Duck hepatitis A virus serotype 1 minigenome: a model for studying the viral 3′UTR effect on viral translation

Abstract

To date, the genetic replication and translation mechanisms as well as the pathogenesis of duck hepatitis A virus type 1 (DHAV-1) have not been adequately characterized due to the lack of a reliable and efficient cell culture system. Although the full-length infections clone system is the best platform to manipulate the virus, it is relatively difficult to assemble this system due to the lack of a suitable cell line. It has been proven that the minigenome system an efficient reverse genetics system for the study of RNA viruses. In some cases, it can be used to displace the infectious clone of RNA viruses. Here, we generated a minigenome for DHAV-1 with two luciferase reporter genes, firefly luciferase (Fluc) and Renilla luciferase (Rluc). The Rluc gene was used as a reference gene for the normalization of the Fluc gene expression in transfected cells, which provided a platform for studying the regulatory mechanisms of DHAV-1. Furthermore, to investigate the role of DHAV-3'UTR in the regulation of viral protein translation, deletions in the 3'UTR were introduced into the DHAV-1 minigenome. Luciferase activity, an indicator of virus translation, was then determined. These results showed that a minigenome system for DHAV-1 was successfully constructed for the first time and that the complete or partial deletion of the DHAV-3'UTR did not affect the expression level of the reporter gene, indicating that DHAV-1 translation may not be modulated by the viral genomic 3'UTR sequence.

The Functional Role of the 3′ Untranslated Region and Poly(A) Tail of Duck Hepatitis A Virus Type 1 in Viral Replication and Regulation of IRES-Mediated Translation

“…However, to PV, the 3′ UTR was not absolutely essential for viral replication, and mutated PV without the complete 3′ UTR only exhibited a moderate deficiency in RNA synthesis ( Brown et al, 2005 ). A previous study also showed that DHAV-1 translation initiation was 3′ UTR-independent ( Liang et al, 2015 ). Nevertheless, in the present study, we showed that viral DHAV-1 viral RNA lacking the complete 3′ UTR was unable to replicate in DEFs ( Figure 4A ).…”

Cleavage of poly(A)-binding protein by duck hepatitis A virus 3C protease

“…DHAV contains the longest 3′UTR in the Picornavirus family. However, the deletion of the 3′UTR had no effect on DHAV IRES-mediated translation initiation with a bicistronic reporter minigenome system 20 .…”

“…Reportedly, the DHAV IRES is distinct from type IV IRESs, although it does share common features with type IV IRES elements of picornaviruses 12 . However, the translation of DHAV might not be modulated by its 3′UTR sequence 20 . In addition, there are three structurally different 2A proteins in the polyprotein of DHAV 21 , including an aphthovirus-like 2A1 22 , a conserved avrRpt2-induced gene (AIGI) protein 2A2 21 , and a parechovirus-like 2A3 23 .…”