Potential Role of Epigenetic Mechanism in Manganese Induced Neurotoxicity

Manganese is a vital nutrient and is maintained at an optimal level (2.5–5 mg/day) in human body. Chronic exposure to manganese is associated with neurotoxicity and correlated with the development of various neurological disorders such as Parkinson’s disease. Oxidative stress mediated apoptotic cell death has been well established mechanism in manganese induced toxicity. Oxidative stress has a potential to alter the epigenetic mechanism of gene regulation. Epigenetic insight of manganese neurotoxicity in context of its correlation with the development of parkinsonism is poorly understood. Parkinson’s disease is characterized by the �-synuclein aggregation in the form of Lewy bodies in neuronal cells. Recent fndings illustrate that manganese can cause overexpression of �-synuclein. �-Synuclein acts epigenetically via interaction with histone proteins in regulating apoptosis. �-Synuclein also causes global DNA hypomethylation through sequestration of DNA methyltransferase in cytoplasm. An individual genetic difference may also have an influence on epigenetic susceptibility to manganese neurotoxicity and the development of Parkinson’s disease. Tis review presents the current state of fndings in relation to role of epigenetic mechanism in manganese induced neurotoxicity, with a special emphasis on the development of Parkinson’s disease

Global DNA methylation profiling of manganese-exposed human neuroblastoma SH-SY5Y cells reveals epigenetic alterations in Parkinson’s disease-associated genes

Manganese (Mn) is an essential trace element required for optimal functioning of cellular biochemical pathways in the central nervous system. Elevated exposure to Mn through environmental and occupational exposure can cause neurotoxic effects resulting in manganism, a condition with clinical symptoms identical to idiopathic Parkinson’s disease. Epigenetics is now recognized as a biological mechanism involved in the etiology of various diseases. Here, we investigated the role of DNA methylation alterations induced by chronic Mn (100 µM) exposure in human neuroblastoma (SH-SY5Y) cells in relevance to Parkinson’s disease. A combined analysis of DNA methylation and gene expression data for Parkinson’s disease-associated genes was carried out. Whole-genome bisulfite conversion and sequencing indicate epigenetic perturbation of key genes involved in biological processes associated with neuronal cell health. Integration of DNA methylation data with gene expression reveals epigenetic alterations to PINK1, PARK2 and TH genes that play critical roles in the onset of Parkinsonism. The present study suggests that Mn-induced alteration of DNA methylation of PINK1–PARK2 may influence mitochondrial function and promote Parkinsonism. Our findings provide a basis to further explore and validate the epigenetic basis of Mn-induced neurotoxicity

Manganese exposure: linking down-regulation of miRNA-7 and miRNA-433 with α-synuclein overexpression and risk of idiopathic Parkinson's disease

Manganese is an essential trace element however elevated environmental and occupational exposure to this element has been correlated with neurotoxicity symptoms clinically identical to idiopathic Parkinson's disease. In the present study we chronically exposed human neuroblastoma SH-SY5Y cells to manganese (100 μM) and carried out expression profiling of miRNAs known to modulate neuronal differentiation and neurodegeneration. The miRNA PCR array results reveal alterations in expression levels of miRNAs, which have previously been associated with the regulation of synaptic transmission and apoptosis. The expressions of miR-7 and miR-433 significantly reduced upon manganese exposure. By in silico homology analysis we identified SNCA and FGF-20as targets of miR-7 and miR-433. We demonstrate an inverse correlation in expression levels where reduction in these two miRNAs causes increases in SNCA and FGF-20. Transient transfection of SH-SY5Y cells with miR-7 and miR-433 mimics resulted in down regulation of SNCA and FGF-20 mRNA levels. Our study is the first to uncover the potential link between manganese exposure, altered miRNA expression and parkinsonism: manganese exposure causes overexpression of SNCA and FGF-20 by diminishing miR-7 and miR-433 levels. These miRNAs may be considered critical for protection from manganese induced neurotoxic mechanism and hence as potential therapeutic targets

Characterization and optimization of production of exopolysaccharide from Chlamydomonas reinhardtii.

