The Study Of CSFV Non-coding Region Function

Classical swine fever(Classical swine fever, CSF) is a highly contagious viral infection of pigs and boar, with high pathogenic and high mortality rates. Because of the characteristics, CSF had caused great economic losses to the pig industry in many countries. Therefore, swine fever has been classified as a Class A OIE notifiable infectious diseases.Classical swine fever is a viral infection caused by swine fever virus(Classical swine fever virus, CSFV) of the genus Pestivirus in the family Flaviviridae. CSFV is a positive strand RNA virus, with an outer envelope structure. The diameter of CSFV is about 40 ~ 60 nm, its genome size is about 12.3 kb. Genome structure of CSFV is mainly include of three parts, followed by the 5'untranslated region(5'untranslated region, 5'UTR), poly-protein open reading frame(ORF) capable of encoding approximately 3900 amino acid residues and 3' coding region(3'untranslated region, 3'UTR). 5'UTR and 3'UTR of viral genomic RNA replication and translation have played an important role.According to sequence alignment of the virulent strain Shimen and the attenuated strain HCLV of CSFV, 12 nucleotides insert in the 3'UTR of attenuated strain HCLV between 58-69 sites more than virulent strain Shimen, which is significant differences caused by the intensity of toxicity. Using the Dual-Luciferase Reporter Assay System, 5'UTR and 3'UTR of Shimen and HCLV strains were compared. The results showed that, virulent strain Shimen's 5'UTR and 3'UTR luciferase relative activity were higher, indicating the strength of CSFV toxicity indeed associated with non-coding region.In order to study the mechanism that 3'UTR CSFV in translation process of CSFV regulating, a series of truncated experiments to 3'UTR were proformed. The area of inhibiting translation effect mainly located in the region of nucleotide 80-106, with rich AU, which is designed as the ARE areas. By using biological information technology analysis, we found the potential microRNA ssc-let-7 binding site located at 112-139 points of downstream of the ARE. Our study showed that the ssc-let-7 combination area had no obvious inhibition effort to 3 'UTR, but it inhibitory effect let7 to 3' UTR is significantly enhanced by the ARE. Suggesting that only in present of ARE, ssc-let-7 has the function of synergistic inhibition of CSFV translation.The inhibitory effect of 3'UTR can enhance without ARE area, suggesting that some of the host cell binding proteins for regulating the translation of CSFV. When the HuR protein overexpressed, the inhibition effect of ARE area is significantly reduced. On the other hand, when after intervention of HuR, inhibition effect enhanced. Therefore, after the combination of HuR of host cell and CSFV 3'UTR area, can enhance translation function of CSFV.All in all, 3 'UTR of CSFV can inhibit its translation process in host cells, and ARE area plays a major role in process of virus translation regulating. CSFV can also hire some of the host cell proteins, such as HuR to adjust its protein translation.