The Experts below are selected from a list of 1701 Experts worldwide ranked by ideXlab platform
Qiwei Qin - One of the best experts on this subject based on the ideXlab platform.
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identification of major capsid protein as a potential biomarker of grouper Iridovirus infected cells using aptamers selected by selex
2019Co-Authors: Mingzhu Liu, Xiaohong Huang, Hehe Xiao, Shina Wei, Qiwei QinAbstract:Biomarkers have important roles in disease pathogenesis, and serve as important disease indicators for developing novel diagnostic and therapeutic approaches. Grouper Iridovirus is a nucleocytoplasmic DNA virus, which not only causes great economic losses in mariculture but also seriously threatens the global biodiversity. However, a lack of biomarkers has limited the progress in clarifying Iridovirus pathogenesis. Here, we report novel molecular probes, aptamers, for specific identification of biomarkers in grouper Iridovirus-infected cells. Aptamers are selected by SELEX, which is a completely different approach from conventional antibody-based methods for biomarkers discovery. Aptamer-based technology is the unique efficient selection for cell-specific target molecules, and helps find out new biomarkers without the knowledge of characteristics of proteins expressed on virus-infected cell surface. With the implementation of a two-step strategy (aptamer selection and biomarker discovery), combined with mass spectrometry, grouper Iridovirus major capsid protein was ultimately identified as a potential biomarker of aptamer Q5 for grouper Iridovirus infection. The specific interactions of aptamer Q5 and MCP were experimentally validated by several assays, including EMSA, co-localization of fluorescence by LSCM, binding competition tests, and siRNA silencing tests by flow cytometry. This aptamer-based method for biomarkers discovery developed with grouper Iridovirus-infected cells could be applicable to other types of virus infection, markedly improve our studies of biomarker discovery and virus pathogenesis, and further facilitate the development of diagnostic tools and therapeutic approaches to treat virus infection.
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soft shelled turtle Iridovirus enters cells via cholesterol dependent clathrin mediated endocytosis as well as macropinocytosis
2018Co-Authors: Youhua Huang, Shaowen Wang, Xiaohong Huang, Qiwei QinAbstract:Ranaviruses are nucleoplasmic large DNA viruses that can cause major economic losses in the aquaculture industry and pose a severe threat to global ecological diversity. The available literature demonstrates that classifiable members of the genus Ranavirus enter cells via multiple and complicated routes. Here, we demonstrated the underlying cellular entry mechanism of soft-shelled turtle Iridovirus (STIV) using green fluorescence tagged recombinant virus. Treatment with chlorpromazine, sucrose, ethyl-isopropyl amiloride, chloroquine or bafilomycin A1 all significantly decreased STIV infection, suggesting that STIV uses clathrin-mediated endocytosis and macropinocytosis to enter cells via a pH-dependent pathway. Depletion of cellular cholesterol with methyl-β-cyclodextrin significantly inhibited STIV entry, but neither filipin III nor nystatin did, suggesting that STIV entry was cholesterol dependent but caveola independent. Treatment with dynasore, genistein, ML-7 or cytochalasin D all significantly inhibited STIV infection, indicating that Rac GTPase and myosin II activity were required for the macropinocytosis-like pathway as well as actin polymerization. Our findings suggest that the molecular events involved in STIV entry are not identical to those of other ranavirus isolates. Our results also extend our understanding of the molecular mechanism of Iridovirus entry and pathogenesis.