Chlamydomonas reinhardtii strain RAC was isolated based on its ability to secrete large amount of exopolysaccharide (EPS). The purified EPS had a molecular weight of 2.25�10(5)Da, and showed fibrillar structure with surfaces having sheet-like appearance. Chemical analysis showed the presence of galacturonic acid, ribose, arabinose, xylose, glucose, galactose and rhamnose sugars. The production of EPS was optimized by the classical one-at-a-time approach and Plackett-Burman design, followed by response surface methodology. The resulting response surface model was statistically significant (p<0.5) and predicted maximum EPS production of 628 mg/L. The optimum production medium consisted of CaCl2 - 74, NaNO3 - 422, K2HPO4 - 10 and MgSO4 - 200mg/L with a pH 7. The EPS showed significant antioxidant activity, which can have several industrial applications. This is the first report on characterization and production of EPS from a Chlamydomonas strain isolated from India. Its differences from the earlier reported EPS are discussed

Diversity and metabolic potential of culturable root-associated bacteria from Origanum vulgare in sub-Himalayan region.

Study of rhizospheric bacteria from important plants is very essential, as they are known to influence plant growth and productivity, and also produce industrially important metabolites. Origanum vulgare is a perennial medicinal aromatic plant rich in phenolic antioxidants. Present study investigates the diversity of culturable root-associated bacteria from this plant in Palampur, India, which constitutes a unique ecosystem due to high rain fall, wide temperature fluctuations and acidic soil. Both root endophytes and rhizospheric soil bacteria were isolated, which resulted in a total of 120 morphologically different isolates. They were found to group into 21 phylotypes based on restriction fragment length polymorphism analysis. Growth medium composition had significant effect on the diversity of the isolated bacterial populations. The isolates were characterized for various metabolic, plant growth promoting (PGP) and other biotechnologically useful activities, based on which they were clustered into groups by principal component analysis. Majority of the isolates belonged to γ-Proteobacteria and Firmicutes. Pseudomonas and Stenotrophomonas were the most dominant species and together constituted 27.5 % of the total isolates. Many isolates, especially γ-Proteobacteria, showed very high PGP activities. Few isolates exhibited very high antioxidant activity, which may find potential applications in food and health industries. Firmicutes were catabolically the most versatile group and produced several hydrolytic enzymes. To the best of our knowledge, it is the first study describing rhizospheric microbial community of O. vulgare

Lateral gene transfer in phylogeny of azoreductase enzyme.

This paper attempts to reconstruct the phylogeny of azoreductase enzyme from different organisms and compare it with the small subunit rRNA-based phylogeny of the organisms. The two phylogenies were found to be incongruent, indicating several events of lateral transfer of azoreductase gene between phylogenetically diverse organisms. However, the phylogenetic analysis methods have several limitations and a single method may not give the true pattern. Hence, it is necessary to corroborate the results with other complementary analysis tools. We used several tools to test our hypothesis of lateral transfer and found that it was supported not only by the analysis of the whole sequences, but also by the conserved motifs detected in these sequences. There were ample evidences for lateral transfer of azoreductase gene among enteric bacteria. There were also indications that azoreductase probably evolved in prokaryotes and then it was laterally transferred to eukaryotes in multiple events, resulting in some sequence variation among eukaryotic azoreductases. Finally, profile HMMs and conserved motifs extracted from these azoreductase sequences were found to provide sensitive tools for identifying potential azoreductases from the database. The analysis techniques used in this study can be extended to other gene trees to verify their evolutionary histories

Mercuric reductase activity of multiple heavy metal-resistant Lysinibacillus sphaericus G1

