Genetic variation and gene expression of anthocyanin synthesis and transport related enzymes in Oryza sativa against thiocyanate.

Plants exposed to environmental contaminants often synthesize anthocyanins (ATHs) as an approach to safeguard themselves from adverse impact. However, the overload of ATHs in plant cells can threaten their growth and development through proteins oxidization and intercalating with DNAs inside cells. In the present study, a microcosm hydroponic experiment was conducted using rice seedlings to investigate the molecular signaling pathways involved in regulating and controlling ATHs synthesis and transport exposed to thiocyanate (SCN-). Our results indicated that SCN- exposure significantly (p < 0.05) increased the expression of ATHs synthesis related genes (i.e., PAL, CHS, ANS, UFGT genes) in rice tissues, altered the activities of these ATHs synthesis related enzymes, and consequently elevated the ATHs content. However, SCN- exposure significantly decreased the expression of ATHs transport related genes (i.e., GST, ABC, MATE genes) in rice seedlings, suggesting that SCN- exposure have restrained ATHs transport from cytosol to vacuole in cells, eventually posing a significant adverse effect on cells survival. Our findings highlight on one of the plant aspects in managing the toxicity triggered by secondary metabolites under stress conditions.

Transcriptomic analysis of cytochrome P450 genes and pathways involved in chromium toxicity in Oryza sativa.

In plants, cytochrome P450 monooxygenase (CYP) plays an important role in detoxifying xenobiotic chemicals and coordinating abiotic stresses. Agilent 44 K rice microarray has been used to focus on the transcriptional profile of osCYP genes in rice seedling exposed to Cr solution containing K2CrO4 or Cr(NO3)3. Our study showed that expression profiles of 264 osCYP genes identified were tissue, dose and stimulus specific in rice seedlings. Comparative genomics analysis revealed that more differentially expressed osCYP genes were discovered in roots than in shoots under both Cr exposures. Results from Venn diagram analysis of differentially expressed osCYP genes demonstrated that there were common osCYP genes and unique osCYP genes present in different rice tissue as well as in different Cr treatments, which may control and/or regulate involvement of different CYP isoenzymes under Cr exposure individually or combinedly. KEGG analysis indicated that significant up- and down-regulated osCYP genes in rice tissues were chiefly related to "biosynthesis of secondary metabolites". However, involvements of osCYP genes mapped in the "biosynthesis of secondary metabolites" were tissue and dose specific, implying their distinctly responsive and adaptive mechanisms during Cr exposure. Overall, our findings are evident to describe and clarify their individual roles of specific osCYP genes in regulating involvement of CYP isoforms in Cr detoxification by rice seedlings.

Regulation Network of Sucrose Metabolism in Response to Trivalent and Hexavalent Chromium in Oryza sativa.

The presence of chromium (Cr) in cultivated fields affects carbohydrate metabolism of rice (Oryza sativa L.) and weakens its productivity. Little is known about the molecular mechanism of sucrose metabolism underlying Cr stress response in rice plants. In the present study, the transcriptome map of sucrose metabolism in rice seedlings exposed to both trivalent and hexavalent chromium was investigated using Agilent 4 × 44K rice microarray analysis. Results indicated that Cr exposure (3 days) significantly (p < 0.05) improved sucrose accumulation, and altered the activities of sucrose synthetase, sucrose phosphate phosphatase, and amylosynthease in rice tissues. We identified 119 differentially regulated genes involved in 17 sucrose metabolizing enzymes and found that gene responses in roots were significantly (p < 0.05) stronger than in shoots under both Cr(III) and Cr(VI) treatment. The network maps of gene regulation responsible for sucrose metabolism in rice plants provide a theoretical basis for further cultivating Cr-resistant rice cultivars through molecular genetic improvement.

Impact of water quality on the microbial diversity in the surface water along the Three Gorge Reservoir (TGR), China.

