Yuee Zhi

The role of NOS-mediated ROS accumulation in an early phase Cu-induced acute cytotoxicity in MCF-7 cells

Copper (Cu) ion is essential for the biological systems, however, high level of CuCl2 exposure causes detrimental effects, which leads to cell apoptosis. Nitric oxide (NO) is an efficient cell signal messenger, which plays an important role in cell apoptosis. However, the potential mechanism of an early phase Cu-induced acute cytotoxicity through the nitric oxide synthase (NOS) signaling pathway and its interaction has not been studied. In this report, we provide data showing that high level of CuCl2 could rapidly decrease the NO production with the release of Ca2+ and Zn2+, and then modulate the transcriptional and translational expression of NOSs in MCF-7 cells. The reactive oxygen species (ROS) level in cells was increased after high level of CuCl2 exposure, which led to the alpha subunit of eukaryotic initiation factor 2 phosphorylation. By using the free radical scavenger N-acetyl-l-cysteine or the NOS substrate l-arginine, it demonstrated that NOS played a critical role on the Cu-induced ROS generation, which further led to the oxidative stress and cell apoptosis. These results suggested that Cu-induced apoptosis was associated with the oxidative stress, and through the NOS-mediated signaling pathway.

Lignocellulose degradation by the isolate of Streptomyces griseorubens JSD-1

Streptomyces griseorubens JSD-1 is an isolate that can utilize lignocellulose (straw) as its sole carbon source for growth, and these lignocellulolytic genes involved in this biotransformation are expected to be crucial. However, little is known about the genetic basis related to this process. To further investigate the lignocellulose-degrading mechanisms, genome sequencing was carried out using MiSeq platform. After obtaining its draft genome, the key lignocellulolytic genes such as multicopper oxidase, exo-1, 4-β-glucanase, endo-1,4-β-glucanase, and β-xylosidase were identified and characterized. Multiple sequence alignments were performed to find out the identities of these analyzed proteins to those of their similar species. Signal peptide cleavage sites were predicted by SignalP 4.1 to reflect the cellular localization of their mature proteins. Besides, predicted 3D structures of these proteins were modeled by Phyre2, which showed to be highly identical to the templates in the PDB database. Finally, quantitative real-time PCR revealed that expression levels of all analyzed enzymes were significantly and generally up-regulated during the whole cultivation, indicating that they all contributed to the biodegradation.

The identification of the nitrate assimilation related genes in the novel Bacillus megaterium NCT-2 accounts for its ability to use nitrate as its only source of nitrogen.

Bacillus megaterium NCT-2 is a novel bacterium that can utilize nitrate as its only nitrogen source for growth. The nitrate assimilation related genes that are involved in this process would be expected to be crucial. However, little is known about the genomic background of this bacterium, let alone the sequences of the nitrate assimilation related genes. In order to further investigate the nitrate assimilation function of the NCT-2, genome sequencing was performed. After obtaining the fine map of the NCT-2 genome, which was submitted to the NCBI GenBank (AHTF00000000), the sequences of the nitrate assimilation related genes (the nitrate reductase electron transfer subunit nasB and the nitrate reductase catalytic subunit nasC, the nitrite reductase [NAD(P)H] large subunit nasD and the nitrite reductase [NAD(P)H] small subunit nasE, and the glutamine synthetase glnA) were identified. Multiple alignments were performed to find out the sequence identities of the nitrate assimilation related genes to that of their similar species. Through KEGG signaling mapping search, the nitrate assimilation related genes were revealed to be located in the nitrogen metabolism signaling pathway. The putative 3D protein structures of these genes were modeled by SWISS MODEL, and shown to be highly similar to the nitrate assimilation related genes in the PDB database. Finally, the sequence validity of the nitrate assimilation related genes was verified by PCR with specifically designed primers.

Biological pretreatment of rice straw with Streptomyces griseorubens JSD-1 and its optimized production of cellulase and xylanase for improved enzymatic saccharification efficiency.

