(loid)s in composting, and they had the ability to transform heavy metals into stable or low-toxic speciation by regulating enzyme transport, redox, methylation, etc. This study summarized the heavy metal metabolism pathway mediated by HMRGs, providing a new perspective for understanding the transformation of heavy metals in the composting process.

Unveiling Metal Tolerance Mechanisms in Leersia hexandra Swartz under Cr/Ni Co-Pollution by Studying Endophytes and Plant Metabolites.

Heavy metal pollution poses significant environmental challenges, and understanding how plants and endophytic bacteria interact to mitigate these challenges is of utmost importance. In this study, we investigated the roles of endophytic bacteria, particularly Chryseobacterium and Comamonas, in Leersia hexandra Swartz (L. hexandra) in response to chromium and nickel co-pollution. Our results demonstrated the remarkable tolerance of Chryseobacterium and Comamonas to heavy metals, and their potential to become dominant species in the presence of co-pollution. We observed a close relationship between these endophytic bacteria and the significant differences in metabolites, particularly carbohydrates, flavonoids, and amino acids in L. hexandra. These findings shed light on the potential of endophytic bacteria to promote the production of aspartic acid and other metabolites in plants as a response to abiotic stressors. Furthermore, our study presents a new direction for plant and bioremediation strategies in heavy metal pollution and enhances our understanding of L. hexandra's mechanisms for heavy metal tolerance.

Unravelling the Mechanisms of Heavy Metal Tolerance: Enhancement in Hydrophilic Antioxidants and Major Antioxidant Enzymes Is Not Crucial for Long-Term Adaptation to Copper in Chlamydomonas reinhardtii.

Understanding of the mechanisms of heavy metal tolerance in algae is important for obtaining strains that can be applied in wastewater treatment. Cu is a redox-active metal directly inducing oxidative stress in exposed cells. The Cu-tolerant Chlamydomonas reinhardtii strain Cu2, obtained via long-term adaptation, displayed increased guaiacol peroxidase activity and contained more lipophilic antioxidants, i.e., α-tocopherol and plastoquinol, than did non-tolerant strain N1. In the present article, we measured oxidative stress markers; the content of ascorbate, soluble thiols, and proline; and the activity of superoxide dismutase (SOD), catalase (CAT), and ascorbate peroxidase (APX) in N1 and Cu2 strains grown in the absence or presence of excessive Cu. The Cu2 strain displayed less pronounced lipid peroxidation and increased APX activity compared to N1. The amount of antioxidants was similar in both strains, while SOD and CAT activity was lower in the Cu2 strain. Exposure to excessive Cu led to a similar increase in proline content in both strains and a decrease in ascorbate and thiols, which was more pronounced in the N1 strain. The Cu2 strain was less tolerant to another redox-active heavy metal, namely chromium. Apparently other mechanisms, probably connected to Cu transport, partitioning, and chelation, are more important for Cu tolerance in Cu2 strain.

Insights into heavy metal tolerance mechanisms of Brassica species: physiological, biochemical, and molecular interventions.

Heavy metal (HM) contamination of soil due to anthropogenic activities has led to bioaccumulation and biomagnification, posing toxic effects on plants by interacting with vital cellular biomolecules such as DNA and proteins. Brassica species have developed complex physiological, biochemical, and molecular mechanisms for adaptability, tolerance, and survival under these conditions. This review summarizes the HM tolerance strategies of Brassica species, covering the role of root exudates, microorganisms, cell walls, cell membranes, and organelle-specific proteins. The first line of defence against HM stress in Brassica species is the avoidance strategy, which involves metal ion precipitation, root sorption, and metal exclusion. The use of plant growth-promoting microbes, Pseudomonas, Psychrobacter, and Rhizobium species effectively immobilizes HMs and reduces their uptake by Brassica roots. The roots of Brassica species efficiently detoxify metals, particularly by flavonoid glycoside exudation. The composition of the cell wall and callose deposition also plays a crucial role in enhancing HMs resistance in Brassica species. Furthermore, plasma membrane-associated transporters, BjCET, BjPCR, BjYSL, and BnMTP, reduce HM concentration by stimulating the efflux mechanism. Brassica species also respond to stress by up-regulating existing protein pools or synthesizing novel proteins associated with HM stress tolerance. This review provides new insights into the HM tolerance mechanisms of Brassica species, which are necessary for future development of HM-resistant crops.

