Functional Analyses of RNA Structures Shared between the Internal Ribosome Entry Sites of Hepatitis C Virus and the Picornavirus Porcine Teschovirus 1 Talfan

Abstract

The internal ribosome entry site (IRES) of porcine teschovirus 1 (PTV-1), a member of the Picornaviridae family, is quite distinct from other well-characterized picornavirus IRES elements, but it displays functional similarities to the IRES from hepatitis C virus (HCV), a member of the Flaviviridae family. In particular, a dominant negative mutant form of eIF4A does not inhibit the activity of the PTV-1 IRES. Furthermore, there is a high level (ca. 50%) of identity between the PTV-1 and HCV IRES sequences. A secondary-structure model of the whole PTV-1 IRES has been derived which includes a pseudoknot. Validation of specific features within the model has been achieved by mutagenesis and functional assays. The differences and similarities between the PTV-1 and HCV IRES elements should assist in defining the critical features of this type of IRES.Porcine teschovirus 1 (PTV-1) Talfan is the prototype member of the Teschovirus genus within the Picornaviridae family. PTV infection results in polioencephalomyelitis in swine, and there have been recent incidents of disease (resulting in paralysis or mortality) in both the United States and Japan (28,42). Multiple serotypes of teschoviruses have been identified, and nearly complete genome sequences are available for a variety of strains (6,14,20,43). The 5Ј-terminal region of the genome sequence is missing in each case, and it is possible that a poly(C) tract is present within the 5Ј untranslated region of the viral RNA in at least some strains (6, 43).Picornavirus genomes are infectious and function as mRNAs. Initiation of protein synthesis on picornavirus RNA is dependent on an internal ribosome entry site (IRES) (see reference 2 for a review). Several different classes of picornavirus IRES element have been described. With the exception of the PTV-1 IRES (see below), they are all large, complex RNA structures of about 450 nucleotides (nt) which contain a polypyrimidine tract located about 20 nt upstream of an AUG codon at the 3Ј end of the element. (Note that in the cardio-and aphthovirus elements, this AUG is an initiation codon, but in the enteroand rhinoviruses, the AUG codon is not usually recognized and initiation occurs at the next AUG codon.) The poliovirus (PV) and human rhinovirus elements represent one class of IRES; these elements function poorly in the rabbit reticulocyte lysate (RRL) in vitro translation system unless it is supplemented with additional proteins (e.g., from HeLa cell extracts). In contrast, the cardio-and aphthovirus IRES elements (e.g., from encephalomyocarditis virus [EMCV] and foot-and-mouth disease virus [FMDV]) do function very efficiently in the standard RRL translation system and they have a secondary structure different from that of the entero-and rhinovirus IRES elements. The hepatitis A virus IRES is distinct again; most notably, it requires the intact translation initiation complex eIF4F (1, 3), whereas the other picornavirus IRES elements function when the eIF4G component of this complex has been cleaved or the cap-binding pro...

Picornavirus IRES elements: RNA structure and host protein interactions

“…Mutations within the loop of domain IIIe strongly inhibited IRES function. Similarly, disruption of the pseudoknot inhibited IRES activity, and conversely, mutants inserting compensatory mutations in S1 and S2 stems displayed enhanced IRES function (Chard et al, 2006b).…”

“…Concerning the HCV-like picornavirus IRES elements, early studies carried out on the PTV-1 IRES showed that the functional element spans only 280 nts (Chard et al, 2006b). A secondary structure model for the PTV-1 IRES (Chard et al, 2006b) predicted domains II and III, but in contrast to the HCV IRES, domain IV was lacking.…”

“…A secondary structure model for the PTV-1 IRES (Chard et al, 2006b) predicted domains II and III, but in contrast to the HCV IRES, domain IV was lacking. In turn, domain III included subdomains IIId and IIIe, and a pseudoknot (Pk) in subdomain IIIf (Fig.…”

Genus-Specific Substitution Rate Variability among Picornaviruses

“…Human picornaviruses produce symptoms ranging from mild respiratory illness to hemorrhagic conjunctivitis, myocarditis, acute flaccid paralysis, and neonatal organ failure (19,27,28,33,40,41). Veterinary picornaviruses, such as foot-and-mouth disease virus (FMDV), encephalomyocarditis virus (EMCV), and porcine teschovirus (PTV), can have devastating effects on livestock (5,8,21).…”

“…Human picornaviruses produce symptoms ranging from mild respiratory illness to hemorrhagic conjunctivitis, myocarditis, acute flaccid paralysis, and neonatal organ failure (19,27,28,33,40,41). Veterinary picornaviruses, such as foot-and-mouth disease virus (FMDV), encephalomyocarditis virus (EMCV), and porcine teschovirus (PTV), can have devastating effects on livestock (5,8,21).Although picornaviruses such as poliovirus (PV) are known to evolve more rapidly than other viruses with single-stranded RNA (ssRNA) genomes (9, 29), little research has been conducted to investigate how they evolve more rapidly than other viruses with similarly error-prone RNA-dependent RNA polymerases (29, 57) or if certain picornaviruses evolve more rapidly than others. Understanding the evolutionary potentials and constraints of these important pathogens is imperative for the development of durable vaccines and effective treatment plans for individual pathogens (42).…”