HCV and CSFV IRES domain II mediate eIF2 release during 80S ribosome assembly

Locker, Nicolas; Easton, Laura E.; Lukavsky, Peter J.

doi:10.1038/sj.emboj.7601549

Cited by 132 publications

(195 citation statements)

References 52 publications

(117 reference statements)

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“…Following elution of the complexes, the presence of initiation factors was analyzed by Western blotting. As expected and in agreement with previous studies (Locker et al 2007), complexes assembled onto the HCV IRES contained eIF3 but not eIF4E or eIF4G (Fig. 4B).…”

Section: Inactivation Of Eif4e Inhibits Vflip Ires Activitysupporting

confidence: 80%

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Functional analysis of Kaposi's sarcoma–associated herpesvirus vFLIP expression reveals a new mode of IRES-mediated translation

Othman

Sulaiman

Willcocks

et al. 2014

RNA

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Kaposi's sarcoma-associated herpesvirus (KSHV) is an oncogenic virus, the etiological agent of Kaposi's sarcoma (KS) and primary effusion lymphoma (PEL). One of the key viral proteins that contributes to tumorigenesis is vFLIP, a viral homolog of the FLICE inhibitory protein. This KSHV protein interacts with the NFκB pathway to trigger the expression of antiapoptotic and proinflammatory genes and ultimately leads to tumor formation. The expression of vFLIP is regulated at the translational level by an internal ribosomal entry site (IRES) element. However, the precise mechanism by which ribosomes are recruited internally and the exact location of the IRES has remained elusive. Here we show that a 252-nt fragment directly upstream of vFLIP, within a coding region, directs translation. We have established its RNA structure and demonstrate that IRES activity requires the presence of eIF4A and an intact eIF4G. Furthermore, and unusually for an IRES, eIF4E is part of the complex assembled onto the vFLIP IRES to direct translation. These molecular interactions define a new paradigm for IRES-mediated translation.

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Section: Inactivation Of Eif4e Inhibits Vflip Ires Activitysupporting

confidence: 80%

“…www.rnajournal.org 1807 recruitment, while we previously demonstrated that a loop structure within domain II mediates the 60S subunit joining event (Locker et al 2007). Therefore, a deep understanding of the structures accessible to RNA is required to decipher IRES-mediated regulation during translation.…”

Section: Vflip Ires Activity Requires Eif4g and Eif4ementioning

confidence: 99%

Functional analysis of Kaposi's sarcoma–associated herpesvirus vFLIP expression reveals a new mode of IRES-mediated translation

Othman

Sulaiman

Willcocks

et al. 2014

RNA

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“…HCV IRES domain II enhances binding of the coding sequence in the mRNA-binding cleft of 40S subunits by inducing conformational changes, including rotation of the head relative to the body, opening of the entry latch of the mRNA-binding cleft, and closure of the cleft at the exit side (Kolupaeva et al 2000a, Spahn et al 2001. The larger size of domain II of the SPV9 IRES compared to HCV and CSFV-like IRESs (Lukavsky et al 2003;Hellen and de Breyne 2007;Locker et al 2007) may result in it making additional contacts with the 40S subunit resulting in additional conformational changes in the bound ribosome compared to those caused by binding of the HCV IRES (Spahn et al 2001) which may account for the ability of the SPV9 IRES to promote eIF2-independent binding of Met-tRNA i Met to the P site. We suggested that HP-like IRESs are modular elements that have exchanged between flaviviruses (which include HCV and CSFV) and some picornaviruses (such as SPV9) by recombination .…”

Section: The Mechanism Of Initiation On the Spv9 Iresmentioning

confidence: 99%

“…HP IRESs consist of the z100-nt-long domain II, which enhances 48S complex formation and subunit joining, domain III, which contains a pseudoknot and several branched hairpins that together bind eIF3 and the 40S subunit, and a sequence between the pseudoknot and the initiation codon that in CSFV is unstructured and in HCV forms the hairpin domain IV (Reynolds et al 1995;Honda et al 1996;Pestova et al 1998b;Sizova et al 1998;Kolupaeva et al 2000a,b;Kieft et al 2001;Pisarev et al 2005;Locker et al 2007). The coding sequences adjacent to HCV and CSFV IRESs are unrelated but both unstructured, which contributes to initiation efficiency (Reynolds et al 1995;Fletcher et al 2002).…”

