Free ISG15 Inhibits the Replication of Peste des Petits Ruminants Virus by Breaking the Interaction of Nucleoprotein and Phosphoprotein.

Peste des petits ruminants virus (PPRV) causes a highly contagious disease in small ruminants and severe economic losses in developing countries. PPRV infection can stimulate high levels of interferon (IFN) and many IFN-stimulated genes (ISGs), such as ISG15, which may play a key role in the process of viral infection. However, the role of ISG15 in PPRV infection and replication has not yet been reported. In this study, we found ISG15 expression to be significantly upregulated after PPRV infection of caprine endometrial epithelial cells (EECs), and ISG15 inhibits the proliferation of PPRV. Further analysis showed that free ISG15 could inhibit PPRV proliferation. Moreover, ISG15 does not affect the binding, entry, and transcription but does suppress the replication of PPRV. A detailed analysis revealed that ISG15 interacts and colocalizes with both viral N and P proteins and that its interactive regions are all located in the N-terminal domain. Further studies showed that ISG15 can competitively interact with N and P proteins and significantly interfere with their binding. Finally, through the construction of the C-terminal mutants of ISG15 with different lengths, it was found that amino acids (aa) 77 to 101 play a key role in inhibiting the binding of N and P proteins and that interaction with the P protein disappears after the deletion of 77 to 101 aa. The present study revealed a novel mechanism of ISG15 in disrupting the activity of the N0-P complex to inhibit viral replication. IMPORTANCE PPRV, a widespread and fatal disease of small ruminants, is one of the most devastating animal diseases in Africa, the Middle East, and Asia, causing severe economic losses. IFNs play an important role as a component of natural immunity against pathogens, yet the role of ISG15, an IFN-stimulated gene, in protecting against PPRV infection is currently unknown. We demonstrated, for the first time, that free ISG15 inhibits PPRV proliferation by disrupting the activity of the N0-P complex, a finding that has not been reported in other viruses. Our results provide important insights that can further understand the pathogenesis and innate immune mechanisms of PPRV.

Zinc finger antiviral protein (ZAP) inhibits small ruminant morbillivirus replication in vitro.

Small ruminant morbillivirus (SRMV) is a highly contagious and economically important viral disease of small domestic and wild ruminants. Difficulty with its stable proliferation in ovis aries-derived cells has led to a relative lag in the study of its natural immunity and pathogenesis. Here we report the antiviral properties of ZAP against SRMV, a single-stranded negative-stranded RNA virus of the genus Morbillivirus. ZAP expression was significantly induced in sheep endometrial epithelial cells following SRMV infection. ZAP inhibited SRMV replication in cells after infection, while its overexpression in Vero-SLAM cells significantly increased their resistance to SRMV replication. The ZAP protein co-localized with SRMV RNA in the cytoplasm and ZAP-responsive elements were mapped to the 5' untranslated region of SRMV nucleocapsid, phosphoprotein, matrix, and fusion. In summary, ZAP confers resistance to SRMV infection by directly targeting viral RNA and inhibiting viral replication. Our findings further extend the ranges of viral targets of ZAP and help elucidate the mechanism of SRMV replication.

RPS5 interacts with the rabbit hemorrhagic disease virus 3' extremities region and plays a role in virus replication.

Rabbit hemorrhagic disease virus (RHDV), a member of Caliciviridae family, causes a highly contagious disease in rabbits. The RHDV replication mechanism is poorly understood due to the lack of a suitable culture system in vitro. This study identified RHDV 5' and 3' extremities (Ex) RNA binding proteins from the rabbit kidney cell line RK-13 based on a pull-down assay by applying a tRNA scaffold streptavidin aptamer. Using mass spectrometry (MS), several host proteins were discovered which interact with RHDV 5' and 3' Ex RNA. The ribosomal protein S5 (RPS5) was shown to interact with RHDV 3' Ex RNA directly by RNA-pulldown and confocal microscopy. To further investigate the role of RPS5 in RHDV replication, small interfering RNAs for RPS5 and RPS5 eukaryotic expression plasmids were used to change the expression level of RPS5 in RK-13 cells and the results showed that the RHDV replication and translation levels were positively correlated with the expression level of RPS5. It was also verified that RPS5 promoted RHDV replication by constructing RPS5 stable overexpression cell lines and RPS5 knockdown cell lines. In summary, it has been identified that RPS5 interacted with the RHDV 3' Ex RNA region and played a role in virus replication. These results will help to understand the mechanism of RHDV replication.