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Étude structurale et fonctionnelle de l’élément NRS régulateur négatif de l’épissage de l’ARN du virus du Sarcome de Rous

Abstract : Retroviruses require both spliced and unspliced RNAs for productive replication. Accumulation of unspliced RNA in Rous Sarcoma Virus (RSV) depends on the NRS element, (Negative Regulator of Splicing). The NRS element is bipartite. Its 5’ terminal part is considered as an ESE that binds SR proteins and its 3’ part contains a decoy 5’-splice site (ss), which inhibits splicing at the bona fide 5’ ss. Only the 3D structure of a small NRS fragment including the decoy 5’ ss had been experimentally studied. Here, by chemical and enzymatic probing of entire RSV NRS, we determine its 2D structure. By comparative analysis of 2D structures of functional and non-functional avian NRS variants and of all sequenced avian NRSs, we bring strong arguments for a biological significance of the established structure. By in vitro splicing assays, we show a crucial role of two of the established stem-loop structures and by affinity purification of complexes formed by WT and truncated NRSs in HeLa cell nuclear extract, we demonstrate their importance for SR protein and U1 snRNP association. We define a new small NRS element retaining splicing inhibitory properties and finally demonstrate the capability of the SR protein 9G8 to increase NRS activity in vitro and in cellulo
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https://hal.univ-lorraine.fr/tel-01746321
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  • HAL Id : tel-01746321, version 1

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Aileen Bar. Étude structurale et fonctionnelle de l’élément NRS régulateur négatif de l’épissage de l’ARN du virus du Sarcome de Rous. Médecine humaine et pathologie. Université Henri Poincaré - Nancy 1, 2011. Français. ⟨NNT : 2011NAN10153⟩. ⟨tel-01746321⟩

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