The Three Gorges Reservoir (TGR), one of the world's largest reservoirs, has crucial roles in flood control, power generation, and navigation. The TGR is contaminated because of the human activities, and how the contaminated water influences the distribution of the microbial community have not been well studied. In this study, we collected 41 freshwater samples from 13 main dwelling districts along the TGR to investigate the water quality, the distribution of the microbial community, and how water quality affects the microbial community structure. The sampling sites cover the whole TGR along the stream, with 670 km distance. Our results show that both water quality and the compositions of bacterial community vary along the TGR. The distribution of bacterial community is closely related to the local water quality. There is the highest concentration of chemical oxygen demand (COD), the highest relative abundance of Firmicutes, and the highest relative abundance of Bacillus in the upstream, compared to the middle and down streams. Redundancy analysis (RDA) showed that PO43- and COD were the main environmental factors influencing on the structure of bacterial community. The relative abundance of nitrification and denitrification functional genes also altered along the streams. These findings provide the basic data for water quality, the distribution of bacterial community, the link of environmental factors, and the bacterial community structure along the TGR, which guides the local environmental protection agency to launch protection strategy for maintaining the ecosystem health of the TGR.

Role of cytochrome c in modulating chromium-induced oxidative stress in Oryza sativa.

Cytochrome c (CYTc) is one of the redox-active molecules responsible for electron transport in plant mitochondria. It is important in the interaction of metal ions with mitochondrial proteome, it is also essential for insights into electron transport during oxidative stress. This study focused on the clarification of interactions of CYTc with metal accumulation and associated modulation of CYTc with reactive oxygen species (ROS) generation in rice seedlings exposed to chromium either hexavalent Cr(VI) or trivalent Cr(III). The result indicates that exposures to both Cr species resulted in significant accumulation of Cr in rice tissues. ROS-dependent growth inhibition of rice seedlings was also evident from both Cr treatments. Changes of CYTc content in roots and shoots maintained almost the same pattern after both Cr exposures. However, dynamically transcriptional levels of rice CYTc genes conducted by real-time quantitative RT-PCR were inconsistent in rice tissues. The results presented here demonstrate that oxidative stresses induced by ROS accumulation do not equally and/or detrimentally influence the perturbations of CYTc content and expression patterns of CYTc genes in rice tissues after Cr exposure, suggesting that the decrease of CYTc levels in roots did not originate ROS accumulation, it may be due to metal-binding properties, while enhancement of CYTc content in shoots was probably a signal of severe growth inhibition owing to ROS accumulation.

Microarray-based expression analysis of phytohormone-related genes in rice seedlings during cyanide metabolism.

Plants exhibit highly coordinated, dynamic reactions to various abiotic stressors. As cyanide is a non-essential element for plant growth, entry inside plants can exert toxicity at multiple levels. In plant, hormone plays a pivot role under stress conditions. The fluctuations of stress-responsive hormones help in altering cellular dynamics and hence play a central role in coordination and adaptation growth responses under stress. This study focusses on uptake of cyanide in Oryza sativa seedlings and its effect on physiological and on genetic level. Microarray approach has been focused on transcriptional profiling of genes which are involved in systemic acquired resistance for cyanide. Our study shows that the change in different hormonal contents maintained almost the same pattern in roots and shoots upon CN exposure, except for SA. However, the hormone-related gene expression pattern conducted by microarray analysis was inconsistent in both plant materials (root/shoots). Comparison of gene expression between root/shoots showed a total of 29 in roots and 16 DEGs, respectively, indicating that hormone-related genes in roots were more responsive than those in shoots during exogenous CN metabolism. These results showed a remarkable change at transcript level of plant hormone-related genes, including biosynthesis, degradation, induction, and signal transduction under cyanide stress.

Potential adverse outcome pathway (AOP) of silver nanoparticles mediated reproductive toxicity in zebrafish.

Recently, the augmented utilization of silver nanoparticles (AgNPs) resulted in increasingrates of its release to aquatic environment, which potentially caused adverse effects to aquatic organisms. Therefore, this study investigated - reproductive toxicity and associated potential adverse outcome pathway (AOP) in zebrafish after chronic exposure to AgNPs. To serve the purpose, three-month-old adult zebrafish were exposed to different concentrations (0, 10, 33 and 100 µg/L) of AgNPs for five weeks. Exposure to 33 and 100 µg/L of AgNPs significantly decreased the fecundity in female zebrafish, accompanied by increasing apoptotic cells in the ovarian and testicular tissue using TUNEL assay. Increasing tissue burdens of AgNPs and reactive oxygen species (ROS) production were also found in both ovary and testis after five-week exposure to AgNPs. To explore the mechanism of the apoptotic pathway, the transcription levels of various genes (bax, bcl-2, caspase-3, and caspase-9) associated with the mitochondrion-mediated apoptosis pathway were examined in zebrafish after exposure to AgNPs. The results showed that the expression patterns of all the investigated genes were altered to some extent. These findings demonstrated that AgNPs exposure caused oxidative stress, induced germ cells apoptosis via mitochondrial-dependent pathway, and ultimately impaired the reproduction in zebrafish.

