Abstract
The NS5A protein of classical swine fever virus (CSFV) is involved in the RNA synthesis and viral replication. However, the NS5A-interacting cellular proteins engaged in the CSFV replication are poorly defined. Using yeast two-hybrid screen, the eukaryotic elongation factor 1A (eEF1A) was identified to be an NS5A-binding partner. The NS5A–eEF1A interaction was confirmed by coimmunoprecipitation, glutathione S-transferase (GST) pulldown and laser confocal microscopy assays. The domain I of eEF1A was shown to be critical for the NS5A–eEF1A interaction. Overexpression of eEF1A suppressed the CSFV growth markedly, and conversely, knockdown of eEF1A enhanced the CSFV replication significantly. Furthermore, eEF1A, as well as NS5A, was found to reduce the translation efficiency of the internal ribosome entry site (IRES) of CSFV in a dose-dependent manner, as demonstrated by luciferase reporter assay. Streptavidin pulldown assay revealed that eEF1A could bind to the CSFV IRES. Collectively, our results suggest that eEF1A interacts with NS5A and negatively regulates the growth of CSFV.
Highlights
Classical swine fever virus (CSFV), together with bovine viral diarrhea virus (BVDV) and border disease virus (BDV), is a member of the genus Pestivirus within the family Flaviviridae and an important pathogen of pigs that often causes huge economic losses to the pig industry worldwide
S-transferase (GST) pulldown assay was performed with the glutathione S-transferase (GST)-tagged NS5A protein expressed in Escherichia coli and the Myc-tagged eukaryotic elongation factor 1A (eEF1A) protein expressed in HEK293T cells
The results showed that GST-NS5A but not GST interacts with eEF1A (Figure 1B)
Summary
Classical swine fever virus (CSFV), together with bovine viral diarrhea virus (BVDV) and border disease virus (BDV), is a member of the genus Pestivirus within the family Flaviviridae and an important pathogen of pigs that often causes huge economic losses to the pig industry worldwide. CSFV is a small enveloped virus with a single-stranded, positive-sense RNA genome of 12.3 kb. The RNA genome contains a single large open reading frame (ORF) coding for a polyprotein, flanked by a 51 -untranslated region (UTR) and a 31 -UTR. Under cellular and viral protease processing, the polyprotein is processed into 11 mature proteins, including four structural proteins (C, Erns , E1 and E2) and seven nonstructural proteins (Npro , p7, NS2-3, NS4A, NS4B, NS5A and NS5B) [1,2,3]. The 51 -UTR contains an internal ribosome entry site (IRES) that is responsible for translation initiation of the viral genome [4].
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