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development and characterization of aptamer based enzyme linked apta sorbent assay for the detection of singapore grouper Iridovirus infection
2016Co-Authors: Lingli Zhou, Qiwei Qin, Shina Wei, Jingguang Wei, Min YangAbstract:AbstractAims:Singapore grouper Iridovirus (SGIV) is a devastating aquaculture virus responsible for heavy economic losses to grouper, Epinephelus sp. aquaculture. The aim of this study was to develop a rapid and sensitive detection method for SGIV infections in infected groupers.Methods and Results:
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antiviral role of grouper sting against Iridovirus infection
2015Co-Authors: Youhua Huang, Xiaohong Huang, Zhengliang Ouyang, Shina Wei, Jingguang Wei, Wei Wang, Sheng Zhou, Qiwei QinAbstract:Stimulator of interferon genes (STING, also known as MITA, ERIS, MPYS or TMEM173) has been identified as a central component in the innate immune response to cytosolic DNA and RNA derived from different pathogens. However, the detailed role of STING during fish Iridovirus infection still remained largely unknown. Here, the STING homolog from grouper Epinephelus coioides (EcSTING) was cloned and its effects on IFN response and antiviral activity were investigated. The full-length EcSTING cDNA was composed of 1590 bp and encoded a polypeptide of 409 amino acids with 80% identity to STING homolog from large yellow croaker. Amino acid alignment analysis indicated that EcSTING contained 4 predicated transmembrane motifs (TMs) in the N terminal, and a C-terminal domain (CTD) which consisted of a dimerization domain (DD), c-di-GMP-binding domain (CBD) and a C-terminal tail (CTT). Expression profile analysis revealed that EcSTING was abundant in gill, spleen, brain, skin, and liver. Upon different stimuli in vivo, the EcSTING transcript was dramatically up-regulated after challenging with Singapore grouper Iridovirus (SGIV), lipopolysaccharide (LPS) and polyinosin-polycytidylic acid (poly I:C). Reporter gene assay showed that EcSTING activated ISRE, zebrafish type I IFN and type III IFN promoter in vitro. Mutant analysis showed that IFN promoter activity was mostly mediated by the phosphorylation sites at serine residue S379 and S387. Moreover, EcSTING induced type I and III IFN promoter activity could be impaired by overexpression of EcIRF3-DN or EcIRF7-DN, suggesting that EcSTING mediated IFN response in IRF3/IRF7 dependent manner. In addition, the cytopathic effect (CPE) progression of SGIV infection and viral protein synthesis was significantly inhibited by overexpression of EcSTING, and the inhibitory effect was abolished in serine residue S379 and S387 mutant transfected cells. Together, our results demonstrated that EcSTING might be an important regulator of grouper innate immune response against Iridovirus infection.
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entry of a novel marine dna virus singapore grouper Iridovirus into host cells occurs via clathrin mediated endocytosis and macropinocytosis in a ph dependent manner
2014Co-Authors: Shaowen Wang, Youhua Huang, Xiaohong Huang, Xian Hao, Mingjun Cai, Hongda Wang, Qiwei QinAbstract:Iridoviruses are nucleocytoplasmic DNA viruses which cause great economic losses in the aquaculture industry but also show significant threat to global biodiversity. However, a lack of host cells has resulted in poor progress in clarifying Iridovirus behavior. We investigated the crucial events during virus entry using a combination of single-virus tracking and biochemical assays, based on the established virus-cell infection model for Singapore grouper Iridovirus (SGIV). SGIV infection in host cells was strongly inhibited when cells were pretreated with drugs blocking clathrin-mediated endocytosis, including sucrose and chlorpromazine. Inhibition of key regulators of macropinocytosis, including Na+/H+ exchanger, Rac1 GTPase, p21-activated kinase 1 (PAK1), protein kinase C (PKC), and myosin II, significantly reduced SGIV uptake. Cy5-labeled SGIV particles were observed to colocalize with clathrin and macropinosomes. In contrast, disruption of cellular cholesterol by methyl-β-cyclodextrin and nystatin had no effect on virus infection, suggesting that SGIV entered grouper cells via the clathrin-mediated endocytic pathway and macropinocytosis but not via caveola-dependent endocytosis. Furthermore, inhibitors of endosome acidification such as chloroquine and bafilomycin A1 blocked virus infection, indicating that SGIV entered cells in a pH-dependent manner. In addition, SGIV particles were observed to be transported along both microtubules and actin filaments, and intracellular SGIV motility was remarkably impaired by depolymerization of microtubules or actin filaments. The results of this study for the first time demonstrate that not only the clathrin-dependent pathway but also macropinocytosis are involved in fish DNA enveloped virus entry, thus providing a convenient tactic for exploring the life cycle of DNA viruses. IMPORTANCE Virus entry into host cells is critically important for initiating infections and is usually recognized as an ideal target for the design of antiviral strategies. Iridoviruses are large DNA viruses which cause serious threats to ecological diversity and the aquaculture industry worldwide. However, the current understanding of Iridovirus entry is limited and controversial. Singapore grouper Iridovirus (SGIV) is a novel marine fish DNA virus which belongs to genus Ranavirus, family Iridoviridae. Here, using single-virus tracking technology in combination with biochemical assays, we investigated the crucial events during SGIV entry and demonstrated that SGIV entered grouper cells via the clathrin-mediated endocytic pathway in a pH-dependent manner but not via caveola-dependent endocytosis. Furthermore, we propose for the first time that macropinocytosis is involved in Iridovirus entry. Together, this work not only contributes greatly to understating Iridovirus pathogenesis but also provides an ideal model for exploring the behavior of DNA viruses in living cells.
Youhua Huang - One of the best experts on this subject based on the ideXlab platform.
