Subcellular redistribution and sequential recruitment of macromolecular components during SGIV assembly

Abstract

Virus infection consists of entry, synthesis of macromolecular components, virus assembly and release. Understanding of the mechanisms underlying each event is necessary for the intervention of virus infection in human healthcare and agriculture. Here we report the visualization of Singapore grouper iridovirus (SGIV) assembly in the medaka haploid embryonic stem (ES) cell line HX1. SGIV is a highly infectious DNA virus that causes a massive loss in marine aquaculture. Ectopic expression of VP88GFP, a fusion between green fluorescent protein and the envelope protein VP088, did not compromise the ES cell properties and susceptibility to SGIV infection. Although VP88GFP disperses evenly in the cytoplasm of non-infected cells, it undergoes aggregation and redistribution in SGIV-infected cells. Real-time visualization revealed multiple key stages of VP88GFP redistribution and the dynamics of viral assembly site (VAS). Specifically, VP88GFP entry into and condensation in the VAS occurred within a 6-h duration, a similar duration was observed also for the release of VP88GFP-containing SGIV out of the cell. Taken together, VP088 is an excellent marker for visualizing the SGIV infection process. Our results provide new insight into macromolecular component recruitment and SGIV assembly.

Keywords

ES cell / medaka / orf088 / SGIV / viral assembly

References

[1] Carette JE, Guimaraes CP, Varadarajan M, Park AS, Wuethrich I, Godarova A, Kotecki M, Cochran BH, Spooner E, Ploegh HL (2009) Haploid genetic screens in human cells identify host factors used by pathogens. Science 326:1231–1235

[2] Chen LM, Wang F, Song W, Hew CL (2006) Temporal and differential gene expression of Singapore grouper iridovirus. J Gen Virol 87:2907–2915

[3] Dimmock NJ, Easton AJ, Leppard K (2007) Introduction to modern virology, 6th edn. Blackwell, Malden

[4] Ge XY, Li JL, Yang XL, Chmura AA, Zhu G, Epstein JH, Mazet JK, Hu B, Zhang W, Peng C (2013) Isolation and characterization of a bat SARS-like coronavirus that uses the ACE2 receptor. Nature 503:535–538

[5] Heath CM, Windsor M, Wileman T (2001) Aggresomes resemble sites specialized for virus assembly. J Cell Biol 153:449–455

[6] Hong Y, Schartl M (2006) Isolation and differentiation of medaka embryonic stem cells. Methods Mol Biol 329:3–16

[7] Hong Y, Winkler C, Schartl M (1996) Pluripotency and differentiation of embryonic stem cell lines from the medakafish (Oryzias latipes). Mech Dev 60:33–44

[8] Hong Y, Winkler C, Schartl M (1998) Production of medakafish chimeras from a stable embryonic stem cell line. Proc Natl Acad Sci USA 95:3679–3684

[9] Hong Y, Chen S, Gui J, Schartl M (2004a) Retention of the developmental pluripotency in medaka embryonic stem cells after gene transfer and long-term drug selection for gene targeting in fish. Transgenic Res 13:41–50

[10] Hong Y, Liu T, Zhao H, Xu H, Wang W, Liu R, Chen T, Deng J, Gui J (2004b) Establishment of a normal medakafish spermatogonial cell line capable of sperm production in vitro. Proc Natl Acad Sci USA 101:8011–8016

[11] Hong N, Li M, Zeng Z, Yi M, Deng J, Gui J, Winkler C, Schartl M, Hong Y (2010) Accessibility of host cell lineages to medaka stem cells depends on genetic background and irradiation of recipient embryos. Cell Mol Life Sci 67:1189–1202

[12] Huang X, Huang Y, Gong J, Yan Y, Qin Q (2008) Identification and characterization of a putative lipopolysaccharide-induced TNFalpha factor (LITAF) homolog from Singapore grouper iridovirus. Biochem Biophys Res Commun 373:140–145

[13] Huang X, Huang Y, Sun J, Han X, Qin Q (2009) Characterization of two grouper Epinephelus akaara cell lines: application to studies of Singapore grouper iridovirus (SGIV) propagation and virushost interaction. Aquaculture 292:172–179

[14] Huang X, Huang Y, Ouyang Z, Xu L, Yan Y, Cui H, Han X, Qin Q (2011) Singapore grouper iridovirus, a large DNA virus, induces nonapoptotic cell death by a cell type dependent fashion and evokes ERK signaling. Apoptosis 16:831–845

