The 3′-Terminal Hexamer Sequence of Classical swine fever virus RNA Plays a Role in Negatively Regulating the IRES-Mediated Translation

Abstract

The 3′ untranslated region (UTR) is usually involved in the switch of the translation and replication for a positive-sense RNA virus. To understand the 3′ UTR involved in an internal ribosome entry site (IRES)-mediated translation in Classical swine fever virus (CSFV), we first confirmed the predicted secondary structure (designated as SLI, SLII, SLIII, and SLIV) by enzymatic probing. Using a reporter assay in which the luciferase expression is under the control of CSFV 5′ and 3′ UTRs, we found that the 3′ UTR harbors the positive and negative regulatory elements for translational control. Unlike other stem loops, SLI acts as a repressor for expression of the reporter gene. The negative cis -acting element in SLI is further mapped to the very 3′-end hexamer CGGCCC sequence. Further, the CSFV IRES-mediated translation can be enhanced by the heterologous 3′-ends such as the poly(A) or the 3′ UTR of Hepatitis C virus (HCV). Interestingly, such an enhancement was repressed by flanking this hexamer to the end of poly(A) or HCV 3′ UTR. After sequence comparison and alignment, we have found that this hexamer sequence could hypothetically base pair with the sequence in the IRES IIId1, the 40 S ribosomal subunit binding site for the translational initiation, located at the 5′ UTR. In conclusion, we have found that the 3′-end terminal sequence can play a role in regulating the translation of CSFV.

The Untranslated Regions of Classic Swine Fever Virus RNA Trigger Apoptosis

“…4C). This is related to our previous results showing that SLI of the CSFV 3′UTR repressed expression of a reporter gene, which was further mapped to the hexamer CGGCCC of the 3′ end of SLI possibly forming a base pair with the sequence in the IRES IIId1 of 5′ UTR, the 40 S ribosomal subunit binding site [14]. These results support a possible interaction of SLI of 3′UTR with 5′UTR RNA being involved in the induction of apoptosis.…”

Hepatitis C virus 3′UTR regulates viral translation through direct interactions with the host translation machinery

“…In some plant viruses, 3′-cap-independent translation elements are thought to bind and deliver translation initiation factors to the 5′end of the viral genome through long-range RNA–RNA contacts, thereby stimulating translation initiation (3–6). Similarly, the conserved 3′ untranslated regions (3′ UTRs) of animal RNA viruses including classical swine fever virus (7), foot-and-mouth disease virus (8), dengue virus (9) and hepatitis C virus (HCV) (10) have been reported to regulate viral translation. However, in these cases, the molecular mechanisms by which the 3′UTRs function remain to be elucidated.…”

Functional long-range RNA–RNA interactions in positive-strand RNA viruses

“…Negative regulators of viral IRES activity have also been identified. The genome of the pestivirus classical swine fever virus (CSFV) lacks both a 5ʹ cap and a 3ʹ poly(A) tail, and the 3ʹ UTR inhibits the translational activity of the IRES in the 5ʹ UTR 34 . The negative regulatory sequence that affected IRES activity was mapped to a 3ʹ-terminal RNA hairpin that ends with CGGCCC-OH.…”

“…This terminal sequence was also found to be complementary to a sequence that is located in the ribosomebinding region of the IRES (FIG. 2d), which suggests that a CGGCCC-IRES base-pairing interaction inhibits ribosome recruitment 34 .…”