Reproductive effects linked to DNA methylation in male zebrafish chronically exposed to environmentally relevant concentrations of di-(2-ethylhexyl) phthalate.

Di-(2-ethylhexyl) phthalate (DEHP) possesses the potential to interfere with the male reproductive endocrine system in mammals; however, its reproductive toxicity in male zebrafish and associated epigenetic studies have not been explored. In this study, three-month-old male zebrafish were exposed to environmentally relevant concentrations of DEHP (0, 10, 33 and 100 µg/L) for 3 months, and then the impact on the reproduction of males and the underlying mechanism were investigated. Histological testing showed that an exposure concentration of 100 µg/L DEHP significantly inhibited spermatogenesis, with an associated decline in capability to fertilize untreated oocytes. Electron microscopic examinations also revealed noticeable damage to the testicular ultrastructure at the 100 µg/L DEHP exposure level. In addition, exposure to 33 and 100 µg/L of DEHP resulted in a decline of circulating testosterone (T) and an increase in the level of 17ß-estradiol (E2), both of which were possibly derived from the downregulation of cyp17a1 and hsd17b3 genes and the upregulation of the cyp19a1a gene in the gonads. The DNA methylation statuses of these genes were altered within their promoter regions. A significant increase in global DNA methylation in both the male testes and their offspring larvae was observed at higher exposure concentration of DEHP. Our findings demonstrate that exposure to environmentally relevant concentrations of DEHP can damage the testes, disturbe the sex hormones production, and inhibite spermatogenesis, which ultimately impairs the reproduction of male zebrafish.

Identification and expression analysis of CYS-A1, CYS-C1, NIT4 genes in rice seedlings exposed to cyanide.

Involvement of genes (CYS-A1, CYS-C1 and NIT4) encoded with cysteine synthase, ß-cyanoalanine synthase, nitrilase and cyanide metabolisms are evident in Arabidopsis. In the present study, identifications of CYS-A1, CYS-C1 and NIT4, predictions of conserved motifs, and constructions of phylogenetic relationships, based on their amino acid sequences in rice, were conducted. In order to elucidate the transcriptional responses of these cyanide-degrading genes, two candidate homologues were selected for each gene to test their expression changes upon exposure to exogenous KCN in rice seedlings using RT-PCR. Results showed that all selected candidate homologous genes were differentially expressed at different exposure points in roots and shoots of rice seedlings, suggesting their distinct roles during cyanide assimilation. Both candidate homologues for CYS-A1 constantly exhibited more abundant transcripts in comparison to control. However, only one candidate homologue for CYS-C1 and NIT4 showed a remarkable up-regulation during KCN exposure. Analysis of both tissue and solution cyanide indicated that rice seedlings were quickly able to metabolize exogenous KCN with minor accumulation in plant tissues. In conclusion, significant up-regulation of CYS-A1 suggested that the endogenous pool of cysteine catalyzed by cysteine synthase does not restrict the conversion of exogenous KCN into cyanoalanine through the ß-cyanoalanine pathway. However, insufficient responses of the transcription level of NIT4 suggested that NIT enzyme may be a limiting factor for cyanoalanine assimilation by rice seedlings.

Graphene oxide nanosheets induce DNA damage and activate the base excision repair (BER) signaling pathway both in vitro and in vivo.