Functional characterization of rice metallothionein OsMT-I-Id: Insights into metal binding and heavy metal tolerance mechanisms

Metallothioneins (MTs) are cysteine-rich proteins known for their strong metal-binding capabilities, making them effective in detoxifying heavy metals (HMs). This study focuses on characterizing the functional properties of OsMT-I-Id, a type-I Metallothionein found in rice. Using a HM-responsive yeast cup1Δ (DTY4), ycf1∆ (for cadmium), and acr3∆ mutants (for trivalent arsenic), we assessed the impact of OsMT-I-Id on metal accumulation and cellular resilience. Our results demonstrated that yeast cells expressing OsMT-I-Id showed increased tolerance and accumulated higher levels of copper (Cu), arsenic (As), and cadmium (Cd), compared to control cells. This can be attributed to the protein's ability to chelate and bind HMs. Site-directed mutagenesis was employed to investigate the specific contributions of cysteine residues. The study revealed that yeast cells with a mutated C-domain displayed heightened HM sensitivity, while cells with a mutated N-domain exhibited reduced sensitivity. This underscores the critical role of C-cysteine-rich domains in metal binding and tolerance of type-I rice MTs. Furthermore, the study identified the significance of the 12th cysteine position at the N-domain and the 68th and 72nd cysteine positions at the C-domain in influencing OsMT-I-Id metal-binding capacity. This research provides novel insights into the structure-function relationship and metal binding properties of type-I plant MTs.

24-Epibrassinolide confers zinc stress tolerance in watermelon seedlings through modulating antioxidative capacities and lignin accumulation.

Zinc (Zn) is an important element in plants, but over-accumulation of Zn is harmful. It is well-known that brassinolide (BR) plays a key role in the regulation of abiotic stress responses in plants. However, the effects of brassinolide on alleviating Zn phytotoxicity in watermelon (Citrullus lanatus L.) seedlings are not clear. The purpose of this study was to study the effect of 24-epibrassinolide (EBR, one of the bioactive BRs) on Zn tolerance of watermelon seedlings and its potential resistance mechanism. Exposure to excessive Zn significantly inhibited shoot and root fresh weight of watermelon, but this could be significantly alleviated by the optimum 0.05 μM EBR. Exogenous spraying EBR increased the pigments and alleviated oxidative damage caused by Zn through reducing Zn accumulation and the levels of reactive oxygen species (ROS) and malonaldehyde (MDA) and increasing the activities of antioxidant enzymes and contents of ascorbic acid (AsA) and glutathione (GSH). Importantly, the relative mRNA levels of antioxidant genesincluding Cu/Zn-superoxidedismutase (Cu-Zn SOD), catalase (CAT), ascorbic acid peroxidase (APX), and glutathione reductase (GR) were significantly induced after EBR treatment. In addition, EBR pre-treatment induced lignin accumulation under Zn stress, and the activity of phenylalanine ammonia-lyase (PAL) and 4-coumaric ligase (4CL), two key enzymes regulating lignin synthesis, also tended to be consistent. Collectively, the present research proves the beneficial effects of EBR in response to Zn stress through enhancing antioxidant defense and lignin accumulation and provides a new insight into the mechanism of BR-enhancing heavy metal tolerance.

Physiological and DNA methylation analysis provides epigenetic insights into kenaf cadmium tolerance heterosis

Soil heavy metal pollution is one of the most challenging problems. Kenaf is an important natural fiber crop with strong heterosis and a higher tolerance to heavy metals. However, little is known about the molecular mechanisms of kenaf heavy metal tolerance, especially the mechanism of genomic DNA methylation regulating heterosis. In this study, kenaf cultivars CP085, CP089, and their hybrid F1 seedlings were subjected to 300 µM cadmium stress and found obvious heterosis of cadmium resistance in morphology and antioxidant enzyme activity of F1 hybrid seedlings. Through methylation-sensitive amplification polymorphism (MSAP) analysis, we highlighted that the total DNA methylation level under cadmium decreased by 16.9 % in F1 and increased by 14.0 % and 3.0 % in parents CP085 and CP089, respectively. The hypomethylation rate was highest (21.84 %), but hypermethylation was lowest (17.24 %) in F1 compared to parent cultivars. In particular, principal coordinates analysis (PCoA) indicates a significant epigenetic differentiation between F1 and its parents under cadmium. Furthermore, 21 differentially methylated DNA fragments (DMFs) were analyzed. Especially, the expression of NPF2.7, NADP-ME, NAC71, TPP-D, LRR-RLKs, and DHX51 genes were changed due to cadmium stress and related to cytosine methylation regulation. Finally, the knocked-down of the differentially methylated gene NPF2.7 by virus-induced gene silencing (VIGS) resulted in increased sensitivity of kenaf seedlings under cadmium stress. It is speculated that low DNA methylation levels can regulate gene expression that led to the heterosis of cadmium tolerance in kenaf.

