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

Romero-López, Cristina; Berzal‐Herranz, Alfredo

doi:10.1261/rna.1680809

Cited by 112 publications

(160 citation statements)

References 83 publications

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“…We confirm the secondary structure of SLIIId as previously described and validated by experimental data (Supplemental Fig. 1A; Brown et al 1992;Kolupaeva et al 2000;Lukavsky et al 2000;Odreman-Macchioli et al 2000;Romero-López and Berzal-Herranz 2009;Pang et al 2011;Hajdin et al 2013). These secondary structures are additionally stabilized by an LRI introduced in Romero-López and Berzal-Herranz (2012) (No.…”

supporting

confidence: 89%

“…Some cryo-electron microscopy analyses do not provide sufficient resolution to unambiguously identify the known SLIIId structure (Spahn et al 2001;Boehringer et al 2005;Siridechadilok et al 2005). However, experimental data strongly argue for the classical known SLIIId structure to be present at least when the IRES RNA is in solution (Brown et al 1992;OdremanMacchioli et al 2000;Romero-López and Berzal-Herranz 2009;Pang et al 2011;Hajdin et al 2013), and nuclease footprints in the apical loop of the classical version of SLIIId disappear upon binding of the small 40S ribosomal subunit (Kolupaeva et al 2000;Lukavsky et al 2000). Moreover, the classical version of SLIIId is strongly supported by data showing a four-way junction including SLIII, SLIIIe, and SLIIIf (Berry et al 2011;Yamamoto et al 2014).…”

Section: Resultsmentioning

confidence: 99%

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Conserved RNA secondary structures and long-range interactions in hepatitis C viruses

Fricke

Dünnes

Zayas

et al. 2015

RNA

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Hepatitis C virus (HCV) is a hepatotropic virus with a plus-strand RNA genome of ∼9.600 nt. Due to error-prone replication by its RNA-dependent RNA polymerase (RdRp) residing in nonstructural protein 5B (NS5B), HCV isolates are grouped into seven genotypes with several subtypes. By using whole-genome sequences of 106 HCV isolates and secondary structure alignments of the plus-strand genome and its minus-strand replication intermediate, we established refined secondary structures of the 5 ′ untranslated region (UTR), the cis-acting replication element (CRE) in NS5B, and the 3 ′ UTR. We propose an alternative structure in the 5 ′ UTR, conserved secondary structures of 5B stem-loop (SL)1 and 5BSL2, and four possible structures of the X-tail at the very 3 ′ end of the HCV genome. We predict several previously unknown long-range interactions, most importantly a possible circularization interaction between distinct elements in the 5 ′ and 3 ′ UTR, reminiscent of the cyclization elements of the related flaviviruses. Based on analogy to these viruses, we propose that the 5 ′ -3 ′ UTR base-pairing in the HCV genome might play an important role in viral RNA replication. These results may have important implications for our understanding of the nature of the cis-acting RNA elements in the HCV genome and their possible role in regulating the mutually exclusive processes of viral RNA translation and replication.

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supporting

confidence: 89%

Section: Resultsmentioning

confidence: 99%

Conserved RNA secondary structures and long-range interactions in hepatitis C viruses

Fricke

Dünnes

Zayas

et al. 2015

RNA

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“…The HCV genome contains well-defined RNA structural elements in its UTRs. Previous studies demonstrated that the protein-independent interaction of domain IIId and short stem-loop 5BSL3.2 is essential for replication (55). Also, previously reported data indicated the participation of different cellular factors in inducing interactions between the HCV 5= and 3= UTRs (38,56).…”

Section: Discussionmentioning

confidence: 93%

Human La Protein Interaction with GCAC near the Initiator AUG Enhances Hepatitis C Virus RNA Replication by Promoting Linkage between 5′ and 3′ Untranslated Regions

Kumar

Ray

2013

J Virol

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Human La protein has been implicated in facilitating the internal initiation of translation as well as replication of hepatitis C virus (HCV) RNA. Previously, we demonstrated that La interacts with the HCV internal ribosome entry site (IRES) around the GCAC motif near the initiator AUG within stem-loop IV by its RNA recognition motif (RRM) (residues 112 to 184) and influences HCV translation. In this study, we have deciphered the role of this interaction in HCV replication in a hepatocellular carcinoma cell culture system. We incorporated mutation of the GCAC motif in an HCV monocistronic subgenomic replicon and a pJFH1 construct which altered the binding of La and checked HCV RNA replication by reverse transcriptase PCR (RT-PCR). The mutation drastically affected HCV replication. Furthermore, to address whether the decrease in replication is a consequence of translation inhibition or not, we incorporated the same mutation into a bicistronic replicon and observed a substantial decrease in HCV RNA levels. Interestingly, La overexpression rescued this inhibition of replication. More importantly, we observed that the mutation reduced the association between La and NS5B. The effect of the GCAC mutation on the translation-to-replication switch, which is regulated by the interplay between NS3 and La, was further investigated. Additionally, our analyses of point mutations in the GCAC motif revealed distinct roles of each nucleotide in HCV replication and translation. Finally, we showed that a specific interaction of the GCAC motif with human La protein is crucial for linking 5= and 3= ends of the HCV genome. Taken together, our results demonstrate the mechanism of regulation of HCV replication by interaction of the cis-acting element GCAC within the HCV IRES with human La protein. Hepatitis C virus (HCV), a major etiologic agent of posttransfusion and sporadic non-A, non-B hepatitis (NANBH), belongs to the Hepacivirus genus of the Flaviviridae family (1, 2). The HCV genome consists of a 9.6-kb single-stranded positive-sense RNA containing an open reading frame (ORF) encoding an ϳ3,000-residue polyprotein precursor flanked at both ends by highly structured and conserved untranslated regions (UTRs) (1-4). HCV translation is mediated by an internal ribosome entry site (IRES) located mostly within the 341-nucleotide (nt) 5= UTR and extending to 30 to 40 nucleotides downstream of the initiator AUG (iAUG) codon. Sequence elements that are directly involved in HCV replication are located mostly in the 3= UTR (1, 4-6). The HCV 3= UTR varies between 200 and 235 nt in length, including a short variable region, a poly(U/UC) tract (with an average length of 80 nt), and an invariant 98-nt X-tail region (7-9). Initiation of HCV replication takes place by formation of a ribonucleoprotein (RNP) complex at the 3= UTR of the viral genome. The newly synthesized negative-strand RNA then serves as a template for the production of the plus-strand copies of viral RNA (10, 11).Some reports have revealed the presence of cis-acting elements in ...

