title: Characterization of the termini of the West Nile virus genome and their interactions with the small isoform of the 2' 5'-oligoadenylate synthetase family.
abstract: 2' 5'-Oligoadenylate synthetases (OAS) are interferon-stimulated proteins that act in the innate immune response to viral infection. Upon binding viral double-stranded RNA, OAS enzymes produce 2'-5'-linked oligoadenylates that stimulate RNase L and ultimately slow viral propagation. Truncations/mutations in the smallest human OAS isoform, OAS1, results in susceptibility to West Nile virus (WNV). We have previously demonstrated in vitro the interaction between OAS1 and the 5'-terminal region of the WNV RNA genome. Here we report that the 3'-terminal region is also able to mediate specific interaction with and activation of OAS1. Binding and kinetic experiments identified a specific stem loop within the 3'-terminal region that is sufficient for activation of the enzyme. The solution conformation of the 3'-terminal region was determined by small angle X-ray scattering, and computational models suggest a conformationally restrained structure comprised of a helix and short stem loop. Structural investigation of the 3'-terminal region in complex with OAS1 is also presented. Finally, we show that genome cyclization by base pairing between the 5'- and 3'-terminal regions, a required step for replication, is not sufficient to protect WNV from OAS1 recognition in vitro. These data provide a physical framework for understanding recognition of the highly structured terminal regions of a flaviviral genome by an innate immune enzyme.
authors: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors: Soumya
authors: S
authors: Deo
authors: School of Biosciences, University of Birmingham, Birmingham B152TT, UK.
authors: Trushar R
authors: TR
authors: Patel
authors: Laboratory of Bioinformatics and Protein Engineering, International Institute of Molecular and Cell Biology, ul. Ks. Trojdena 4, 02-109 Warsaw, Poland.
authors: Grzegorz
authors: G
authors: Chojnowski
authors: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors: Amit
authors: A
authors: Koul
authors: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors: Edis
authors: E
authors: Dzananovic
authors: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada; Manitoba Institute for Materials, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors: Kevin
authors: K
authors: McEleney
authors: Laboratory of Bioinformatics and Protein Engineering, International Institute of Molecular and Cell Biology, ul. Ks. Trojdena 4, 02-109 Warsaw, Poland; Laboratory of Bioinformatics, Institute of Molecular Biology and Biotechnology, Faculty of Biology, Adam Mickiewicz University, ul. Umultowska 89, 61-614 Poznan, Poland.
authors: Janusz M
authors: JM
authors: Bujnicki
authors: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada; Department of Biochemistry and Medical Genetics, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada. Electronic address: sean.mckenna@umanitoba.ca.
authors: Sean A
authors: SA
authors: McKenna
mesh_terms: D015088:2',5'-Oligoadenylate Synthetase / Q000235:genetics* / Q000378:metabolism; D020029:Base Pairing; D001483:Base Sequence; D002151:Calorimetry; D000067493:Dynamic Light Scattering; D024202:Electrophoretic Mobility Shift Assay; D016679:Genome, Viral / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics* / Q000378:metabolism; D053838:Scattering, Small Angle; D013728:Terminator Regions, Genetic / Q000235:genetics*; D014902:West Nile virus / Q000235:genetics*
chemical_list: D011994:Recombinant Proteins; C475085:OAS1 protein, human; D015088:2',5'-Oligoadenylate Synthetase
keywords: Computational structure determination; Oligoadenylate synthetase; RNA–protein interaction; Small angle X-ray scattering; Terminal region; West Nile virus