Strasbourg University, CNRS, RNA Architecture and Reactivity, UPR 9002, Strasbourg The IRES 5΄UTR of the dicistrovirus cricket paralysis virus is a type III IRES containing an essential pseudoknot structure Nucleic Acids Research 2017 45:8993–9004 doi: 10.1093/nar/gkx622 Lauriane Gross, Quentin Vicens, Evelyne Einhorn, Audrey Noireterre, Laure Schaeffer, Lauriane Kuhn, Jean-Luc Imler, Gilbert Eriani, Carine Meignin and Franck Martin
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Currently in the third year of her PhD, Lauriane GROSS obtained a Bachelor's and a Master's degree in Molecular Biology and Genetics at Strasbourg University. For her Master M2 internship she joined the team of Dr. Gilbert ERIANI "Evolution of translation initiation systems in eukaryotes" at IBMC in Strasbourg. She is now doing her PhD in the same team under the direction of Dr. Franck MARTIN, where she is studying the molecular mechanisms used by the Cricket Paralysis Virus (CrPV) to hijack the host's translational machinery and synthesize its own viral proteins. This dicistrovirus possesses, in its genomic RNA, two open reading frames (ORF). The translation initiation of the two ORFs is controlled by structural elements called 'internal ribosome entry sites' (IRES). These RNA structures allow the virus to attract and hijack the host ribosomes and translation initiation factors in order to translate its viral proteins in a rapid and very efficient way. The translation of each CrPV ORF is guided by an IRES. The intergenic region IRES governing the ORF2 has been extensively studied. In this article, the authors have the structure of this IRES and the molecular mechanisms that allow the IRES to direct the ORF1 protein synthesis. They also identify the host cell initiation factors required for translation by this IRES. All these results allowed the authors to conclude that this IRES belongs to the IRES class III.
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CNRS UPR 9002 - RNA Architecture and Reactivity
Gilbert ERIANI Team
IBMC, 15 rue René Descartes
67084 Strasbourg Cedex
Abstract
Cricket paralysis virus (CrPV) is a dicistrovirus. Its positive-sense single-stranded RNA genome contains two internal ribosomal entry sites (IRESs). The 5' untranslated region (5'UTR) IRES5'UTR mediates translation of non-structural proteins encoded by ORF1 whereas the well-known intergenic region (IGR) IRESIGR is required for translation of structural proteins from open reading frame 2 in the late phase of infection. Concerted action of both IRES is essential for host translation shut-off and viral translation. IRESIGR has been extensively studied, in contrast the IRES5'UTR remains largely unexplored. Here, we define the minimal IRES element required for efficient translation initiation in drosophila S2 cell-free extracts. We show that IRES5'UTR promotes direct recruitment of the ribosome on the cognate viral AUG start codon without any scanning step, using a Hepatitis-C virus-related translation initiation mechanism. Mass spectrometry analysis revealed that IRES5'UTR recruits eukaryotic initiation factor 3, confirming that it belongs to type III class of IRES elements. Using Selective 2'-hydroxyl acylation analyzed by primer extension and DMS probing, we established a secondary structure model of 5'UTR and of the minimal IRES5'UTR. The IRES5'UTR contains a pseudoknot structure that is essential for proper folding and ribosome recruitment. Overall, our results pave the way for studies addressing the synergy and interplay between the two IRES from CrPV.