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soft shelled turtle Iridovirus enters cells via cholesterol dependent clathrin mediated endocytosis as well as macropinocytosis
2018Co-Authors: Youhua Huang, Shaowen Wang, Xiaohong Huang, Qiwei QinAbstract:Ranaviruses are nucleoplasmic large DNA viruses that can cause major economic losses in the aquaculture industry and pose a severe threat to global ecological diversity. The available literature demonstrates that classifiable members of the genus Ranavirus enter cells via multiple and complicated routes. Here, we demonstrated the underlying cellular entry mechanism of soft-shelled turtle Iridovirus (STIV) using green fluorescence tagged recombinant virus. Treatment with chlorpromazine, sucrose, ethyl-isopropyl amiloride, chloroquine or bafilomycin A1 all significantly decreased STIV infection, suggesting that STIV uses clathrin-mediated endocytosis and macropinocytosis to enter cells via a pH-dependent pathway. Depletion of cellular cholesterol with methyl-β-cyclodextrin significantly inhibited STIV entry, but neither filipin III nor nystatin did, suggesting that STIV entry was cholesterol dependent but caveola independent. Treatment with dynasore, genistein, ML-7 or cytochalasin D all significantly inhibited STIV infection, indicating that Rac GTPase and myosin II activity were required for the macropinocytosis-like pathway as well as actin polymerization. Our findings suggest that the molecular events involved in STIV entry are not identical to those of other ranavirus isolates. Our results also extend our understanding of the molecular mechanism of Iridovirus entry and pathogenesis.
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antiviral role of grouper sting against Iridovirus infection
2015Co-Authors: Youhua Huang, Xiaohong Huang, Zhengliang Ouyang, Shina Wei, Jingguang Wei, Wei Wang, Sheng Zhou, Qiwei QinAbstract:Stimulator of interferon genes (STING, also known as MITA, ERIS, MPYS or TMEM173) has been identified as a central component in the innate immune response to cytosolic DNA and RNA derived from different pathogens. However, the detailed role of STING during fish Iridovirus infection still remained largely unknown. Here, the STING homolog from grouper Epinephelus coioides (EcSTING) was cloned and its effects on IFN response and antiviral activity were investigated. The full-length EcSTING cDNA was composed of 1590 bp and encoded a polypeptide of 409 amino acids with 80% identity to STING homolog from large yellow croaker. Amino acid alignment analysis indicated that EcSTING contained 4 predicated transmembrane motifs (TMs) in the N terminal, and a C-terminal domain (CTD) which consisted of a dimerization domain (DD), c-di-GMP-binding domain (CBD) and a C-terminal tail (CTT). Expression profile analysis revealed that EcSTING was abundant in gill, spleen, brain, skin, and liver. Upon different stimuli in vivo, the EcSTING transcript was dramatically up-regulated after challenging with Singapore grouper Iridovirus (SGIV), lipopolysaccharide (LPS) and polyinosin-polycytidylic acid (poly I:C). Reporter gene assay showed that EcSTING activated ISRE, zebrafish type I IFN and type III IFN promoter in vitro. Mutant analysis showed that IFN promoter activity was mostly mediated by the phosphorylation sites at serine residue S379 and S387. Moreover, EcSTING induced type I and III IFN promoter activity could be impaired by overexpression of EcIRF3-DN or EcIRF7-DN, suggesting that EcSTING mediated IFN response in IRF3/IRF7 dependent manner. In addition, the cytopathic effect (CPE) progression of SGIV infection and viral protein synthesis was significantly inhibited by overexpression of EcSTING, and the inhibitory effect was abolished in serine residue S379 and S387 mutant transfected cells. Together, our results demonstrated that EcSTING might be an important regulator of grouper innate immune response against Iridovirus infection.
