Duck enteritis virus UL54 is an IE protein primarily located in the nucleus

Abstract

BackgroundThe UL54 protein of Duck Enteritis Virus (DEV) is a homolog of herpes simplex virus-1 (HSV-1) immediate-early infectious cell protein 27 (ICP27), a multifunctional protein essential for viral infection. Nonetheless, there is little information on the UL54 protein of DEV.MethodsThe UL54 gene was cloned into the pPAL7 vector, and the recombinant protein, expressed in the E. coli Rosetta, was used to produce a specific antibody. Using this antibody, Western blotting and indirect immunofluorescence analysis (IFA) were used to analyze the expression level and intracellular localization, respectively, of UL54 in DEV-infected cells at different times. Real-time quantitative reverse transcription PCR (RT-PCR) and the pharmacological inhibition test were utilized to ascertain the kinetic class of the UL54 gene.ResultsUL54 was expressed as a fusion protein of approximately 66.0 kDa using the prokaryotic expression system, and this protein was used to generate the specific anti-UL54 antibody. The UL54 protein was initially diffusely distributed throughout the cytoplasmic region; then, after 2 h, it gradually distributed into the nucleus, peaking at 24 h, and complete localization to the nucleus was observed thereafter. The UL54 transcript was detected as early as 0.5 h, and peak expression was observed at 24 h. The UL54 gene was insensitive to the DNA polymerase inhibitor Ganciclovir (GCV) and the protein synthesis inhibitor Cycloheximide (CHX), both of which confirmed that UL54 was an immediate early gene.ConclusionsThe DEV UL54 gene was expressed in a prokaryotic expression system and characterized for expression level, intracellular localization and gene kinetic class. We propose that these results will provide the foundation for further functional analyses of this gene.

Duck virus enteritis (duck plague) – a comprehensive update

“…Researchers have prepared oil emulsified inactivated vaccine against Salmonella Typhimurium (ST), DP and Duck Hepatitis Virus (DHV) which was found safe, potent and capable of protecting ducklings against ST, DP and DHV infections effectively (Hanan et al 2014;. Researchers are investigating the role of UL16 and UL54 genes also and their expressed proteins from DEV, so that they can also be utilized in planning prophylactic strategy against DEV virus in susceptible bird populations (He et al 2012;Liu et al 2015;Liu, Cheng, et al 2016). A DEV-vectored vaccine containing a recombinant DEV (rDEV-ΔUL2-HA) inserted with the HA gene from duck-origin H9 subtype (H9N2) AIV into the UL2 gene by homologous recombination has been developed recently.…”

Duck enteritis virus UL21 is a late gene encoding a protein that interacts with pUL16

“…IE genes are expressed in the presence of CHX although the transcription of E and L genes is suppressed [31]. We also treated DEV-infected cells with GCV and examined UL54 (an IE gene) [32], US2 (an L gene) [33], UL13 (an E gene) [34], and β-actin as controls. We found that UL21 gene expression was inhibited in the groups treated with GCV and CHX.…”

Molecular characterization of the duck enteritis virus US10 protein

“…The reactivity of the antibody was determined using western blot analysis [ 18 ]. On the one hand, the lysates of bacteria transformed with pET-32a(+) vs. pET-32a(+)/US10 were blotted to test the cross-reactivity of anti-US10 serum.…”

“…IFA was conducted using a standard procedure [ 18 ]. Briefly, DEV-infected cells were harvested at 12, 24, 36, and 48 h post-infection, plated onto coverslips, and fixed with 4% paraformaldehyde for 30 min.…”