Riboproteomics of the Hepatitis C Virus Internal Ribosomal Entry Site

Lu, Henry; Li, Weiqun; Noble, William Stafford; Payan, Donald G.; Anderson, Dave

doi:10.1021/pr0499592

Cited by 74 publications

(60 citation statements)

References 65 publications

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“…Mass spectrometry identified 39 proteins bound to Tob-HCV sORF mRNA. Among others, we found hnRNP C, polypyrimidine tract binding protein 1, and nucleolin, previously described to interact with the HCV 59UTR (Gontarek et al 1999;Ito and Lai 1999;Lu et al 2004). We also identified NF90/NFAR-1, NF45, and RNA helicase A, which interact with both the HCV 59UTR and HCV 39UTR, and function in HCV replication (Isken et al 2007).…”

Section: Resultssupporting

confidence: 66%

“…Interestingly, we also isolated IGF2BP1 that was identified before as an HCV IRES-interacting protein (CRDBP) (Lu et al 2004), but not characterized regarding its function in HCV IRES-mediated translation. A role of IGF2BP1 in the regulation of IGF-II mRNA and b-actin mRNA translation was described before (Nielsen et al 1999;Hüttelmaier et al 2005).…”

Section: Resultsmentioning

confidence: 99%

“…By using an mRNA that bears both the HCV 59UTR and HCV 39UTR as an affinity matrix, we specifically purified proteins that have been shown to interact with either the HCV 59UTR or HCV 39UTR, or both. The proteins IGF2BP1, hnRNP C, hnRNP D, hnRNP Q, polypyrimidine tract binding protein 1, PAI-1 mRNA binding protein 1, nucleolin, NF90/NFAR, RNA helicase A, and five subunits of eIF 3 (out of nine identified) match with proteins isolated from cytoplasmic Huh7 cell extract employing an HCV 59UTR that lacks domain I and represents the HCV IRES consisting of domains II to IV (Kim et al 2004;Lu et al 2004). HnRNP C, YB-1, polypyrimidine tract binding protein 1, DDX3, NF90/NFAR, and RNA helicase A have been identified as HCV 39UTR binding proteins in Huh7 extract (Gontarek et al 1999;Harris et al 2006;Isken et al 2007).…”

Section: Discussionmentioning

confidence: 97%

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IGF2BP1 enhances HCV IRES-mediated translation initiation via the 3′UTR

Weinlich¹,

Hüttelmaier²,

Schierhorn³

et al. 2009

RNA

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The positive-strand RNA genome of the Hepatitis C virus (HCV) contains an internal ribosome entry site (IRES) in the 59untranslated region (59UTR) and structured sequence elements within the 39UTR, but no poly(A) tail. Employing a limited set of initiation factors, the HCV IRES coordinates the 59cap-independent assembly of the 43S pre-initiation complex at an internal initiation codon located in the IRES sequence. We have established a Huh7 cell-derived in vitro translation system that shows a 39UTR-dependent enhancement of 43S pre-initiation complex formation at the HCV IRES. Through the use of tobramycin (Tob)-aptamer affinity chromatography, we identified the Insulin-like growth factor-II mRNA-binding protein 1 (IGF2BP1) as a factor that interacts with both, the HCV 59UTR and 39UTR. We report that IGF2BP1 specifically enhances translation at the HCV IRES, but it does not affect 59cap-dependent translation. RNA interference against IGF2BP1 in HCV replicon RNA-containing Huh7 cells reduces HCV IRES-mediated translation, whereas replication remains unaffected. Interestingly, we found that endogenous IGF2BP1 specifically co-immunoprecipitates with HCV replicon RNA, the ribosomal 40S subunit, and eIF3. Furthermore eIF3 comigrates with IGF2BP1 in 80S ribosomal complexes when a reporter mRNA bearing both the HCV 59UTR and HCV 39UTR is translated. Our data suggest that IGF2BP1, by binding to the HCV 59UTR and/or HCV 39UTR, recruits eIF3 and enhances HCV IRES-mediated translation.

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Section: Resultssupporting

confidence: 66%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 97%

See 1 more Smart Citation

IGF2BP1 enhances HCV IRES-mediated translation initiation via the 3′UTR

Weinlich¹,

Hüttelmaier²,

Schierhorn³

et al. 2009

RNA

View full text Add to dashboard Cite

show abstract

“…Proteins that remained binding to the RNA under stringent conditions were subsequently analyzed by SDS-PAGE and silver staining and, following extraction and digestion with trypsin, identified by mass spectrometry analysis (see Materials and Methods). Thus, among others, we identified the proteins hnRNP C, polypyrimidine tract binding (PTB) protein, and nucleolin, which were earlier reported to bind to the HCV NTRs (Gontarek et al 1999;Ito and Lai 1999;Lu et al 2004). Most interestingly, we also found the NF/ NFAR proteins NF90/NFAR-1, NF45, and RNA helicase A (RHA) specifically binding to the HCV NTRs (Fig.…”

Section: Nf/nfar Proteins Specifically Bind To the Hcv 59-and 39ntrsupporting

confidence: 53%

Nuclear factors are involved in hepatitis C virus RNA replication

Isken¹,

Baroth²,

Grassmann³

et al. 2007

RNA

145

171

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Unraveling the molecular basis of the life cycle of hepatitis C virus (HCV), a prevalent agent of human liver disease, entails the identification of cell-encoded factors that participate in the replication of the viral RNA genome. This study provides evidence that the so-called NF/NFAR proteins, namely, NF90/NFAR-1, NF110/NFAR-2, NF45, and RNA helicase A (RHA), which mostly belong to the dsRBM protein family, are involved in the HCV RNA replication process. NF/NFAR proteins were shown to specifically bind to replication signals in the HCV genomic 59 and 39 termini and to promote the formation of a looplike structure of the viral RNA. In cells containing replicating HCV RNA, the generally nuclear NF/NFAR proteins accumulate in the cytoplasmic viral replication complexes, and the prototype NFAR protein, NF90/NFAR-1, stably interacts with a viral protein.HCV replication was inhibited in cells where RNAi depleted RHA from the cytoplasm. Likewise, HCV replication was hindered in cells that contained another NF/NFAR protein recruiting virus. The recruitment of NF/NFAR proteins by HCV is assumed to serve two major purposes: to support 59-39 interactions of the viral RNA for the coordination of viral protein and RNA synthesis and to weaken host-defense mechanisms.

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“…Several cellular RNA binding proteins have been identified by proteomics that could be involved in this process and in different steps of HCV replication. For instance, 26 host proteins were found to specifically interact with the IRES (Lu et al 2004) and more than 70 cellular factors were identified to interact with the 3 ′ UTR of the plusstrand . Accordingly, it has been speculated that a RNA circularization could be a general replication mechanism for all plus-strand RNA viruses (Herold and Andino 2001).…”

Section: Possible Circularization Of Hcvmentioning

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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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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Riboproteomics of the Hepatitis C Virus Internal Ribosomal Entry Site

Cited by 74 publications

References 65 publications

IGF2BP1 enhances HCV IRES-mediated translation initiation via the 3′UTR

IGF2BP1 enhances HCV IRES-mediated translation initiation via the 3′UTR

Nuclear factors are involved in hepatitis C virus RNA replication

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

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