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entry of a novel marine dna virus singapore grouper Iridovirus into host cells occurs via clathrin mediated endocytosis and macropinocytosis in a ph dependent manner
2014Co-Authors: Shaowen Wang, Youhua Huang, Xiaohong Huang, Xian Hao, Mingjun Cai, Hongda Wang, Qiwei QinAbstract:Iridoviruses are nucleocytoplasmic DNA viruses which cause great economic losses in the aquaculture industry but also show significant threat to global biodiversity. However, a lack of host cells has resulted in poor progress in clarifying Iridovirus behavior. We investigated the crucial events during virus entry using a combination of single-virus tracking and biochemical assays, based on the established virus-cell infection model for Singapore grouper Iridovirus (SGIV). SGIV infection in host cells was strongly inhibited when cells were pretreated with drugs blocking clathrin-mediated endocytosis, including sucrose and chlorpromazine. Inhibition of key regulators of macropinocytosis, including Na+/H+ exchanger, Rac1 GTPase, p21-activated kinase 1 (PAK1), protein kinase C (PKC), and myosin II, significantly reduced SGIV uptake. Cy5-labeled SGIV particles were observed to colocalize with clathrin and macropinosomes. In contrast, disruption of cellular cholesterol by methyl-β-cyclodextrin and nystatin had no effect on virus infection, suggesting that SGIV entered grouper cells via the clathrin-mediated endocytic pathway and macropinocytosis but not via caveola-dependent endocytosis. Furthermore, inhibitors of endosome acidification such as chloroquine and bafilomycin A1 blocked virus infection, indicating that SGIV entered cells in a pH-dependent manner. In addition, SGIV particles were observed to be transported along both microtubules and actin filaments, and intracellular SGIV motility was remarkably impaired by depolymerization of microtubules or actin filaments. The results of this study for the first time demonstrate that not only the clathrin-dependent pathway but also macropinocytosis are involved in fish DNA enveloped virus entry, thus providing a convenient tactic for exploring the life cycle of DNA viruses. IMPORTANCE Virus entry into host cells is critically important for initiating infections and is usually recognized as an ideal target for the design of antiviral strategies. Iridoviruses are large DNA viruses which cause serious threats to ecological diversity and the aquaculture industry worldwide. However, the current understanding of Iridovirus entry is limited and controversial. Singapore grouper Iridovirus (SGIV) is a novel marine fish DNA virus which belongs to genus Ranavirus, family Iridoviridae. Here, using single-virus tracking technology in combination with biochemical assays, we investigated the crucial events during SGIV entry and demonstrated that SGIV entered grouper cells via the clathrin-mediated endocytic pathway in a pH-dependent manner but not via caveola-dependent endocytosis. Furthermore, we propose for the first time that macropinocytosis is involved in Iridovirus entry. Together, this work not only contributes greatly to understating Iridovirus pathogenesis but also provides an ideal model for exploring the behavior of DNA viruses in living cells.
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Identification of lymphocystis disease virus from paradise fish Macropodus opercularis (LCDV-PF)
2014Co-Authors: Juan Feng, Youhua HuangAbstract:Iridoviruses are large DNA viruses that are subdivided into five genera: Ranavirus, Megalocytivirus, Lymphocystivirus, ChlorIridovirus and Iridovirus. The Iridovirus lymphocystis disease virus (LCDV) is an important fish pathogen that can infect marine and freshwater fish worldwide. In this study, we have identified the pathogen in paradise fish (Macropodus opercularis) with lymphocystis. On the skin and fins of diseased paradise fish, a large number of nodules were observed. H&E staining showed that the nodules were composed of encapsulated hypertrophied cells. Using electron microscopy, numerous virus particles with a diameter of >210 nm and with hexagonal profiles were observed in the cytoplasm. Phylogenetic analysis based on the major capsid protein (MCP), DNA polymerase and myristylated membrane protein (MMP) genes revealed that LCDV from paradise fish (LCDV-PF) was closely related to lymphocystis disease virus from China (LCDV-C), followed by lymphocystis disease virus 1 (LCDV-1). Taken together, our data provide the first molecular evidence that, in addition to megalocytivirus, LCDV is an important iridoviral pathogen in paradise fish besides megalocytivirus.
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characterization of an envelope gene vp19 from singapore grouper Iridovirus
2013Co-Authors: Xiaohong Huang, Youhua Huang, Shaowen Wang, Jie Gong, Zhengliang Ouyang, Xiuli Chen, Qiwei QinAbstract:Background Viral envelope proteins are always proposed to exert important function during virus infection and replication. Vertebrate Iridoviruses are enveloped large DNA virus, which can cause great economic losses in aquaculture and ecological destruction. Although numerous Iridovirus envelope proteins have been identified using bioinformatics and proteomic methods, their roles in virus infection remained largely unknown.
Xiaohong Huang - One of the best experts on this subject based on the ideXlab platform.
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identification of major capsid protein as a potential biomarker of grouper Iridovirus infected cells using aptamers selected by selex
2019Co-Authors: Mingzhu Liu, Xiaohong Huang, Hehe Xiao, Shina Wei, Qiwei QinAbstract:Biomarkers have important roles in disease pathogenesis, and serve as important disease indicators for developing novel diagnostic and therapeutic approaches. Grouper Iridovirus is a nucleocytoplasmic DNA virus, which not only causes great economic losses in mariculture but also seriously threatens the global biodiversity. However, a lack of biomarkers has limited the progress in clarifying Iridovirus pathogenesis. Here, we report novel molecular probes, aptamers, for specific identification of biomarkers in grouper Iridovirus-infected cells. Aptamers are selected by SELEX, which is a completely different approach from conventional antibody-based methods for biomarkers discovery. Aptamer-based technology is the unique efficient selection for cell-specific target molecules, and helps find out new biomarkers without the knowledge of characteristics of proteins expressed on virus-infected cell surface. With the implementation of a two-step strategy (aptamer selection and biomarker discovery), combined with mass spectrometry, grouper Iridovirus major capsid protein was ultimately identified as a potential biomarker of aptamer Q5 for grouper Iridovirus infection. The specific interactions of aptamer Q5 and MCP were experimentally validated by several assays, including EMSA, co-localization of fluorescence by LSCM, binding competition tests, and siRNA silencing tests by flow cytometry. This aptamer-based method for biomarkers discovery developed with grouper Iridovirus-infected cells could be applicable to other types of virus infection, markedly improve our studies of biomarker discovery and virus pathogenesis, and further facilitate the development of diagnostic tools and therapeutic approaches to treat virus infection.