Abscisic-Acid-Regulated Responses to Alleviate Cadmium Toxicity in Plants.

High levels of cadmium (Cd) in soil can cause crop yield reduction or death. Cadmium accumulation in crops affects human and animal health as it passes through the food chain. Therefore, a strategy is needed to enhance the tolerance of crops to this heavy metal or reduce its accumulation in crops. Abscisic acid (ABA) plays an active role in plants' response to abiotic stress. The application of exogenous ABA can reduce Cd accumulation in shoots of some plants and enhance the tolerance of plants to Cd; therefore, ABA may have good application prospects. In this paper, we reviewed the synthesis and decomposition of ABA, ABA-mediated signal transduction, and ABA-mediated regulation of Cd-responsive genes in plants. We also introduced physiological mechanism underlying Cd tolerance because of ABA. Specifically, ABA affects metal ion uptake and transport by influencing transpiration and antioxidant systems, as well as by affecting the expression of metal transporter and metal chelator protein genes. This study may provide a reference for further research on the physiological mechanism of heavy metal tolerance in plants.

Insights from comparative transcriptome analysis in the responses of Pb-tolerant fungi Curvularia tsudae to Pb stress

The fungus Curvularia tsudae can survive in environments that are extremely contaminated by heavy metals; however, the underlying molecular mechanisms of heavy metal tolerance are not clear. In this study, we determined the effects of lead (Pb) stress on the growth of C. tsudae and used RNA-Seq to identify significant genes and biological processes involved. The present study showed that C. tsudae had an outstanding resistant capacity to Pb stress and could survive at a concentration of 1600 mg L−1 Pb. Although an obvious inhibition on the growth was observed, the fungus exhibited tolerance as it continued to grow at a Pb concentration of 1600 mg L−1 for seven days. A total of 9997 (9020 up and 977 down) differentially expressed genes (DEGs) were detected in the mycelium of C. tsudae at Pb free (0 mg L−1) and Pb stressed samples. Pathway enrichment analysis identified several biological processes for managing Pb stress. Genes involved in carbohydrate metabolism tended to be modulated in response to Pb stress, while amino acids and the lipid metabolism would also be induced by Pb stress, and up-regulated genes involved in antioxidant substances and ABC transporters may be committed to high Pb tolerance. Our study contributes to the current literature on C. tsudae response to Pb stress and provides a useful reference for fungi as bioremediators in heavy metal-contaminated environments.

Genomic and Transcriptomic Analysis Reveal Multiple Strategies for the Cadmium Tolerance in Vibrio parahaemolyticus N10-18 Isolated from Aquatic Animal Ostrea gigas Thunberg.

The waterborne Vibrio parahaemolyticus can cause acute gastroenteritis, wound infection, and septicemia in humans. Pollution of heavy metals in aquatic environments is proposed to link high incidence of the multidrug-resistant (MDR) pathogen. Nevertheless, the genome evolution and heavy metal tolerance mechanism of V. parahaemolyticus in aquatic animals remain to be largely unveiled. Here, we overcome the limitation by characterizing an MDR V. parahaemolyticus N10-18 isolate with high cadmium (Cd) tolerance using genomic and transcriptomic techniques. The draft genome sequence (4,910,080 bp) of V. parahaemolyticus N10-18 recovered from Ostrea gigas Thunberg was determined, and 722 of 4653 predicted genes had unknown function. Comparative genomic analysis revealed mobile genetic elements (n = 11) and heavy metal and antibiotic-resistance genes (n = 38 and 7). The bacterium significantly changed cell membrane structure to resist the Cd2+ (50 μg/mL) stress (p < 0.05). Comparative transcriptomic analysis revealed seven significantly altered metabolic pathways elicited by the stress. The zinc/Cd/mercury/lead transportation and efflux and the zinc ATP-binding cassette (ABC) transportation were greatly enhanced; metal and iron ABC transportation and thiamine metabolism were also up-regulated; conversely, propanoate metabolism and ribose and maltose ABC transportation were inhibited (p < 0.05). The results of this study demonstrate multiple strategies for the Cd tolerance in V. parahaemolyticus.