A culture was isolated from an industrial mercuric salt-contaminated soil, which could tolerate Cd, Co, Zn, Cr, and Hg up to 190, 525, 350, 935, and 370 μM, respectively. The isolate was identified as Lysinibacillus sphaericus by 16S rRNA gene sequencing. It bioaccumulated Cd, Co, and Zn, and reductively detoxified Cr and Hg. Chromate reductase and mercuric reductase (MerA) activities in the cell extract were 2.4 and 0.13 units mg-1 protein, respectively. The study also describes designing of broad-specificity primers based on firmicute merA genes. These primers were successfully used to amplify a 440 bp merA fragment from the current isolate. Based on the partial sequence, complete merA ORF of 1641 bp was amplified. It showed 99% similarity to a putative merA gene from distantly related Streptococcus agalactiae, but only 72% identity with the well-characterized merA from a more closely related Bacillus cereus RC607. The gene sequence possessed all the features required for the functioning of MerA enzyme, and its function was confirmed by recombinant expression in E. coli. To the best of our knowledge, this is the first report of full length merA gene from L. sphaericus

Functional and phylogenetic diversity of root-associated bacteria of Ajuga bracteosa in Kangra valley.

Present study investigates the cultivable diversity of root-associated bacteria from a medicinal plant Ajuga bracteosa in the Kangra valley, in order to determine their plant growth promoting (PGP) and biotechnological potential. The plant was found to exhibit a positive rhizosphere effect of 1.3-1.5. A total of 123 morphologically different bacteria were isolated from the rhizospheric soil and roots of the plant. Medium composition was found to have significant effect on the composition of isolated bacterial populations. Majority of the rhizospheric soil isolates belonged to α- and γ-Proteobacteria, with Pseudomonas constituting the most dominant species. Endophytic bacterial community, on other hand, consisted almost exclusively of Firmicutes. Majority of the isolates showed PGP activity by producing siderophores and indole acetic acid. Several isolates were found to exhibit very high antioxidant activity in the culture medium. A significant proportion of isolates also demonstrated other ecologically important activities like phosphate solubilization, nitrogen fixation, and production of hydrolytic enzymes including amylase, protease, lipase, chitinase, cellulase, pectinase and phosphatase. Firmicutes were found to be metabolically the most versatile group and performed multiple enzyme activities. This is the first systematic study of culturable bacterial community from the rhizosphere of A. bracteosa, particularly in the Kangra valley region

Structural and functional characterization of mercuric reductase from Lysinibacillus sphaericus strain G1

In response to the widespread presence of inorganic Hg in the environment, bacteria have evolved resistance systems with mercuric reductase (MerA) as the key enzyme. MerA enzymes have still not been well characterized from gram positive bacteria. Current study reports physico-chemical, kinetic and structural characterization of MerA from a multiple heavy metal resistant strain of Lysinibacillus sphaericus, and discusses its implications in bioremediation application. The enzyme was homodimeric with subunit molecular weight of about 60 kDa. The K-m and V-max were found to be 32 A mu M of HgCl2 and 18 units/mg respectively. The enzyme activity was enhanced by beta-mercaptoethanol and NaCl up to concentrations of 500 A mu M and 100 mM respectively, followed by inhibition at higher concentrations. The enzyme showed maximum activity in the pH range of 7-7.5 and temperature range of 25-50 degrees C, with melting temperature of 67 degrees C. Cu2+ exhibited pronounced inhibition of the enzyme with mixed inhibition pattern. The enzyme contained FAD as the prosthetic group and used NADPH as the preferred electron donor, but it showed slight activity with NADH as well. Structural characterization was carried out by circular dichroism spectrophotometry and X-ray crystallography. X-ray confirmed the homodimeric structure of enzyme and gave an insight on the residues involved in catalytic binding. In conclusion, the investigated enzyme showed higher catalytic efficiency, temperature stability and salt tolerance as compared to MerA enzymes from other mesophiles. Therefore, it is proposed to be a promising candidate for Hg2+ bioremediation