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soft shelled turtle Iridovirus enters cells via cholesterol dependent clathrin mediated endocytosis as well as macropinocytosis
2018Co-Authors: Youhua Huang, Shaowen Wang, Xiaohong Huang, Qiwei QinAbstract:Ranaviruses are nucleoplasmic large DNA viruses that can cause major economic losses in the aquaculture industry and pose a severe threat to global ecological diversity. The available literature demonstrates that classifiable members of the genus Ranavirus enter cells via multiple and complicated routes. Here, we demonstrated the underlying cellular entry mechanism of soft-shelled turtle Iridovirus (STIV) using green fluorescence tagged recombinant virus. Treatment with chlorpromazine, sucrose, ethyl-isopropyl amiloride, chloroquine or bafilomycin A1 all significantly decreased STIV infection, suggesting that STIV uses clathrin-mediated endocytosis and macropinocytosis to enter cells via a pH-dependent pathway. Depletion of cellular cholesterol with methyl-β-cyclodextrin significantly inhibited STIV entry, but neither filipin III nor nystatin did, suggesting that STIV entry was cholesterol dependent but caveola independent. Treatment with dynasore, genistein, ML-7 or cytochalasin D all significantly inhibited STIV infection, indicating that Rac GTPase and myosin II activity were required for the macropinocytosis-like pathway as well as actin polymerization. Our findings suggest that the molecular events involved in STIV entry are not identical to those of other ranavirus isolates. Our results also extend our understanding of the molecular mechanism of Iridovirus entry and pathogenesis.
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antiviral role of grouper sting against Iridovirus infection
2015Co-Authors: Youhua Huang, Xiaohong Huang, Zhengliang Ouyang, Shina Wei, Jingguang Wei, Wei Wang, Sheng Zhou, Qiwei QinAbstract:Stimulator of interferon genes (STING, also known as MITA, ERIS, MPYS or TMEM173) has been identified as a central component in the innate immune response to cytosolic DNA and RNA derived from different pathogens. However, the detailed role of STING during fish Iridovirus infection still remained largely unknown. Here, the STING homolog from grouper Epinephelus coioides (EcSTING) was cloned and its effects on IFN response and antiviral activity were investigated. The full-length EcSTING cDNA was composed of 1590 bp and encoded a polypeptide of 409 amino acids with 80% identity to STING homolog from large yellow croaker. Amino acid alignment analysis indicated that EcSTING contained 4 predicated transmembrane motifs (TMs) in the N terminal, and a C-terminal domain (CTD) which consisted of a dimerization domain (DD), c-di-GMP-binding domain (CBD) and a C-terminal tail (CTT). Expression profile analysis revealed that EcSTING was abundant in gill, spleen, brain, skin, and liver. Upon different stimuli in vivo, the EcSTING transcript was dramatically up-regulated after challenging with Singapore grouper Iridovirus (SGIV), lipopolysaccharide (LPS) and polyinosin-polycytidylic acid (poly I:C). Reporter gene assay showed that EcSTING activated ISRE, zebrafish type I IFN and type III IFN promoter in vitro. Mutant analysis showed that IFN promoter activity was mostly mediated by the phosphorylation sites at serine residue S379 and S387. Moreover, EcSTING induced type I and III IFN promoter activity could be impaired by overexpression of EcIRF3-DN or EcIRF7-DN, suggesting that EcSTING mediated IFN response in IRF3/IRF7 dependent manner. In addition, the cytopathic effect (CPE) progression of SGIV infection and viral protein synthesis was significantly inhibited by overexpression of EcSTING, and the inhibitory effect was abolished in serine residue S379 and S387 mutant transfected cells. Together, our results demonstrated that EcSTING might be an important regulator of grouper innate immune response against Iridovirus infection.
