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      Identification of Glutathione S-Transferase (GST) Genes from a Dark Septate Endophytic Fungus ( Exophiala pisciphila) and Their Expression Patterns under Varied Metals Stress

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          Abstract

          Glutathione S-transferases (GSTs) compose a family of multifunctional enzymes that play important roles in the detoxification of xenobiotics and the oxidative stress response. In the present study, twenty four GST genes from the transcriptome of a metal-tolerant dark septate endophyte (DSE), Exophiala pisciphila, were identified based on sequence homology, and their responses to various heavy metal exposures were also analyzed. Phylogenetic analysis showed that the 24 GST genes from E. pisciphila ( EpGSTs) were divided into eight distinct classes, including seven cytosolic classes and one mitochondrial metaxin 1-like class. Moreover, the variable expression patterns of these EpGSTs were observed under different heavy metal stresses at their effective concentrations for inhibiting growth by 50% (EC 50). Lead (Pb) exposure caused the up-regulation of all EpGSTs, while cadmium (Cd), copper (Cu) and zinc (Zn) treatments led to the significant up-regulation of most of the EpGSTs ( p < 0.05 to p < 0.001). Furthermore, although heavy metal-specific differences in performance were observed under various heavy metals in Escherichia coli BL21 (DE3) transformed with EpGSTN-31, the over-expression of this gene was able to enhance the heavy metal tolerance of the host cells. These results indicate that E. Pisciphila harbored a diverse of GST genes and the up-regulated EpGSTs are closely related to the heavy metal tolerance of E. pisciphila. The study represents the first investigation of the GST family in E. pisciphila and provides a primary interpretation of heavy metal detoxification for E. pisciphila.

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          Most cited references31

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          Glutathione transferases.

          This review describes the three mammalian glutathione transferase (GST) families, namely cytosolic, mitochondrial, and microsomal GST, the latter now designated MAPEG. Besides detoxifying electrophilic xenobiotics, such as chemical carcinogens, environmental pollutants, and antitumor agents, these transferases inactivate endogenous alpha,beta-unsaturated aldehydes, quinones, epoxides, and hydroperoxides formed as secondary metabolites during oxidative stress. These enzymes are also intimately involved in the biosynthesis of leukotrienes, prostaglandins, testosterone, and progesterone, as well as the degradation of tyrosine. Among their substrates, GSTs conjugate the signaling molecules 15-deoxy-delta(12,14)-prostaglandin J2 (15d-PGJ2) and 4-hydroxynonenal with glutathione, and consequently they antagonize expression of genes trans-activated by the peroxisome proliferator-activated receptor gamma (PPARgamma) and nuclear factor-erythroid 2 p45-related factor 2 (Nrf2). Through metabolism of 15d-PGJ2, GST may enhance gene expression driven by nuclear factor-kappaB (NF-kappaB). Cytosolic human GST exhibit genetic polymorphisms and this variation can increase susceptibility to carcinogenesis and inflammatory disease. Polymorphisms in human MAPEG are associated with alterations in lung function and increased risk of myocardial infarction and stroke. Targeted disruption of murine genes has demonstrated that cytosolic GST isoenzymes are broadly cytoprotective, whereas MAPEG proteins have proinflammatory activities. Furthermore, knockout of mouse GSTA4 and GSTZ1 leads to overexpression of transferases in the Alpha, Mu, and Pi classes, an observation suggesting they are part of an adaptive mechanism that responds to endogenous chemical cues such as 4-hydroxynonenal and tyrosine degradation products. Consistent with this hypothesis, the promoters of cytosolic GST and MAPEG genes contain antioxidant response elements through which they are transcriptionally activated during exposure to Michael reaction acceptors and oxidative stress.
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            Structure, function and evolution of glutathione transferases: implications for classification of non-mammalian members of an ancient enzyme superfamily.

