Objective:To study the role and possible molecular mechanism of neural precursor cell-expressed developmentally down-regulated gene 4-like ( NEDD4 L) in the replication of hepatitis B virus (HBV). Methods:Small interfering RNA (siRNA) targeting NEDD4 L, plasmid expressing NEDD4 L with hemagglutinin(HA) C-terminal tag (pcDNA3.1- NEDD4 L-HA), plasmid expressing 1.3×HBV genome (pGEM-HBV1.3) and poly (dAT: dAT) were respectively transfected into HepG2 cells using Lipofectamine2000. HepG2.2.15 cells, a cell line that can stably express HBV, were used as control. The mRNA levels of NEDD4 L, interferon (IFN)-α, IFN-β, interferon-stimulated gene 56 ( ISG56), myxovirus resistance protein A ( MxA), oligoadenylate synthetase ( OAS), and the levels of HBV DNA or 3.5 kb HBV RNA were detected by real-time fluorescent quantitative polymerase chain reaction (qRT-PCR). Western blot was used to detect the silence and over-expression of NEDD4 L, and the protein levels of the related signaling molecules. The amount of IFN-β in the cellular supernatant was measured by enzyme linked immunosorbent assay (ELISA). Student t test was used for comparison of continuous data between groups. Results:The levels of NEDD4L mRNA and protein in HepG2.2.15 cells were 10.53±0.47 and 4.17±0.43, respectively, which were both statistically higher than those in HepG2 cells (1.00±0.05, t=3.27, P=0.008 and 1.26±0.25, t=1.68, P=0.030, respectively). In HepG2 cells with knockdown of NEDD4 L, the expression level of HBV DNA in cellular supernatant was 0.32±0.09, which was statistically lower than that in the control (1.00±0.05, t=-0.93, P=0.020), and the expression level of 3.5 kb HBV RNA was 0.49±0.11, which was statistically lower than that in the control (1.00±0.05, t=-0.68, P=0.040), while the mRNA levels of IFN-β and downstream effector molecules ( ISG56, MxA and OAS) were all significantly increased compared with the control ( t=4.66, 9.38, 7.29 and 7.01, respectively, all P<0.01). With poly (dAT: dAT) treatment and vesicular stomatitis virus (VSV) stimulation, the levels of IFN-β in HepG2 cells with knockdown of NEDD4 L were (776.41±115.49) ng/L and (961.21±130.19) ng/L, respectively, which were both statistically higher than those of the control group ((320.15± 56.05) ng/L, t=2.43, P=0.020; (440.17±67.82) ng/L, t=2.85, P=0.030, respectively). With poly (dAT: dAT) treatment and VSV stimulation, the levels of IFN-β in HepG2 cells with overexpression of NEDD4 L were (156.18±26.47) ng/L and (176.67±34.51) ng/L, respectively, which were both statistically lower than those of the control group ((320.38±49.39) ng/L, t=-2.03, P=0.040; (440.59±68.83) ng/L, t=-1.93, P=0.030, respectively). Western blot showed that the replication of HBV reduced the protein level of melanoma differentiation-associated protein 5 (MDA5), a key molecule in upstream of IFN-β, but the down-regulation was not obvious in cells with the knockdown of NEDD4 L. Conclusion:The replication of HBV could promote the up-regulation of NEDD4L protein and subsequently reduce the protein level of MDA5, thereby inhibiting the production of IFN-β, which facilitates HBV to escape the innate immune response.

Enterovirus (EV) includes a group of important RNA viruses with similarity in viral structure and pathogenesis. They can cause various diseases in humans and pose a great threat to human health. The biological events that occur in EV-host interactions are the core topics to be investigated for better understanding the pathogenesis of EV and developing related prevention and treatment strategies. With the development of genomics technologies, RNA modifications and protein post-translational modifications (PTM) in EV and its hosts have drawn much attention and some achievements have been made, which may shed light on developing precise strategies for the prevention and control of EV-associated diseases. In this paper, to better understand the pathogenesis of EV infection and provide reference for clinical translational research, the progress in the roles of RNA modifications and protein PTM in EV-host interactions are summarized.