ABSTRACT Biological pretreatment of rice straw and production of reducing sugars by hydrolysis of bio-pretreated material with Streptomyces griseorubens JSD-1 was investigated. After 10 days of incubation, various chemical compositions of inoculated rice straw were degraded and used for further enzymatic hydrolysis studies. The production of cellulolytic enzyme by S. griseorubens JSD-1 favored the conversion of cellulose to reducing sugars. The culture medium for cellulolytic enzyme production by using agro-industrial wastes was optimized through response surface methodology. According to the response surface analysis, the concentrations of 11.13, 20.34, 4.61, and 2.85 g L−1 for rice straw, wheat bran, peptone, and CaCO3, respectively, were found to be optimum for cellulase and xylanase production. Then the hydrolyzed spent Streptomyces cells were used as a nitrogen source and the maximum filter paper cellulase, carboxymethylcellulase, and xylanase activities of 25.79, 78.91, and 269.53 U mL−1 were achieved. The crude cellulase produced by S. griseorubens JSD-1 was subsequently used for the hydrolysis of bio-pretreated rice straw, and the optimum saccharification efficiency of 88.13% was obtained, indicating that the crude enzyme might be used instead of commercial cellulase during a saccharification process. These results give a basis for further study of bioethanol production from agricultural cellulosic waste.

The role of nitric oxide synthase signaling pathway in the Zn-induced cellular responses in MCF-7 cells

Trace amount zinc plays key roles in biological systems, while in excessive amount it causes toxic effects. Evidence shows that there exists a crosstalk between NO and Zn apoptotic signal transduction pathway. However, the potential mechanism of Zn-induced cellular responses through the NOS signaling pathway has not been determined yet. In this research, trace amount ZnCl2 (1nM) could induce the NO production, however it appears that this influence does not extend to genetic level in MCF-7 cells. Whereas, excess ZnCl2 (100μM, 1mM) could lead to a decreased NO production first with the release of Ca(2+), and then induce the NO production with the transcriptional and translational activation of NOSs. The ROS generation was also induced by excess ZnCl2, causing the elF2α phosphorylation. The alleviation effect of N-acetyl-l-cysteine or l-arginine on the Zn-induced ROS generation and apoptosis suggested that Zn-induced apoptosis was associated with the NOS-mediated oxidative stress.

Three new flavonol triglycosides from Derris trifoliata

Three new flavonol triglycosides, kaempferol-3-O-α-l-rhamnopyranosyl-(1 → 6)-β-d-glucopyranosyl-(1 → 3)-β-d-glucopyranoside (1), quercetin-3-O-α-l-rhamnopyranosyl-(1 → 6)-β-d-glucopyranosyl-(1 → 3)-β-d-glucopyranoside (2), quercetin-3-O-α-l-rhamnopyranosyl-(1 → 6)-β-d-glucopyranosyl-(1 → 2)-β-d-glucopyranoside (3), together with the two known flavonol glycosides, kaempferol-3-O-α-l-rhamnopyranosyl-(1 → 6)-β-d-glucopyranoside and kaempferol-3-O-α-l-rhamnopyranosyl-(1 → 6)-β-d-glucopyranosyl-(1 → 2)-β-d-glucopyranoside, were isolated from the aerial parts of Derris trifoliata. Their structures were determined on the basis of chemical and spectroscopic analyses.

Non-Point Source Pollution Characteristics of Agriculture-Derived Nitrogen in Groundwater in Suburban Area of Shanghai Based on Models

This paper analyzes the characteristics of the NPS nitrogen pollution of the groundwater in Guoyuan Village, Pudong District, Shanghai. And the associated effects on the surface and groundwater around the study area are discussed in detail based on the successive observed data, DNDC and L-THIA model. The results show that both of the surface and groundwater are polluted so seriously that they are not suitable to drink. The average content of total nitrogen in surface water is 6.3 mg/L and 16.85 mg/L in the groundwater, and both of them attribute to Grade V surface water standard (≤2.0 mg/L) according to the national standard(GB 3838-2002). It is concluded that the nitrogen pollution comes mainly from the fertilizer of the peach orchard based on the further modeling analysis. Therefore, reasonable adjustment of fertilization measures and project may be an effective and practical approach to control the nitrogen pollution around the peach orchard.