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entry of a novel marine dna virus singapore grouper Iridovirus into host cells occurs via clathrin mediated endocytosis and macropinocytosis in a ph dependent manner
2014Co-Authors: Shaowen Wang, Youhua Huang, Xiaohong Huang, Xian Hao, Mingjun Cai, Hongda Wang, Qiwei QinAbstract:Iridoviruses are nucleocytoplasmic DNA viruses which cause great economic losses in the aquaculture industry but also show significant threat to global biodiversity. However, a lack of host cells has resulted in poor progress in clarifying Iridovirus behavior. We investigated the crucial events during virus entry using a combination of single-virus tracking and biochemical assays, based on the established virus-cell infection model for Singapore grouper Iridovirus (SGIV). SGIV infection in host cells was strongly inhibited when cells were pretreated with drugs blocking clathrin-mediated endocytosis, including sucrose and chlorpromazine. Inhibition of key regulators of macropinocytosis, including Na+/H+ exchanger, Rac1 GTPase, p21-activated kinase 1 (PAK1), protein kinase C (PKC), and myosin II, significantly reduced SGIV uptake. Cy5-labeled SGIV particles were observed to colocalize with clathrin and macropinosomes. In contrast, disruption of cellular cholesterol by methyl-β-cyclodextrin and nystatin had no effect on virus infection, suggesting that SGIV entered grouper cells via the clathrin-mediated endocytic pathway and macropinocytosis but not via caveola-dependent endocytosis. Furthermore, inhibitors of endosome acidification such as chloroquine and bafilomycin A1 blocked virus infection, indicating that SGIV entered cells in a pH-dependent manner. In addition, SGIV particles were observed to be transported along both microtubules and actin filaments, and intracellular SGIV motility was remarkably impaired by depolymerization of microtubules or actin filaments. The results of this study for the first time demonstrate that not only the clathrin-dependent pathway but also macropinocytosis are involved in fish DNA enveloped virus entry, thus providing a convenient tactic for exploring the life cycle of DNA viruses. IMPORTANCE Virus entry into host cells is critically important for initiating infections and is usually recognized as an ideal target for the design of antiviral strategies. Iridoviruses are large DNA viruses which cause serious threats to ecological diversity and the aquaculture industry worldwide. However, the current understanding of Iridovirus entry is limited and controversial. Singapore grouper Iridovirus (SGIV) is a novel marine fish DNA virus which belongs to genus Ranavirus, family Iridoviridae. Here, using single-virus tracking technology in combination with biochemical assays, we investigated the crucial events during SGIV entry and demonstrated that SGIV entered grouper cells via the clathrin-mediated endocytic pathway in a pH-dependent manner but not via caveola-dependent endocytosis. Furthermore, we propose for the first time that macropinocytosis is involved in Iridovirus entry. Together, this work not only contributes greatly to understating Iridovirus pathogenesis but also provides an ideal model for exploring the behavior of DNA viruses in living cells.
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characterization of an envelope gene vp19 from singapore grouper Iridovirus
2013Co-Authors: Xiaohong Huang, Youhua Huang, Shaowen Wang, Jie Gong, Zhengliang Ouyang, Xiuli Chen, Qiwei QinAbstract:Background Viral envelope proteins are always proposed to exert important function during virus infection and replication. Vertebrate Iridoviruses are enveloped large DNA virus, which can cause great economic losses in aquaculture and ecological destruction. Although numerous Iridovirus envelope proteins have been identified using bioinformatics and proteomic methods, their roles in virus infection remained largely unknown.
Qiya Zhang - One of the best experts on this subject based on the ideXlab platform.
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rana grylio virus rgv envelope protein 2l subcellular localization and essential roles in virus infectivity revealed by conditional lethal mutant
2014Co-Authors: Libo He, Fei Ke, Qiya ZhangAbstract:Rana grylio virus (RGV) is a pathogenic Iridovirus that has resulted in high mortality in cultured frog. Here, an envelope protein gene, 2L, was identified from RGV and its possible role in virus infection was investigated. Database searches found that RGV 2L had homologues in all sequenced Iridoviruses and is a core gene of Iridoviruses. Western blotting detection of purified RGV virions confirmed that 2L protein was associated with virion membrane. Fluorescence localization revealed that 2L protein co-localized with viral factories in RGV infected cells. In co-transfected cells, 2L protein co-localized with two other viral envelope proteins, 22R and 53R. However, 2L protein did not co-localize with the major capsid protein of RGV in co-transfected cells. Meanwhile, fluorescence observation showed that 2L protein co-localized with endoplasmic reticulum, but did not co-localize with mitochondria and Golgi apparatus. Moreover, a conditional lethal mutant virus containing the lac repressor/operator system was constructed to investigate the role of RGV 2L in virus infection. The ability to form plaques and the virus titres were strongly reduced when expression of 2L was repressed. Therefore, the current data showed that 2L protein is essential for virus infection. Our study is the first report, to our knowledge, of co-localization between envelope proteins in Iridovirus and provides new insights into the understanding of envelope proteins in Iridovirus.