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“…In the case of the kl-TSS, ribosomes/subunits are then transferred to the 5= end through the kl-TSS's long-distance kissing-loop interaction that mimics the closed loop topology of canonical mRNAs (28). Maintaining template association with ribosomal subunits following translation termination should enhance protein synthesis by increasing the rate of reinitiation, as was recently shown for a 40S binding element in the 3=UTR of hepatitis C virus (49), which would be positioned proximal to the 5= IRES through nearby long-distance RNA-RNA interactions (50).…”

Section: Discussionmentioning

confidence: 96%

The 3′ Untranslated Region of Pea Enation Mosaic Virus Contains Two T-Shaped, Ribosome-Binding, Cap-Independent Translation Enhancers

Gao

Kasprzak

Szarko

et al. 2014

J Virol

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Many plant viruses without 5=caps or 3= poly(A) tails contain 3= proximal, cap-independent translation enhancers (3=CITEs) that bind to ribosomal subunits or translation factors thought to assist in ribosome recruitment. Most 3=CITEs participate in a longdistance kissing-loop interaction with a 5= proximal hairpin to deliver ribosomal subunits to the 5= end for translation initiation. Pea enation mosaic virus (PEMV) contains two adjacent 3=CITEs in the center of its 703-nucleotide 3= untranslated region (3=UTR), the ribosome-binding, kissing-loop T-shaped structure (kl-TSS) and eukaryotic translation initiation factor 4E-binding Panicum mosaic virus-like translation enhance (PTE). We now report that PEMV contains a third, independent 3=CITE located near the 3= terminus. This 3=CITE is composed of three hairpins and two pseudoknots, similar to the TSS 3=CITE of the carmovirus Turnip crinkle virus (TCV). As with the TCV TSS, the PEMV 3=TSS is predicted to fold into a T-shaped structure that binds to 80S ribosomes and 60S ribosomal subunits. A small hairpin (kl-H) upstream of the 3=TSS contains an apical loop capable of forming a kissing-loop interaction with a 5= proximal hairpin and is critical for the accumulation of full-length PEMV in protoplasts. Although the kl-H and 3=TSS are dispensable for the translation of a reporter construct containing the complete PEMV 3=UTR in vitro, deleting the normally required kl-TSS and PTE 3=CITEs and placing the kl-H and 3=TSS proximal to the reporter termination codon restores translation to near wild-type levels. This suggests that PEMV requires three 3=CITEs for proper translation and that additional translation enhancers may have been missed if reporter constructs were used in 3=CITE identification. IMPORTANCEThe rapid life cycle of viruses requires efficient translation of viral-encoded proteins. Many plant RNA viruses contain 3= capindependent translation enhancers (3=CITEs) to effectively compete with ongoing host translation. Since only single 3=CITEs have been identified for the vast majority of individual viruses, it is widely accepted that this is sufficient for a virus's translational needs. Pea enation mosaic virus possesses a ribosome-binding 3=CITE that can connect to the 5= end through an RNA-RNA interaction and an adjacent eukaryotic translation initiation factor 4E-binding 3=CITE. We report the identification of a third 3=CITE that binds weakly to ribosomes and requires an upstream hairpin to form a bridge between the 3= and 5= ends. Although both ribosome-binding 3=CITEs are critical for virus accumulation in vivo, only the CITE closest to the termination codon of a reporter open reading frame is active, suggesting that artificial constructs used for 3=CITE identification may underestimate the number of CITEs that participate in translation.

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

Cited by 112 publications

References 83 publications

Conserved RNA secondary structures and long-range interactions in hepatitis C viruses

Conserved RNA secondary structures and long-range interactions in hepatitis C viruses

Human La Protein Interaction with GCAC near the Initiator AUG Enhances Hepatitis C Virus RNA Replication by Promoting Linkage between 5′ and 3′ Untranslated Regions

The 3′ Untranslated Region of Pea Enation Mosaic Virus Contains Two T-Shaped, Ribosome-Binding, Cap-Independent Translation Enhancers

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