Transcriptome analysis of trembling aspen (Populus tremuloides) under nickel stress.

Plants have evolved heavy metal tolerance mechanisms to adapt and cope with nickel (Ni) toxicity. Decrypting whole gene expression of Trembling Aspen (Pinus tremuloides) under nickel stress could elucidate the nickel resistance/tolerance mechanisms. The main objectives of the present research were to 1) characterize the P. tremuloides transcriptome, and 2) compare gene expression dynamics between nickel-resistant and nickel-susceptible P. tremuloides genotypes with Whole Transcriptome (WT) sequencing. Illumina Sequencing generated 27–45 million 2X150 paired-end reads of raw data per sample. The alignment performed with StringTie Software added two groups of transcripts to the draft genome annotation. One group contained 32,677 new isoforms that match to 17,254 genes. The second group contained 17,349 novel transcripts that represent 16,157 novel genes. Overall, 52,987 genes were identified from which 36,770 genes were selected as differently expressed. With the high stringency (two-fold change, FDR value ≤ 0.05 and logFC value ≥1 (upregulated) or ≤ -1 (downregulated), after GSEA analysis and filtering for gene set size, 575 gene sets were upregulated and 146 were downregulated in nickel resistant phenotypes compared to susceptible genotypes. For biological process, genes associated with translation were significantly upregulated while signal transduction and cellular protein process genes were downregulated in resistant compared to susceptible genotypes. For molecular function, there was a significant downregulation of genes associated with DNA binding in resistant compared to susceptible lines. Significant upregulation was observed in genes located in ribosome while downregulation of genes in chloroplast and mitochondrion were preponderant in resistant genotypes compared to susceptible. Hence, from a whole transcriptome level, an upregulation in ribosomal and translation activities was identified as the main response to Ni toxicity in the resistant plants. More importantly, this study revealed that a metal transport protein (Potrs038704g29436 –ATOX1-related copper transport) was among the top upregulated genes in resistant genotypes when compared to susceptible plants. Other identified upregulated genes associated with abiotic stress include genes coding for Dirigent Protein 10, GATA transcription factor, Zinc finger protein, Auxin response factor, Bidirectional sugar transporter, and thiamine thiazole synthase.

Reprisal of Schima superba to Mn stress and exploration of its defense mechanism through transcriptomic analysis.

One of the most diverse protein families, ATP-binding cassette (ABC) transporters, play a role in disease resistance, heavy metal tolerance, and food absorption.Differentially expressed genes contribute in the investigation of plant defense mechanisms under varying stress conditions. To elucidate the molecular mechanisms involved in Mn metal stress, we performed a transcriptomic analysis to explore the differential gene expression in Schima superba with the comparison of control. A total of 79.84 G clean data was generated and 6558 DEGs were identified in response to Mn metal stress. Differentially expressed genes were found to be involved in defense, signaling pathways, oxidative burst, transcription factors and stress responses. Genes important in metal transport were more expressive in Mn stress than control plants. The investigation of cis-acting regions in the ABC family indicated that these genes might be targeted by a large variety of trans-acting elements to control a variety of stress circumstances. Moreover, genes involved in defense responses, the mitogen-activated protein kinase (MAPK) signaling and signal transduction in S. superba were highly induced in Mn stress. Twenty ABC transporters were variably expressed on 1st, 5th, and 10th day of Mn treatment, according to the qRT PCR data. Inclusively, our findings provide an indispensable foundation for an advanced understanding of the metal resistance mechanisms. Our study will enrich the sequence information of S. superba in a public database and would provide a new understanding of the molecular mechanisms of heavy metal tolerance and detoxification.

Soil applied silicon and manganese combined with foliar application of 5-aminolevulinic acid mediate photosynthetic recovery in Cd-stressed Salvia miltiorrhiza by regulating Cd-transporter genes.