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a conditional lethal mutation in rana grylio virus orf 53r resulted in a marked reduction in virion formation
2013Co-Authors: Libo He, Fei Ke, Qiya ZhangAbstract:Rana grylio virus (RGV) is a pathogenic Iridovirus that has resulted in high mortality in cultured frog. Here, a recombinant RGV (i53R-RGV-lacIO) containing the inducible lac repressor! operator system was constructed. i53R-RGV-lacIO was a conditional lethal mutant in which the expression of envelope protein 53R was regulated by IPTG. i53R-RGV-lacIO shared characteristics similar to RGV in the presence of IPTG. However, the expression level of 53R, the ability of plaques formation, and the virus titers were strongly reduced in the absence of IPTG. Electron microscopy showed that the number of progeny virus produced by i53R-RGV-lacIO was remarkably reduced without IPTG. Furthermore, over-expression of 53R in vitro could increase titers of i53R-RGV-lacIO in the absence of IPTG. Therefore, the current data suggested that the lac repressor/operator system could regulate gene expression in the recombinant Iridovirus. Our study was thought to be the first report of the system in aquatic virus. (C) 2013 Elsevier B.V. All rights reserved.
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a conditional lethal mutation in rana grylio virus orf 53r resulted in a marked reduction in virion formation
2013Co-Authors: Xiaochan Gao, Qiya ZhangAbstract:Rana grylio virus (RGV) is a pathogenic Iridovirus that has resulted in high mortality in cultured frog. Here, a recombinant RGV (i53R-RGV-lacIO) containing the inducible lac repressor/operator system was constructed. i53R-RGV-lacIO was a conditional lethal mutant in which the expression of envelope protein 53R was regulated by IPTG. i53R-RGV-lacIO shared characteristics similar to RGV in the presence of IPTG. However, the expression level of 53R, the ability of plaques formation, and the virus titers were strongly reduced in the absence of IPTG. Electron microscopy showed that the number of progeny virus produced by i53R-RGV-lacIO was remarkably reduced without IPTG. Furthermore, over-expression of 53R in vitro could increase titers of i53R-RGV-lacIO in the absence of IPTG. Therefore, the current data suggested that the lac repressor/operator system could regulate gene expression in the recombinant Iridovirus. Our study was thought to be the first report of the system in aquatic virus.
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Multiple sequence alignment of 50L homologues in Iridoviruses.
2013Co-Authors: Xiao-ying Lei, Qiya ZhangAbstract:RGV, Rana grylio virus; STIV, soft-shelled turtle Iridovirus; CMTV, common midwife toad ranavirus; EHNV, epizootic hematopoietic necrosis virus; ATV, Ambystoma tigrinum virus; TFV, tiger frog virus; FV3, frog virus 3; RRV, Regina ranavirus; GIV, grouper Iridovirus; SGIV, Singapore grouper Iridovirus; LCDV-1, lymphocystis disease virus 1; LCDV-C, lymphocystis disease virus-China. The completely conserved amino acid residues are indicated by a black background, while the grey background are partially conserved residues with greater than 80% identity, and key amino acid residues in SAP domain are indicated by colorful backgrounds. Conserved motifs are shown by rectangles and labeled as Repeated Sequence, NLS motif and SAP domain above the alignment, respectively. Gaps (dashes) were introduced to maximize the alignment.
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sequencing and analysis of the complete genome of rana grylio virus rgv
2012Co-Authors: Tong Ou, Qiya ZhangAbstract:Infection with Rana grylio virus (RGV), an Iridovirus isolated in China in 1995, resulted in a high mortality rate in frogs. The complete genome sequence of RGV was determined and analyzed. The genomic DNA was 105,791 bp long, with 106 open reading frames (ORFs). Dot plot analysis showed that the gene order of RGV shared colinearity with three completely sequenced ranaviruses. A phylogenetic tree was constructed based on concatenated sequences of Iridovirus 26 core-gene-encoded proteins, and the result showed high bootstrap support for RGV being a member of the genus Ranavirus and that Iridoviruses of other genera also clustered closely. A microRNA (miRNA) prediction revealed that RGV could encode 18 mature miRNAs, many of which were located near genes associated with virus replication. Thirty-three repeated sequences were found in the RGV genome. These results provide insight into the genetic nature of RGV and are useful for laboratory diagnosis for vertebrate Iridoviruses.
Shaowen Wang - One of the best experts on this subject based on the ideXlab platform.
