Hepatitis C virus IRES efficiency is unaffected by the genomic RNA 3′NTR even in the presence of viral structural or non-structural proteins

Abstract

Hepatitis C virus (HCV) translation is mediated by an IRES structure. Instead of a poly(A) tail, the 39 end of the genome contains a tripartite 39NTR composed of a non-conserved region, a polypyrimidine tract and a highly conserved stretch of 98 nt, termed the 39X region. Using a set of bicistronic recombinant DNA constructs expressing two reporter genes separated by the HCV IRES, it was determined whether the HCV 39NTR sequence, in the presence or absence of HCV proteins, played a role in the efficiency of HCV IRES-dependent translation ex vivo. Bicistronic expression cassettes were transfected into hepatic and non-hepatic cell lines. These results show that neither the entire 39NTR nor the 39X sequence alters IRES-dependent translation efficiency, whatever the cell line tested. A potential effect of the 39NTR on IRES-dependent translation in the presence of HCV proteins was investigated further. Neither non-structural nor structural HCV proteins had any effect on the efficiency of IRES in this system. In addition, in order to mimic HCV genome organization, monocistronic expression cassettes containing the IRES and a Core-DsRed fusion gene were constructed with or without the 39NTR. In this context, no effect of the 39NTR on IRES translation efficiency was observed, even in the presence of HCV proteins. These data demonstrate that HCV translation is not modulated by the viral genomic 39NTR sequence, even in the presence of HCV structural or non-structural proteins.

The Hepatitis C Virus RNA 3′-Untranslated Region Strongly Enhances Translation Directed by the Internal Ribosome Entry Site

“…Conflicting results have been reported on the effect of the HCV 3Ј-UTR on IRES-directed translation (9,10,15,19,20,26,34,35,37,56). The reasons for these discrepancies are not directly evident.…”

“…However, a controversial situation arose from the results obtained up to now. Some studies reported a stimulatory effect of the 3Ј-UTR on HCV translation (19,20,34,35); others showed that the HCV 3Ј-UTR has no stimulatory influence on HCV IRES-directed translation (9,10,15,26,56). One study even reported an inhibitory effect of the 3Ј-UTR (37).…”

“…Since HCV replicates primarily in human liver cells, several groups examined a possible effect of the HCV 3Ј-UTR on RNA translation by using transfected human hepatoma cell lines. However, results are controversial (10,15,20,26,34). Most of these studies used transfection and transient expression of complete circular or linearized reporter plasmids.…”

“…In some studies, a positive influence of 3Ј-UTR sequences (sometimes comprising only the 3Ј X region) was shown (19,20,34,35). One study reported an inhibitory effect of the 3Ј-UTR (37), whereas other reports claimed that there is no effect of the HCV 3Ј-UTR on IRES-directed translation (9,10,15,26,56).…”

A long-range RNA–RNA interaction between the 5′ and 3′ ends of the HCV genome

“…This is the case of flavivirus , certain picornaviruses, such as FMDV (Serrano et al 2006), retroviruses (Ooms et al 2007;Kenyon et al 2008), or Dengue virus (Polacek et al 2009), among others. With respect to HCV, conflicting results regarding the role of the 39 UTR in viral protein synthesis have been reported (Ito et al 1998;Ito and Lai 1999;Fang and Moyer 2000;Michel et al 2001;Murakami et al 2001;Kong and Sarnow 2002;McCaffrey et al 2002;Imbert et al 2003;Bradrick et al 2006;Song et al 2006;Lourenco et al 2008). Today it is commonly accepted that the 39 end exerts an enhancer translational effect through the recruitment of viral and cellular protein factors that stimulate IRES activity (McCaffrey et al 2002;Bradrick et al 2006;Song et al 2006;Lourenco et al 2008).…”