Expression and biological functional analysis of major capsid protein(MCP) of Singapore grouper iridovirus

【Objective】The biological function of major capsid protein(MCP) of Singapore grouper iridovirus(SGIV) was analyzed in this study, which could provide data support and theoretical basis for elucidating the pathogenic mechanism of SGIV infection and developing antiviral products.【Method】Real-time fluorescence quantitative PCR was used to perform the transcriptional analysis of MCP gene, and identify the expression of MCP in the spleen, liver, kidney, intestine, stomach and gill tissues of grouper during SGIV infection. SGIV MCP genes were cloned into eukaryotic expression vectors pEGFP-N3 and pcDNA3.1, respectively, to construct the recombinant plasmids pEGFP-N3-MCP and pcDNA3.1-MCP. Then pEGFP-N3-MCP was transfected into grouper spleen cells(GS) for subcellular localization analysis. At the same time, the recombinant plasmid pcDNA3.1-MCP was transfected into fathead minnow cells(FHM) to construct a FHM cell line(FHM-MCP) capable of stably expressing MCP protein, which was used to analyze the effects of MCP protein on host cell growth and proliferation and effects of on cell survival rates and viral replication during SGIV infection.【Result】The specific transcription of MCP gene could be detected 8 h after SGIV infection, which meaned that MCP gene was a late gene. After 4 h of SGIV infection, the expression of MCP gene was the highest in spleen, followed by liver, kidney and intestine, however, the relative expression levels in stomach and gill tissues were low, suggesting that stomach and gill tissues were not the main target organs for SGIV infection. Subcellular localization results showed that MCP proteins of SGIV were mainly located in the cytoplasm near the nucleus. Both cell growth curve and cell count results were confirmed that MCP protein could regulate cell growth and promote cell proliferation. At 24 and 48 h after SGIV infection, the survival rate of FHM cells overexpressing MCP gene(FHM-MCP) was significantly higher than that of FHM-Vector cells(transfected with eukaryotic expression vector pcDNA3.1), and the survival rate was as 1.12 and 1.15 times as that of FHM-Vector cells, respectively. Furthermore, the virus titers of FHM-MCP cells were higher than FHM-Vector cells 24 and 48 h after SGIV infection, that was, overexpression of MCP protein could promote virus replication after SGIV infection.【Conclusion】SGIV MCP gene is a late gene, and its encoded protein is mainly located in the cytoplasm near the nucleus. By promoting the division and proliferation of host cells and virus replication, it ultimately improves the virus titer and infectivity of SGIV.