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soft shelled turtle Iridovirus enters cells via cholesterol dependent clathrin mediated endocytosis as well as macropinocytosis
2018Co-Authors: Youhua Huang, Shaowen Wang, Xiaohong Huang, Qiwei QinAbstract:Ranaviruses are nucleoplasmic large DNA viruses that can cause major economic losses in the aquaculture industry and pose a severe threat to global ecological diversity. The available literature demonstrates that classifiable members of the genus Ranavirus enter cells via multiple and complicated routes. Here, we demonstrated the underlying cellular entry mechanism of soft-shelled turtle Iridovirus (STIV) using green fluorescence tagged recombinant virus. Treatment with chlorpromazine, sucrose, ethyl-isopropyl amiloride, chloroquine or bafilomycin A1 all significantly decreased STIV infection, suggesting that STIV uses clathrin-mediated endocytosis and macropinocytosis to enter cells via a pH-dependent pathway. Depletion of cellular cholesterol with methyl-β-cyclodextrin significantly inhibited STIV entry, but neither filipin III nor nystatin did, suggesting that STIV entry was cholesterol dependent but caveola independent. Treatment with dynasore, genistein, ML-7 or cytochalasin D all significantly inhibited STIV infection, indicating that Rac GTPase and myosin II activity were required for the macropinocytosis-like pathway as well as actin polymerization. Our findings suggest that the molecular events involved in STIV entry are not identical to those of other ranavirus isolates. Our results also extend our understanding of the molecular mechanism of Iridovirus entry and pathogenesis.
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entry of a novel marine dna virus singapore grouper Iridovirus into host cells occurs via clathrin mediated endocytosis and macropinocytosis in a ph dependent manner
2014Co-Authors: Shaowen Wang, Youhua Huang, Xiaohong Huang, Xian Hao, Mingjun Cai, Hongda Wang, Qiwei QinAbstract:Iridoviruses are nucleocytoplasmic DNA viruses which cause great economic losses in the aquaculture industry but also show significant threat to global biodiversity. However, a lack of host cells has resulted in poor progress in clarifying Iridovirus behavior. We investigated the crucial events during virus entry using a combination of single-virus tracking and biochemical assays, based on the established virus-cell infection model for Singapore grouper Iridovirus (SGIV). SGIV infection in host cells was strongly inhibited when cells were pretreated with drugs blocking clathrin-mediated endocytosis, including sucrose and chlorpromazine. Inhibition of key regulators of macropinocytosis, including Na+/H+ exchanger, Rac1 GTPase, p21-activated kinase 1 (PAK1), protein kinase C (PKC), and myosin II, significantly reduced SGIV uptake. Cy5-labeled SGIV particles were observed to colocalize with clathrin and macropinosomes. In contrast, disruption of cellular cholesterol by methyl-β-cyclodextrin and nystatin had no effect on virus infection, suggesting that SGIV entered grouper cells via the clathrin-mediated endocytic pathway and macropinocytosis but not via caveola-dependent endocytosis. Furthermore, inhibitors of endosome acidification such as chloroquine and bafilomycin A1 blocked virus infection, indicating that SGIV entered cells in a pH-dependent manner. In addition, SGIV particles were observed to be transported along both microtubules and actin filaments, and intracellular SGIV motility was remarkably impaired by depolymerization of microtubules or actin filaments. The results of this study for the first time demonstrate that not only the clathrin-dependent pathway but also macropinocytosis are involved in fish DNA enveloped virus entry, thus providing a convenient tactic for exploring the life cycle of DNA viruses. IMPORTANCE Virus entry into host cells is critically important for initiating infections and is usually recognized as an ideal target for the design of antiviral strategies. Iridoviruses are large DNA viruses which cause serious threats to ecological diversity and the aquaculture industry worldwide. However, the current understanding of Iridovirus entry is limited and controversial. Singapore grouper Iridovirus (SGIV) is a novel marine fish DNA virus which belongs to genus Ranavirus, family Iridoviridae. Here, using single-virus tracking technology in combination with biochemical assays, we investigated the crucial events during SGIV entry and demonstrated that SGIV entered grouper cells via the clathrin-mediated endocytic pathway in a pH-dependent manner but not via caveola-dependent endocytosis. Furthermore, we propose for the first time that macropinocytosis is involved in Iridovirus entry. Together, this work not only contributes greatly to understating Iridovirus pathogenesis but also provides an ideal model for exploring the behavior of DNA viruses in living cells.
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characterization of an envelope gene vp19 from singapore grouper Iridovirus
2013Co-Authors: Xiaohong Huang, Youhua Huang, Shaowen Wang, Jie Gong, Zhengliang Ouyang, Xiuli Chen, Qiwei QinAbstract:Background Viral envelope proteins are always proposed to exert important function during virus infection and replication. Vertebrate Iridoviruses are enveloped large DNA virus, which can cause great economic losses in aquaculture and ecological destruction. Although numerous Iridovirus envelope proteins have been identified using bioinformatics and proteomic methods, their roles in virus infection remained largely unknown.