Section: Introductionmentioning

confidence: 99%

Factor requirements for translation initiation on the Simian picornavirus internal ribosomal entry site

Breyne¹,

Yu²,

Pestova³

et al. 2007

RNA

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The Simian picornavirus type 9 (SPV9) 59-untranslated region (59 UTR) has been predicted to contain an internal ribosomal entry site (IRES) with structural elements that resemble domains of hepacivirus/pestivirus (HP) IRESs. In vitro reconstitution of initiation confirmed that this 59 UTR contains an IRES and revealed that it has both functional similarities and differences compared to HP IRESs. Like HP IRESs, the SPV9 IRES bound directly to 40S subunits and eukaryotic initiation factor (eIF) 3, depended on the conserved domain IIId for ribosomal binding and consequently for function, and additionally required eIF2/ initiator tRNA to yield 48S complexes that formed elongation-competent 80S ribosomes in the presence of eIF5, eIF5B, and 60S subunits. Toeprinting analysis revealed that eIF1A stabilized 48S complexes, whereas eIF1 induced conformational changes in the 40S subunit, likely corresponding to partial opening of the entry latch of the mRNA-binding channel, that were exacerbated by eIF3 and suppressed by eIF1A. The SPV9 IRES differed from HP IRESs in that its function was enhanced by eIF4A/eIF4F when the IRES was adjacent to the wild-type coding sequence, but was less affected by these factors or by a dominant negative eIF4A mutant when potentially less structured coding sequences were present. Exceptionally, this IRES promoted binding of initiator tRNA to the initiation codon in the P site of 40S subunits independently of eIF2. Although these 40S/IRES/tRNA complexes could not form active 80S ribosomes, this constitutes a second difference between the SPV9 and HP IRESs. eIF1 destabilized the eIF2-independent ribosomal binding of initiator tRNA.

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“…These IRESes also use the eIF2 Á tRNA Á GTP ternary complex and upon GTP hydrolysis, codon-anticodon dependent release of GDP/phosphate, and the action of eIF5B, the 60S ribosomal subunit binds to form the 80S ribosome-IRES complex, followed by the first peptide bond formation Unbehaun et al 2004;Locker et al 2007). Various IRES RNA secondary structure elements, including a pseudoknot near the 39 end of the IRES (Wang et al 1995), have been shown to influence the 40S subunit, eIF3, and ternary complex recruitment, as well as eIF2 release and 80S ribosome formation (Wang et al 1995;Kieft et al 2001;Ji et al 2004;Otto and Puglisi 2004;Locker et al 2007).…”

Section: Viral Ires Groupsmentioning

confidence: 99%

RNA structure-based ribosome recruitment: Lessons from the Dicistroviridae intergenic region IRESes

Pfingsten

Kieft²

2008

RNA

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In eukaryotes, the canonical process of initiating protein synthesis on an mRNA depends on many large protein factors and the modified nucleotide cap on the 59 end of the mRNA. However, certain RNA sequences can bypass the need for these proteins and cap, using an RNA structure-based mechanism called internal initiation of translation. These RNAs are called internal ribosome entry sites (IRESes), and the cap-independent initiation pathway they support is critical for successful infection by many viruses of medical and economic importance. In this review, we briefly describe and compare mechanistic and structural groups of viral IRES RNAs, focusing on those IRESes that are capable of direct ribosome recruitment using specific RNA structures. We then discuss in greater detail some recent advances in our understanding of the intergenic region IRESes of the Dicistroviridae, which use the most streamlined ribosome-recruitment mechanism yet discovered. By combining these findings with knowledge of canonical translation and the behavior of other IRESes, mechanistic models of this RNA structure-based process are emerging.

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HCV and CSFV IRES domain II mediate eIF2 release during 80S ribosome assembly

Cited by 132 publications

References 52 publications

Functional analysis of Kaposi's sarcoma–associated herpesvirus vFLIP expression reveals a new mode of IRES-mediated translation

Functional analysis of Kaposi's sarcoma–associated herpesvirus vFLIP expression reveals a new mode of IRES-mediated translation

Factor requirements for translation initiation on the Simian picornavirus internal ribosomal entry site

RNA structure-based ribosome recruitment: Lessons from the Dicistroviridae intergenic region IRESes

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