Dataset Information

Visualization of Assembly Intermediates and Budding Vacuoles of Singapore Grouper Iridovirus in Grouper Embryonic Cells.

ABSTRACT: Iridovirid infection is associated with the catastrophic loss in aquaculture industry and the population decline of wild amphibians and reptiles, but none of the iridovirid life cycles have been well explored. Here, we report the detailed visualization of the life cycle of Singapore grouper iridovirus (SGIV) in grouper cells by cryo-electron microscopy (cryoEM) and tomography (ET). EM imaging revealed that SGIV viral particles have an outer capsid layer, and the interaction of this layer with cellular plasma membrane initiates viral entry. Subsequent viral replication leads to formation of a viral assembly site (VAS), where membranous structures emerge as precursors to recruit capsid proteins to form an intermediate, double-shell, crescent-shaped structure, which curves to form icosahedral capsids. Knockdown of the major capsid protein eliminates the formation of viral capsids. As capsid formation progresses, electron-dense materials known to be involved in DNA encapsidation accumulate within the capsid until it is fully occupied. Besides the well-known budding mechanism through the cell periphery, we demonstrate a novel budding process in which viral particles bud into a tubular-like structure within vacuoles. This budding process may denote a new strategy used by SGIV to disseminate viral particles into neighbor cells while evading host immune response.

SUBMITTER: Liu Y

PROVIDER: S-EPMC4698634 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature

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Visualization of Assembly Intermediates and Budding Vacuoles of Singapore Grouper Iridovirus in Grouper Embryonic Cells.

Liu Yang Y Tran Bich Ngoc BN Wang Fan F Ounjai Puey P Wu Jinlu J Hew Choy L CL

Scientific reports 20160104

Iridovirid infection is associated with the catastrophic loss in aquaculture industry and the population decline of wild amphibians and reptiles, but none of the iridovirid life cycles have been well explored. Here, we report the detailed visualization of the life cycle of Singapore grouper iridovirus (SGIV) in grouper cells by cryo-electron microscopy (cryoEM) and tomography (ET). EM imaging revealed that SGIV viral particles have an outer capsid layer, and the interaction of this layer with ce ...[more]

PMID: 26727547

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Project description:BackgroundMicroRNAs (miRNAs) are ubiquitous non-coding RNAs that regulate gene expression at the post-transcriptional level. An increasing number of studies has revealed that viruses can also encode miRNAs, which are proposed to be involved in viral replication and persistence, cell-mediated antiviral immune response, angiogenesis, and cell cycle regulation. Singapore grouper iridovirus (SGIV) is a pathogenic iridovirus that has severely affected grouper aquaculture in China and Southeast Asia. Comprehensive knowledge about the related miRNAs during SGIV infection is helpful for understanding the infection and the pathogenic mechanisms.Methodology/principal findingsTo determine whether SGIV encoded miRNAs during infection, a small RNA library derived from SGIV-infected grouper (GP) cells was constructed and sequenced by Illumina/Solexa deep-sequencing technology. We recovered 6,802,977 usable reads, of which 34,400 represented small RNA sequences encoded by SGIV. Sixteen novel SGIV-encoded miRNAs were identified by a computational pipeline, including a miRNA that shared a similar sequence to herpesvirus miRNA HSV2-miR-H4-5p, which suggests miRNAs are conserved in far related viruses. Generally, these 16 miRNAs are dispersed throughout the SGIV genome, whereas three are located within the ORF057L region. Some SGIV-encoded miRNAs showed marked sequence and length heterogeneity at their 3' and/or 5' end that could modulate their functions. Expression levels and potential biological activities of these viral miRNAs were examined by stem-loop quantitative RT-PCR and luciferase reporter assay, respectively, and 11 of these viral miRNAs were present and functional in SGIV-infected GP cells.ConclusionsOur study provided a genome-wide view of miRNA production for iridoviruses and identified 16 novel viral miRNAs. To the best of our knowledge, this is the first experimental demonstration of miRNAs encoded by aquatic animal viruses. The results provide a useful resource for further in-depth studies on SGIV infection and iridovirus pathogenesis.

Project description:BackgroundOrange-spotted grouper (Epinephelus coioides) is an economically important marine fish cultured in China and Southeast Asian countries. The emergence of infectious viral diseases, including iridovirus and betanodavirus, have severely affected food products based on this species, causing heavy economic losses. Limited available information on the genomics of E. coioides has hampered the understanding of the molecular mechanisms that underlie host-virus interactions. In this study, we used a 454 pyrosequencing method to investigate differentially-expressed genes in the spleen of the E. coioides infected with Singapore grouper iridovirus (SGIV).ResultsUsing 454 pyrosequencing, we obtained abundant high-quality ESTs from two spleen-complementary DNA libraries which were constructed from SGIV-infected (V) and PBS-injected fish (used as a control: C). A total of 407,027 and 421,141 ESTs were produced in control and SGIV infected libraries, respectively. Among the assembled ESTs, 9,616 (C) and 10,426 (V) ESTs were successfully matched against known genes in the NCBI non-redundant (nr) database with a cut-off E-value above 10-5. Gene ontology (GO) analysis indicated that "cell part", "cellular process" and "binding" represented the largest category. Among the 25 clusters of orthologous group (COG) categories, the cluster for "translation, ribosomal structure and biogenesis" represented the largest group in the control (185 ESTs) and infected (172 ESTs) libraries. Further KEGG analysis revealed that pathways, including cellular metabolism and intracellular immune signaling, existed in the control and infected libraries. Comparative expression analysis indicated that certain genes associated with mitogen-activated protein kinase (MAPK), chemokine, toll-like receptor and RIG-I signaling pathway were alternated in response to SGIV infection. Moreover, changes in the pattern of gene expression were validated by qRT-PCR, including cytokines, cytokine receptors, and transcription factors, apoptosis-associated genes, and interferon related genes.ConclusionThis study provided abundant ESTs that could contribute greatly to disclosing novel genes in marine fish. Furthermore, the alterations of predicted gene expression patterns reflected possible responses of these fish to the virus infection. Taken together, our data not only provided new information for identification of novel genes from marine vertebrates, but also shed new light on the understanding of defense mechanisms of marine fish to viral pathogens.

Dataset Information

Visualization of Assembly Intermediates and Budding Vacuoles of Singapore Grouper Iridovirus in Grouper Embryonic Cells.

Publications

Visualization of Assembly Intermediates and Budding Vacuoles of Singapore Grouper Iridovirus in Grouper Embryonic Cells.

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