Graphene oxide (GO) has widespread concerns in the fields of biological sciences and medical applications. Currently, studies have reported that excessive GO exposure can cause cellular DNA damage through reactive oxygen species (ROS) generation. However, DNA damage mediated response of the base excision repair (BER) pathway due to GO exposure is not elucidated yet. Therefore, we exposed HEK293T cells and zebrafish embryos to different concentrations of GO for 24 h, and transcriptional profiles of BER pathway genes, DNA damage, and cell viability were analyzed both in vitro and in vivo. Moreover, the deformation of HEK293T cells before and after GO exposure was also investigated using atomic force microscopy (AFM) to identify the physical changes occurred in the cells' structure. CCK-8 and Comet assay revealed the significant decrease in cell viability and increase in DNA damage in HEK293T cells at higher GO doses (25 and 50 µg/mL). Among the investigated genetic markers in HEK293T cells, BER pathway genes (APEX1, OGG1, CREB1, UNG) were significantly up-regulated upon exposure to higher GO dose (50 µg/mL), however, low exposure concentration (5, 25 µg/mL) failed to induce significant genetic induction except for CREB1 at 25 µg/mL. Additionally, the viscosity of HEK293T cells decreased upon GO exposure. In zebrafish, the results of up-regulated gene expressions (apex1, ogg1, polb, creb1) were consistent with those in the HEK293T cells. Taken all together, the exposure to elevated GO concentration could cause DNA damage to HEK293T cells and zebrafish embryos; BER pathway could be proposed as the possible inner response mechanism.

A novel forward osmosis system in landfill leachate treatment for removing polycyclic aromatic hydrocarbons and for direct fertigation.

Landfill leachate (LL) is harmful to aquatic environment because it contains high concentrations of dissolved organic matter, inorganic components, heavy metals, and other xenobiotics. Thus, the remediation of LL is crucial for environmental conservation. Here, a potential application of the forward osmosis (FO) filtration process with ammonium bicarbonate (NH4HCO3) as a draw solution (DS) was investigated to remediate membrane bioreactor-treated LL (M-LL). After the leachate treatment, the toxicity and removal efficiencies of polycyclic aromatic hydrocarbons (PAHs) were evaluated using zebrafish and cultured human cells. The water recovery rate was improved using the current protocol up to 86.6% and 91.6% by both the pressure retarded osmosis (PRO) mode and the forward osmosis (FO) mode. Water flux increased with the increasing DS concentrations, but solution velocities decreased with the operation time. Toxicity tests revealed that the M-LL treated by NH4HCO3 had no toxic effect on zebrafish and human cells. Moreover, green fluorescent protein (GFP) expression in the transgenic zebrafish Tg(cyp1a:gfp) induced by PAHs was very weak compared to the effects induced by untreated M-LL. Since the diluted DS met local safety requirements of liquid fertilizer, it could be directly applied as the liquid fertilizer for fertigation. In conclusion, this novel FO system using NH4HCO3 as the DS provides a cheap and efficient protocol to effectively remove PAHs and other pollutants in LL, and the diluted DS can be directly applied to crops as a liquid fertilizer, indicating that this technique is effective and eco-friendly for the treatment of different types of LL.

The Effects of Disturbance on Hypothalamus-Pituitary-Thyroid (HPT) Axis in Zebrafish Larvae after Exposure to DEHP.

Di-(2-ethylhexyl) phthalate (DEHP) has the potential to disrupt the thyroid endocrine system, but the underlying mechanism is unknown. In this study, zebrafish (Danio rerio) embryos were exposed to different concentrations of DEHP (0, 40, 100, 200, 400 µg/L) from 2 to 168 hours post fertilization (hpf). Thyroid hormones (THs) levels and transcriptional profiling of key genes related to hypothalamus-pituitary-thyroid (HPT) axis were examined. The result of whole-body thyroxine (T4) and triiodothyronine (T3) indicated that the thyroid hormone homeostasis was disrupted by DEHP in the zebrafish larvae. After exposure to DEHP, the mRNA expressions of thyroid stimulating hormone (tshß) and corticotrophin releasing hormone (crh) genes were increased in a concentration dependent manner, respectively. The expression level of genes involved in thyroid development (nkx2.1 and pax8) and thyroid synthesis (sodium/iodide symporter, nis, thyroglobulin, tg) were also measured. The transcripts of nkx2.1 and tg were significantly increased after DEHP exposure, while those of nis and pax8 had no significant change. Down-regulation of uridinediphosphate-glucuronosyl-transferase (ugt1ab) and up-regulation of thyronine deiodinase (dio2) might change the THs levels. In addition, the transcript of transthyretin (ttr) was up-regulated, while the mRNA levels of thyroid hormone receptors (trα and trß) remained unchanged. All the results demonstrated that exposure to DEHP altered the whole-body thyroid hormones in the zebrafish larvae and changed the expression profiling of key genes related to HPT axis, proving that DEHP induced the thyroid endocrine toxicity and potentially affected the synthesis, regulation and action of thyroid hormones.