Fish miR-146 a promotes Singapore grouper iridovirus infection by regulating cell apoptosis and NFk B activation

miR-146a was reported to participate in various pathophysiological conditions in mammals, such as inflammation and immune responses, oncogenesis and cell damage. However, its function in low vertebrates has not been well elucidated. In this study, we characterized the expression profiles and functions of miR-146a in fish cells during iridovirus infection. We found that the reported fish miR-146a genes encoded an identical mature sequence, which shared high similarity with its mammalian orthologues, suggesting a putative functional conservation of miR-146a between fish and other vertebrates. Using a well-established infection model of Singapore grouper iridovirus (SGIV) in fathead minnow cells, we found that SGIV infection induced the expression of miR-146a to a dramatic extent. More importantly, we found that miR-146a promoted SGIV propagation, as demonstrated by higher expression of viral genes and increased virus titres in miR-146aoverexpressing cells. Mechanistically, we found that miR-146a overexpression suppressed, while miR-146a knockdown promoted, NF-kB activation and SGIV-induced cell apoptosis, two major cellular events involved in SGIV infection. Our study suggested that the induction of miR-146a by SGIV infection may function through a feed-forward mechanism to promote viral infection by restraining anti-viral cellular responses. INTRODUCTION microRNA (miRNA) is a class of small non-coding RNAs (ncRNAs), 18–25 nucleotides in length, which post-transcriptionally regulates gene expression through binding to the 3¢UTRs (untranslated regions) of target mRNAs [1, 2]. Since its first discovery in the late 1990s, there has been increasing evidence demonstrating that this class of ncRNAs participates in various biological processes, including cell proliferation, differentiation, oncogenesis, immuno-inflammatory responses and cell apoptosis [3–7]. miR-146a is one of the first reported miRNAs to negatively regulate the immune signalling pathways [8–10]. For example, in chronic hepatitis B miR-146 is induced significantly, which causes hypo-responsiveness in T-cells by targeting STAT1, a key mediator of IFN-a signalling activation, and thus results in immune defects and immunopathogenesis during chronic viral infection [11]. Park et al. found that miR-146a actively participated in the determination of the fate of dendritic cells (DCs), the key cell type responsible for the activation of lymphocytes to fight against exogenous infection as well as subsequent restriction of excessive autoimmunity. They found that during the differentiation of human monocytes from DCs, the upregulation of miR146a can repress the expression of Bcl-2, which in turn promotes DC apoptosis [12]. While most previous studies concerning miR-146a have focused on its regulation in immuno-inflammatory responses and cell fate determination during tumour development, its roles in viral infection and virus–cell interactions remain largely unclear. Singapore grouper iridovirus (SGIV) is a high pathogenic double-stranded DNA virus isolated from cultural groupers [13]. It belongs to genus Ranavirus, family Iridoviridae, and has a broad infectivity in various cell types of different grouper tissues. After infection, SGIV can cause high mortality and it has caused huge economic losses to the aquaculture industry in China and Southeast Asia [14]. In the last decade, extensive studies have been performed to characterize the pathogenic viral genes related to SGIV infection, and these have greatly improved our understanding of the molecular characteristics of this viral pathogen [15–17]. However, the molecular mechanisms underlying this viral pathogen and the host/cell interactions still require extensive investigation. Received 16 November 2016; Accepted 13 April 2017 Author affiliations: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou, 510301, PR China; University of Chinese Academy of Sciences, Beijing, 100049, PR China; School of Medicine, University of Alabama at Birmingham, Birmingham, AL 35294, USA; College of Marine Sciences, South China Agricultural University, Guangzhou 510642, PR China; Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266000, PR China. *Correspondence: Qiwei Qin, qinqw@scsio.ac. cn