Expression and characterization of a novel endo-1,4-β-xylanase produced by Streptomyces griseorubens JSD-1 isolated from compost-treated soil

Streptomyces griseorubens JSD-1 is an isolate sourced from compost-treated soil that can utilize xylan as the unique carbon source for growth; xylanlytic genes related to this process were expected to be crucial. However, little is known about the genetic basic of utilizing xylan in this isolate. To further investigate the molecular mechanism of xylan metabolism, genome sequencing was carried out. The draft genome obtained has been deposited with GenBank. Endo-1,4-β-xylanase (Xyn) was acquired, followed by characterization of its cellular localization and expression profiles. Xyn proved to be a novel enzyme, differing from xylanases from similar species according to combined analyses from multiple sequence alignments and putative three-dimensional structure modeling. Moreover, recombinant expression was optimized in heterogeneous hosts to further determine optimum pH and temperature conditions as well as residual activity in the presence of metal ions and inhibitors. Finally, substrate specificity and kinetic parameters were also studied. In conclusion, our findings illustrate the genetic background related to xylan metabolism in this isolate. In addition, a novel Xyn that is expected to have potential in the bio-degradation of hemicellulosic materials was expressed, purified and characterized.

The cytotoxic effect of the NOS-mediated oxidative stress in MCF-7 cells after PbCl2 exposure

The potential Pb‐induced cytotoxicity in various tissues and biological systems has been reported. Some evidences also indicate that the Pb‐caused cytotoxicity may be associated with the nitric oxide synthase (NOS). However, there remains uncertainty about the role of the NOS signaling pathway during the Pb‐induced cytotoxicity. In this report, we provide data showing that PbCl2 treatment depresses the expressions of the three distinct NOS isoforms: neuronal nitric oxide synthase (nNOS), endothelial NOS (eNOS), and inducible NOS (iNOS) on both transcriptional and translational levels in MCF‐7 cells. The down‐regulation of NOSs expressions by PbCl2 exposure leads to reduced NOS activity and nitric oxide (NO) production. Meanwhile, the intracellular reactive oxygen species (ROS) level is elevated after PbCl2 exposure, which leads to the alpha subunit of eukaryotic initiation factor 2 (elF2α) phosphorylation. The reduction effects of the free radical scavenger N‐acetyl‐l‐cysteine or the NOS substrate l‐arginine on the Pb‐induced ROS generation suggest that the NOS signaling pathway plays a key role in the Pb‐induced oxidative stress, which further results in the elF2α phosphorylation and cytotoxicity.

Transformation of Nitrate Nitrogen by Three Strains of Bacteria Isolated from Facility Culturing Soils

Three bacterial strains capable of transforming nitrate nitrogen designated as NCT-2, NCT-3 and NCT-4, were isolated from excessive nitrate nitrogen salinization soils. According to the sequence analysis of the partial 16s rRNA gene, it is found that strains NCT-2 and NCT-4 belong to Bacillus sp., and strain NCT-3 belongs to Stenotrophomonas sp. The effects of nitrate nitrogen concentration, pH and temperature on transformation were examined. At a concentration of 50.0 mg L-1, More than 93.3% of nitrate nitrogen was transformed by Bacillus sp. NCT-2 after 5d and 72.3% by Stenotrophomonas sp. NCT-3 after 5d. At a concentration of 400.0 mg L-1, the transformation rate was inhibited. Only 40.3% of nitrate nitrogen can be transformed by Bacillus sp. NCT-4 after 5d. More than 63.3% transformation rate was observed with Stenotrophomonas sp. NCT-3 after 5d. The growth of these three strains was inhibited at high pH buffers. The abiotic transformation was basically the same at different pH conditions. The transformation rate of nitrate nitrogen at 25°C was faster than those at 20°C and 35°C by these strains, except the highest degradation rate of Bacillus sp. NCT-4 was obtained at 35°C.