title: Fish TRIM32 functions as a critical antiviral molecule against iridovirus and nodavirus.
abstract: Tripartite motif-containing 32 (TRIM32) has been demonstrated to pay vital roles in cancer, genetic disorders and antiviral immunity. However, the molecular functions of fish TRIM32 still remained largely unknown. Here, a novel TRIM32 gene from orange spotted grouper (EcTRIM32) was cloned and characterized. EcTRIM32 encoded a 685-aa protein which showed 93%, and 60% identity to large yellow croaker (Larimichthys crocea) and human (Homo sapiens), respectively. Amino acid alignment showed that EcTRIM32 contained a conserved RING-finger domain, a BBOX domain and NHL domain. In healthy grouper, the transcript of EcTRIM32 was predominantly detected in brain, liver, intestine, spleen and skin. After injection with Singapore grouper iridovirus (SGIV) and polyI:C, the relative expression of EcTRIM32 in grouper spleen was differently regulated, suggested that EcTRIM32 was involved in antiviral immune response. In transfected grouper spleen (GS) cells, EcTRIM32 displayed bright fluorescence aggregates or spots in the cytoplasm. Notably, the deletion RING domain altered its precise localization and distributed throughout the cytoplasm in GS cells. In EcTRIM32 overexpressing cells, the replication of SGIV or red-spotted grouper nervous necrosis virus (RGNNV) was significantly inhibited compared to the vector control cells. Moreover, the overexpression of EcTRIM32 positively regulated the interferon immune response, evidenced by the significant increase of the expression level of interferon related signaling molecules, including interferon regulatory factor 3 (IRF3), IRF7, interferon-stimulated gene 15 (ISG15), interferon-induced 35-kDa protein (IFP35), MXI, TIR-domain-containing adaptor-inducing interferon-β (TRIF) and melanoma differentiation-associated protein 5 (MDA5). Further studies showed that overexpression of EcTRIM32 significantly enhanced the MDA5-mediated interferon immune response, but decreased stimulator of interferon genes (STING)-mediated interferon immune response. Meanwhile, the expression levels of pro-inflammation cytokines, including TNFα, IL-6 and IL-8 were up-regulated by the ectopic expression of EcTRIM32. We speculated that the regulation of IRF7, and pro-inflammation cytokines by EcTRIM32 overexpression might contribute critical roles in SGIV infection. In addition, the deletion of RING domain not only significantly weakened the antiviral roles of EcTRIM32, but also obviously affected the regulatory effects of EcTRIM32 on interferon immune and inflammation response. Together, our results firstly demonstrated that fish TRIM32 acted as an antiviral factor against both DNA and RNA virus infection.
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, Beijing, China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China.
authors.forename: Yepin
authors.initials: Y
authors.lastname: Yu
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, Beijing, China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China.
authors.forename: Xiaohong
authors.initials: X
authors.lastname: Huang
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, Beijing, China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China.
authors.forename: Jiaxin
authors.initials: J
authors.lastname: Liu
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, Beijing, China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China.
authors.forename: Jingcheng
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, Beijing, China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China.
authors.forename: Yin
authors.initials: Y
authors.lastname: Hu
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, Beijing, China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, Beijing, China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China. Electronic address: huangyh@scsio.ac.cn.
authors.forename: Youhua
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, Beijing, China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China; College of Marine Sciences, South China Agricultural University, Guangzhou 510642, China. Electronic address: qinqw@scsio.ac.cn.
authors.forename: Qiwei
authors.initials: Q
authors.lastname: Qin
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001492:Bass; D003001:Cloning, Molecular; D004266:DNA Virus Infections / Q000235:genetics / Q000276:immunology / Q000662:veterinary* / Q000821:virology; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D005393:Fish Diseases / Q000235:genetics / Q000276:immunology* / Q000821:virology; D029941:Fish Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D007113:Immunity, Innate / Q000235:genetics*; D030381:Nodaviridae / Q000502:physiology; D010802:Phylogeny; D012327:RNA Virus Infections / Q000235:genetics / Q000276:immunology / Q000662:veterinary* / Q000821:virology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017995:Ranavirus / Q000502:physiology; D017386:Sequence Homology, Amino Acid; D000070584:Tripartite Motif Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*
chemical_list: D018076:DNA, Complementary; D029941:Fish Proteins; D012333:RNA, Messenger; D000070584:Tripartite Motif Proteins
keywords: Antiviral; Grouper; Interferon; MDA5; TRIM32