title: Molecular cloning, expression and functional analysis of ISG15 in orange-spotted grouper, Epinephelus coioides.
abstract: Interferon-stimulated gene 15 (ISG15) is an ubiquitin homolog that is significantly induced by type I interferons or viral infections. Groupers, Epinephelus spp. being maricultured in China and Southeast Asian countries, always suffer from virus infection, including iridovirus and nodavirus. To date, the roles of grouper genes, especially interferon related genes in virus infection remained largely unknown. Here, the ISG15 homolog (EcISG15) was cloned from grouper Epinephelus coioides and its immune response to Singapore grouper iridovirus (SGIV) and grouper nervous necrosis virus (GNNV) was investigated. The full-length EcISG15 cDNA was composed of 948 bp and encoded a polypeptide of 155 amino acids with 37-68% identity with the known ISG15 homologs from other fish species. Amino acid alignment analysis indicated that EcISG15 contained two ubiquitin-like (UBL) domains and an Ub-conjugation domain (LRGG). Expressional analysis showed that EcISG15 was dramatically induced by GNNV infection, poly I:C or poly dA-dT treatment, but no obvious changes were observed during SGIV infection. Immunofluorescence assay showed that EcISG15 localized mainly in the cytoplasm of grouper cells in response to poly I:C stimulation or GNNV infection, but not in mock or SGIV infected cells. Western blot analysis indicated that the ISGylation was absent in SGIV-infected cells, but significantly enhanced in GNNV-infected or poly I:C transfected cells, suggesting that EcISG15 might play different roles in SGIV and GNNV infection. Furthermore, overexpression of EcISG15 in vitro inhibited the transcription of GNNV genes significantly. Taken together, the results indicated that fish ISG15 might exert important roles against RNA virus infection.
authors: Key Laboratory of Marine Bio-resources Sustainable Utilization, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
authors: Xiaohong
authors: X
authors: Huang
authors: Youhua
authors: Y
authors: Huang
authors: Jia
authors: J
authors: Cai
authors: Shina
authors: S
authors: Wei
authors: Zhengliang
authors: Z
authors: Ouyang
authors: Qiwei
authors: Q
authors: Qin
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001492:Bass / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D002478:Cells, Cultured; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D029941:Fish Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D007113:Immunity, Innate; D008969:Molecular Sequence Data; D030381:Nodaviridae / Q000276:immunology; D009928:Organ Specificity; D010802:Phylogeny; D011070:Poly I-C / Q000276:immunology; D011067:Poly dA-dT / Q000276:immunology; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017995:Ranavirus / Q000276:immunology; D016415:Sequence Alignment; D025801:Ubiquitin / Q000737:chemistry / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism
chemical_list: D018076:DNA, Complementary; D029941:Fish Proteins; D012333:RNA, Messenger; D025801:Ubiquitin; D011067:Poly dA-dT; D011070:Poly I-C