[15] Huang X, Gong J, Huang Y, Ouyang Z, Wang S, Chen X, Qin Q (2013) Characterization of an envelope gene VP19 from Singapore grouper iridovirus. Virol J 10:354

[16] Li KS, Guan Y, Wang J, Smith GJ, Xu KM, Duan L, Rahardjo AP, Puthavathana P, Buranathai C, Nguyen TD (2004) Genesis of a highly pathogenic and potentially pandemic H5N1 influenza virus in eastern Asia. Nature 430:209–213

[17] Li M, Hong N, Xu H, Yi M, Li C, Gui J, Hong Y (2009) Medaka vasa is required for migration but not survival of primordial germ cells. Mech Dev 126:366–381

[18] Liu Y, Tran BN, Wang F, Ounjai P, Wu J, Hew CL (2016) Visualization of assembly intermediates and budding vacuoles of Singapore Grouper Iridovirus in Grouper embryonic cells. Sci Rep 6:18696

[19] Novoa RR, Calderita G, Arranz R, Fontana J, Granzow H, Risco C (2005) Virus factories: associations of cell organelles for viral replication and morphogenesis. Biol Cell 97:147–172

[20] Qin QW, Lam TJ, Sin YM, Shen H, Chang SF, Ngoh GH, Chen CL (2001) Electron microscopic observations of a marine fish iridovirus isolated from brown-spotted grouper, Epinephelus tauvina. J Virol Methods 98:17–24

[21] Qin QW, Chang SF, Ngoh-Lim GH, Gibson-Kueh S, Shi C, Lam TJ (2003) Characterization of a novel ranavirus isolated from grouper Epinephelus tauvina. Dis Aquat Organ 53:1–9

[22] Song WJ, Qin QW, Qiu J, Huang CH, Wang F, Hew CL (2004) Functional genomics analysis of Singapore grouper iridovirus: complete sequence determination and proteomic analysis. J Virol 78:12576–12590

[23] Teng Y, Hou Z, Gong J, Liu H, Xie X, Zhang L, Chen X, Qin QW (2008) Whole-genome transcriptional profiles of a novel marine fish iridovirus, Singapore grouper iridovirus (SGIV) in virusinfected grouper spleen cell cultures and in orange-spotted grouper, Epinephulus coioides. Virology 377:39–48

[24] Walker P, Winton JR (2010) Emergin viral diseases of fish and shrimp. Vet Res 41:24

[25] Wang S, Huang X, Huang Y, Hao X, Xu H, Cai M, Wang H, Qin Q (2014) 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. J Virol 88:13047–13063

[26] Williams T, Barbosa-Solomieu V, Chinchar VG (2005) A decade of advances in iridovirus research. Adv Virus Res 65:173–248

[27] Xia L, Cao J, Huang X, Qin Q (2009) Characterization of Singapore grouper iridovirus (SGIV) ORF086R, a putative homolog of ICP18 involved in cell growth control and virus replication. Arch Virol 154:1409–1416

[28] Xia L, Liang H, Huang Y, Ou-Yang Z, Qin Q (2010) Identification and characterization of Singapore grouper iridovirus (SGIV) ORF162L, an immediate-early gene involved in cell growth control and viral replication. Virus Res 147:30–39

[29] Yi M, Hong N, Hong Y (2009) Generation of medaka fish haploid embryonic stem cells. Science 326:430–433

[30] Yi M, Hong N, Hong Y (2010) Derivation and characterization of haploid embryonic stem cell cultures in medaka fish. Nat Protoc 5:1418–1430

[31] Yuan Y, Huang X, Zhang L, Zhu Y, Huang Y, Qin Q, Hong Y (2013) Medaka haploid embryonic stem cells are susceptible to Singapore grouper iridovirus as well as to other viruses of aquaculture fish species. J Gen Virol 94:2352–2359

[32] Yuan Y, Li M, Hong N, Hong Y (2014) Correlative light and electron microscopic analyses of mitochondrial distribution in blastomeres of early fish embryos. FASEB J 28:577–585

[33] Zhou S, Wan Q, Huang Y, Huang X, Cao J, Ye L, Lim TK, Lin Q, Qin Q (2011) Proteomic analysis of Singapore grouper iridovirus envelope proteins and characterization of a novel envelope protein VP088. Proteomics 11:2236–2248

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2016 The Author(s) 2016. This article is published with open access at Springerlink.com and journal.hep.com.cn