            The glutathione transferases (GSTs; also known as glutathione S-transferases) are major phase II detoxification enzymes found mainly in the cytosol. In addition to their role in catalysing the conjugation of electrophilic substrates to glutathione (GSH), these enzymes also carry out a range of other functions. They have peroxidase and isomerase activities, they can inhibit the Jun N-terminal kinase (thus protecting cells against H(2)O(2)-induced cell death), and they are able to bind non-catalytically a wide range of endogenous and exogenous ligands. Cytosolic GSTs of mammals have been particularly well characterized, and were originally classified into Alpha, Mu, Pi and Theta classes on the basis of a combination of criteria such as substrate/inhibitor specificity, primary and tertiary structure similarities and immunological identity. Non-mammalian GSTs have been much less well characterized, but have provided a disproportionately large number of three-dimensional structures, thus extending our structure-function knowledge of the superfamily as a whole. Moreover, several novel classes identified in non-mammalian species have been subsequently identified in mammals, sometimes carrying out functions not previously associated with GSTs. These studies have revealed that the GSTs comprise a widespread and highly versatile superfamily which show similarities to non-GST stress-related proteins. Independent classification systems have arisen for groups of organisms such as plants and insects. This review surveys the classification of GSTs in non-mammalian sources, such as bacteria, fungi, plants, insects and helminths, and attempts to relate them to the more mainstream classification system for mammalian enzymes. The implications of this classification with regard to the evolution of GSTs are discussed.
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              Extensive functional diversification of the Populus glutathione S-transferase supergene family.

              Identifying how genes and their functions evolve after duplication is central to understanding gene family radiation. In this study, we systematically examined the functional diversification of the glutathione S-transferase (GST) gene family in Populus trichocarpa by integrating phylogeny, expression, substrate specificity, and enzyme kinetic data. GSTs are ubiquitous proteins in plants that play important roles in stress tolerance and detoxification metabolism. Genome annotation identified 81 GST genes in Populus that were divided into eight classes with distinct divergence in their evolutionary rate, gene structure, expression responses to abiotic stressors, and enzymatic properties of encoded proteins. In addition, when all the functional parameters were examined, clear divergence was observed within tandem clusters and between paralogous gene pairs, suggesting that subfunctionalization has taken place among duplicate genes. The two domains of GST proteins appear to have evolved under differential selective pressures. The C-terminal domain seems to have been subject to more relaxed functional constraints or divergent directional selection, which may have allowed rapid changes in substrate specificity, affinity, and activity, while maintaining the primary function of the enzyme. Our findings shed light on mechanisms that facilitate the retention of duplicate genes, which can result in a large gene family with a broad substrate spectrum and a wide range of reactivity toward different substrates.
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                Author and article information

                Contributors
                Role: Academic Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                17 April 2015
                2015
                : 10
                : 4
                : e0123418
                Affiliations
                [1 ]Key Laboratory of Conservation and Utilization for Bioresources and Key Laboratory of Microbial Diversity in Southwest China, Ministry of Education, Yunnan University, Kunming, Yunnan, China
                [2 ]School of Agriculture, Yunnan University, Kunming, Yunnan, China
                [3 ]School of Life Science, Yunnan University, Kunming, Yunnan, China
                CSIR-National Botanical Research Institute, INDIA
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Conceived and designed the experiments: ZWZ DKZ. Performed the experiments: MS LL JLW GHC TL. Analyzed the data: MS DKZ QQ. Contributed reagents/materials/analysis tools: QQ TL. Wrote the paper: MS DKZ ZWZ.

                [¤]

                Current address: Cuihu North Road 2#, Key Laboratory of Conservation and Utilization for Bioresources, Yunnan University, Kunming, Yunnan, China

                Article
                PONE-D-14-51781
                10.1371/journal.pone.0123418
                4401685
                25884726
                8a156ce7-f752-4fa6-a873-8e435c57451a
                Copyright @ 2015

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited

                History
                : 26 November 2014
                : 18 February 2015
                Page count
                Figures: 3, Tables: 1, Pages: 13
                Funding
                This work was supported by the National Natural Science Foundation of China (31160009, ZWZ; 31460114, ZWZ; 41161083, TL;41461073, TL, http://npd.nsfc.gov.cn/fundingProjectSearchAction.action!search.action). The funders had the role in study design, data collection and analysis, and decision to publish. This work was also supported by the Key Project of Applied Basic Research of Yunnan(2013FA001, ZWZ)the Postdoctoral Sustentation Fund supported through Yunnan University and the Young Academic and Technical Leader Raising Foundation of Yunnan Province (2012HB006,D KZ), who had the role in preparation of the manuscript.
                Categories
                Research Article
                Custom metadata
                All GST genes listed in the manuscript are available from the database of NCBI. Accession numbers are listed in supporting information file S2 Table.

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                Uncategorized

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