Salvia miltiorrhiza is an important medicinal plant that experiences significant growth and biomass losses when cultivated on cadmium (Cd) contaminated soils. High Cd accumulation in plant tissues also increases the risk of metal entry into the food chain. In this study, we proposed that Cd accumulation in S. miltiorrhiza can be restricted through plant growth regulators and nutrient management. Therefore, S. miltiorrhiza seedlings were transplanted into mixed nutrient soil for two weeks, then treated with 30 mg kg-1 CdCl2, 200 mg kg-1 Na2SiO3·9H2O, and 100 mg kg-1 MnSO4, and simultaneously sprayed with 10 mg L-1 ALA on the leaves one week later. This study showed that elevated Cd accumulation significantly reduced plant growth and biomass. This growth inhibition damaged photosynthetic machinery and impaired carbon assimilation. In contrast, 5-aminolevulinic acid (ALA) significantly promoted the biomass of S. miltiorrhiza, and the dry weight of plants treated with ALA combined with manganese (Mn)/silicon (Si) increased by 42% and 55% as compared with Cd+Mn and Cd+Si treatments. Exogenously applied ALA and Si/Mn significantly activated antioxidant enzymes and promoted the growth recovery of S. miltiorrhiza. Further, exogenous ALA also reduced the Cd concentration in S. miltiorrhiza, especially when combined with Si. Compared with the Cd+Si treatment, the Cd+Si+ALA treatment reduced the Cd concentration in roots and leaves by 59% and 60%, respectively. Gene expression analysis suggested that ALA and Si significantly up-regulated genes associated with Cd transport. Other genes related to heavy metal tolerance mechanisms are also regulated to cope with heavy metal stress. These results indicated that the combined action of ALA and Si/Mn could reduce Cd-toxicity by increasing chlorophyll content and changing oxidative stress and can also affect Cd accumulation by regulating gene expression.

Co-over expression of Ascorbate Glutathione pathway enzymes improve mercury tolerance in tomato

The genetic modification of plants for the removal of inorganic pollutants from contaminated soil and water bodies is an emerging area for addressing environmental concerns. This approach is based on the ability of plants to take up and accumulate heavy metals, with efficiency being dependent on the underlying mechanisms of heavy metal accumulation and tolerance. A robust antioxidant pathway is determinantal for heavy metal uptake and accumulation and, therefore, in this study, we evaluated the transgenic tomato plants installed with Ascorbate Glutathione (ASA-GSH) pathway genes for uptake, accumulation, and response to mercury (Hg). We observed that ASA-GSH overexpressing lines were resilient to Hg stress as they displayed higher photosynthetic activity and increased photosynthetic gas exchange parameters with a concomitant decrease in ion leakage under Hg stress. Additionally, transgenic lines accumulated high osmolytes and showed enhanced activity of antioxidant enzymes. Moreover, the results of SEM and confocal microscopy confirmed least damage to plant tissue in ASA-GSH overexpressing lines compared to wild-type under Hg-stress which was further supported by Atomic absorption study that revealed a significant decline in Hg accumulation in the leaves of transgenic lines compared to wild-type under stress conditions. In conclusion, pyramiding of ASA-GSH pathway genes in tomato plants is an efficient approach for the development of Hg-resistant tomato plants and the reclamation of Hg-contaminated sites.

Phytoremediation potential evaluation of three rhubarb species and comparative analysis of their rhizosphere characteristics in a Cd- and Pb-contaminated soil

Screening or breeding exceptional plant species for heavy metal phytoremediation is as important as adopting feasible measures to enhance phytoremediation efficiency, which are largely based on clarifying the mechanisms of heavy metal tolerance and accumulation by plants. In this study, cadmium (Cd) and lead (Pb) tolerance and accumulation characteristics of Rheum officinale, R. palmatum, and R. tanguticum were analysed to assess their phytoremediation potential. The seed germination test indicated that these three rhubarb species could tolerate 10 mg L−1 Cd and 100 mg L−1 Pb. However, when sown in Cd- and Pb-contaminated soil, all three rhubarb species exhibited a relatively high Cd accumulation capacity but a considerably low Pb accumulation capacity according to the bioconcentration factors of Cd (0.42–0.47 in shoots and 0.11–0.15 in roots) and Pb (0.004–0.008 in shoots and 0.007–0.013 in roots). The high Cd translocation factors (3.04–4.24) indicated that these three rhubarb species were suitable for Cd phytoextraction. The changes in rhizospheric physicochemical indices were generally similar among the three rhubarb plants in comparison with those of the unplanted soil. However, differential indicator rhizobacteria were identified for the three rhubarb plants, which may be primarily attributed to their different root system characteristics. These enriched rhizobacteria included many plant growth-promoting bacteria, and several of them were also involved in regulating heavy metal uptake by plants, indicating that three rhubarb species likely recruit differentially beneficial rhizobacteria to maintain plant growth and vitality and to regulate heavy metal uptake in the Cd- and Pb-polluted soil. This study identifies new candidate plant resources for the phytoremediation of Cd-polluted soils and provides novel insights into understanding the interactions among heavy metals, rhizobacteria, and plants.

