IGF2BP1 Significantly Enhances Translation Efficiency of Duck Hepatitis A Virus Type 1 without Affecting Viral Replication

Abstract

As a disease characterized by severe liver necrosis and hemorrhage, duck viral hepatitis (DVH) is mainly caused by duck hepatitis A virus (DHAV). The positive-strand RNA genome of DHAV type 1 (DHAV-1) contains an internal ribosome entry site (IRES) element within the 5′ untranslated region (UTR), structured sequence elements within the 3′ UTR, and a poly(A) tail at the 3′ terminus. In this study, we first examined that insulin-like growth factor-2 mRNA-binding protein-1 (IGF2BP1) specifically interacted with the DHAV-1 3′ UTR by RNA pull-down assay. The interaction between IGF2BP1 and DHAV-1 3′ UTR strongly enhanced IRES-mediated translation efficiency but failed to regulate DHAV-1 replication in a duck embryo epithelial (DEE) cell line. The viral propagation of DHAV-1 strongly enhanced IGF2BP1 expression level, and viral protein accumulation was identified as the key point to this increment. Collectively, our data demonstrated the positive role of IGF2BP1 in DHAV-1 viral proteins translation and provided data support for the replication mechanism of DHAV-1.

In vivo structural characterization of the SARS-CoV-2 RNA genome identifies host proteins vulnerable to repurposed drugs

“…These 11 RBPs include proteins that are known to regulate virus life cycles. For example, IGF2BP1 facilitates the translation of HCV (Weinlich et al, 2009) and duck hepatitis A virus (DHAV) (Chen et al, 2019) from the internal ribosome entry sites (IRES). In addition, our predicted list shows significant overlap with the proteins detected as SARS-CoV-2 interactors by RNA antisense purification (Schmidt et al, 2020) (p = 3.0e-5, one-sided Fisher's exact test) as well as interactors by ribonucleoprotein capture (Lee et al, 2020) (p = 1.5e-11, onesided Fisher's exact test).…”

Interleukin-2 enhancer binding factor 2 negatively regulates the replication of duck hepatitis A virus type 1 by disrupting the RNA-dependent RNA polymerase activity of 3D polymerase

“…The isolated products were analyzed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF–MS). The MALDI-TOF–MS results indicated numerous cellular proteins that might interact with 3'-UTR of DHAV-1, and ILF2 was chosen for further analysis due to its high index [ 30 ]. Next, an in vitro RNA pull-down assay was performed using the biotin-labeled RNA probe of the 3′-UTR and the purified ILF2-His fusion protein to investigate whether ILF2 specifically interacted with the 3′-UTR of DHAV-1, as described in a previous study [ 30 ].…”

“…The MALDI-TOF–MS results indicated numerous cellular proteins that might interact with 3'-UTR of DHAV-1, and ILF2 was chosen for further analysis due to its high index [ 30 ]. Next, an in vitro RNA pull-down assay was performed using the biotin-labeled RNA probe of the 3′-UTR and the purified ILF2-His fusion protein to investigate whether ILF2 specifically interacted with the 3′-UTR of DHAV-1, as described in a previous study [ 30 ]. The biotin-labeled RNA probe of the reverse sequence of the 3'-UTR and purified ILF2-His fusion protein were used as the negative control.…”

“…The in vitro RNA pull-down assay was performed as described previously [ 30 ]. In brief, the Pierce magnetic RNA–protein pull-down kit (Thermo Fisher Scientific, Waltham, MA, USA) was used to perform the in vitro RNA pull-down assay.…”

“…The recombinant plasmid pDHAV-3′UTR-A 25 containing the T7 promoter, the 5′—UTR of DHAV-1, the Firefly luciferase gene, and the 3′—UTR and poly(A) 25 tail of DHAV-1 was constructed [ 30 ]. The recombinant plasmid pDHAV-3′UTR-A 25 was digested with restriction enzyme XhoI (TaKaRa, Dalian, China) and gel purified, and then used for in vitro transcription using the T7 RiboMAX Express large-scale RNA production system (TaKaRa).…”