Abstract
BackgroundIridoviruses are large DNA viruses that cause diseases in fish, amphibians and insects. Singapore grouper iridovirus (SGIV) is isolated from cultured grouper and characterized as a ranavirus. ICP46 is defined to be a core gene of the family Iridoviridae and SGIV ICP46 was demonstrated to be an immediate-early (IE) gene associated with cell growth control and could contribute to virus replication in previous research.MethodsThe transcription start site (TSS) and 5′-untranslated region (5′-UTR) of SGIV ICP46 were determined using 5′ RACE. The core promoter elements of ICP46s were analyzed by bioinformatics analysis. The core promoter region and the regulation model of SGIV ICP46 promoter were revealed by the construction of serially deleted promoter plasmids, transfections, drug treat and luciferase reporter assays. The identification of virion-associated transcriptional transactivator (VATT) that interact with SGIV ICP46 promoter and their binding site on promoter were performed by electrophoretic mobility shift assays (EMSA), DNA pull-down assays and mass spectrometry (MS).ResultsSGIV ICP46 was found to have short 5′-UTR and a presumptive downstream promoter element (DPE), AGACA, which locates at + 36 to + 39 nt downstream of the TSS. The core promoter region of SGIV ICP46 located from − 22 to + 42 nt relative to the TSS. VATTs were involved in the promoter activation of SGIV ICP46 and further identified to be VP12, VP39, VP57 and MCP. A 10-base DNA sequence “ATGGCTTTCG” between the TSS and presumptive DPE was determined to be the binding site of the VATTs.ConclusionOur study showed that four VAATs (VP12, VP39, VP57 and MCP) might bind with the SGIV ICP46 promoter and be involved in the promoter activation. Further, the binding site of the VATTs on promoter was a 10-base DNA sequence between the TSS and presumptive DPE.
Highlights
Iridoviruses are large DNA viruses that cause diseases in fish, amphibians and insects
Our previous study demonstrated that Singapore grouper iridovirus (SGIV) ICP46 is an IE gene associated with cell growth control and contributes to virus replication, the transcription of SGIV ICP46 starts at 2 h p.i. and shows a high transcription level throughout the SGIV infection cycle [11]
We revealed the core region of SGIV ICP46 promoter, investigated the regulation model of SGIV ICP46 promoter associated with viral protein, identified the virion-associated transcriptional transactivator (VATT) that interact with the SGIV ICP46 promoter, and determined the binding site of VATTs on SGIV ICP46 promoter
Summary
Iridoviruses are large DNA viruses that cause diseases in fish, amphibians and insects. ICP46 is defined to be a core gene of the family Iridoviridae and SGIV ICP46 was demonstrated to be an immediate-early (IE) gene associated with cell growth control and could contribute to virus replication in previous research. Iridoviruses are large DNA viruses that cause diseases in fish, amphibians, reptiles and insects, and result in significant economic and ecological losses [1, 2]. Our previous study demonstrated that SGIV ICP46 is an IE gene associated with cell growth control and contributes to virus replication, the transcription of SGIV ICP46 starts at 2 h p.i. and shows a high transcription level throughout the SGIV infection cycle [11]. The promoter sequence and transcriptional regulation of SGIV ICP46 are still unclear
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