Zn tolerance in the evergreen shrub, Aucuba japonica, naturally growing at a mine site: Cell wall immobilization, aucubin production, and Zn adsorption on fungal mycelia.

Aucuba japonica Thunb. is an evergreen understory shrub that grows naturally at a mine site. The mine soil contains high concentrations of heavy metals, and A. japonica appears to maintain detoxification mechanisms against heavy metals in the study site’s understory. This study aimed to investigate the heavy metal tolerance mechanisms in A. japonica, considering the possible roles of arbuscular mycorrhizal and root-endophytic fungi. We conducted fieldwork in summer (canopy-foliation season) and winter (canopy-defoliation season) to measure the heavy metal concentrations in leaves, branches, and roots and analyze possible detoxicants in the roots. The infection rates of arbuscular mycorrhizal and root-endophytic fungi were evaluated via microscopic observation, and heavy metal (Zn) localization in A. japonica roots was observed using confocal laser scanning microscopy. Field analysis showed that A. japonica accumulated excessive Zn and produced aucubin and citric acid in the roots in both summer and winter. Zn localization observations clarified that Zn was distributed in thickened epidermal and cortical cell walls, suggesting that the cell walls functioned as Zn deposition sites, reducing Zn toxicity. It was further clarified that Zn was contained within cortical cells, indicating that Zn might be detoxified by aucubin and citric acid. Arbuscular mycorrhizal and root-endophytic fungi within cortical cells adsorbed Zn on fungal cell walls, indicating that these fungi would reduce Zn content within root cells and might alleviate Zn toxicity. Our results indicated that A. japonica would maintain Zn tolerance in both summer and winter via Zn immobilization in the cell walls and production of aucubin and citric acid, and that arbuscular mycorrhizal and root-endophytic fungi might play important roles in the Zn tolerance of A. japonica.

The Endophytic Fungus Piriformospora Indica-Assisted Alleviation of Cadmium in Tobacco.

Increasing evidence suggests that the endophytic fungus Piriformospora indica helps plants overcome various abiotic stresses, especially heavy metals. However, the mechanism of heavy metal tolerance has not yet been elucidated. Here, the role of P. indica in alleviating cadmium (Cd) toxicities in tobacco was investigated. It was found that P. indica improved Cd tolerance to tobacco, increasing Cd accumulation in roots but decreasing Cd accumulation in leaves. The colonization of P. indica altered the subcellular repartition of Cd, increasing the Cd proportion in cell walls while reducing the Cd proportion in membrane/organelle and soluble fractions. During Cd stress, P. indica significantly enhanced the peroxidase (POD) activity and glutathione (GSH) content in tobacco. The spatial distribution of GSH was further visualized by Raman spectroscopy, showing that GSH was distributed in the cortex of P. indica-inoculated roots while in the epidermis of the control roots. A LC-MS/MS-based label-free quantitative technique evaluated the differential proteomics of P. indica treatment vs. control plants under Cd stress. The expressions of peroxidase, glutathione synthase, and photosynthesis-related proteins were significantly upregulated. This study provided extensive evidence for how P. indica enhances Cd tolerance in tobacco at physiological, cytological, and protein levels.

Identification of novel heavy metal detoxification proteins in Solanum tuberosum: Insights to improve food security protection from metal ion stress.

With increasingly serious environmental pollution problems, research has focused on identifying functional genes within plants that can help ensure food security and soil governance. In particular, plants seem to have been able to evolve specific functional genes to respond to environmental changes by losing partial gene functions, thereby representing a novel adaptation mechanism. Herein, a new category of functional genes was identified and investigated, providing new directions for understanding heavy metal detoxification mechanisms. Interestingly, this category of proteins appears to exhibit specific complexing functions for heavy metals. Further, a new approach was established to evaluate ATP-binding cassette (ABC) transporter family functions using microRNA targeted inhibition. Moreover, mutant and functional genes were identified for future research targets. Expression profiling under five heavy metal stress treatments provided an important framework to further study defense responses of plants to metal exposure. In conclusion, the new insights identified here provide a theoretical basis and reference to better understand the mechanisms of heavy metal tolerance in potato plants. Further, these new data provide additional directions and foundations for mining gene resources for heavy metal tolerance genes to improve safe, green crop production and plant treatment of heavy metal soil pollution.