title: High-affinity interaction of hnRNP A1 with conserved RNA structural elements is required for translation and replication of enterovirus 71.
abstract: Human Enterovirus 71 (EV71) is an emerging pathogen of infectious disease and a serious threat to public health. Currently, there are no antivirals or vaccines to slow down or prevent EV71 infections, thus underscoring the urgency to better understand mechanisms of host-enterovirus interactions. EV71 uses a type I internal ribosome entry site (IRES) to recruit the 40S ribosomal subunit via a pathway that requires the cytoplasmic localization of hnRNP A1, which acts as an IRES trans-activating factor. The mechanism of how hnRNP A1 trans activates EV71 RNA translation is unknown, however. Here, we report that the UP1 domain of hnRNP A1 interacts specifically with stem loop II (SLII) of the IRES, via a thermodynamically well-defined biphasic transition that involves conserved bulge 5'-AYAGY-3' and hairpin 5'-RY(U/A)CCA-3' loops. Calorimetric titrations of wild-type and mutant SLII constructs reveal these structural elements are essential to form a high-affinity UP1-SLII complex. Mutations that alter the bulge and hairpin primary or secondary structures abrogate the biphasic transition and destabilize the complex. Notably, mutations within the bulge that destabilize the complex correlate with a large reduction in IRES-dependent translational activity and impair EV71 replication. Taken together, this study shows that a conserved SLII structure is necessary to form a functional hnRNP A1-IRES complex, suggesting that small molecules that target this stem loop may have novel antiviral properties.
authors.affiliation: Department of Chemistry; Case Western Reserve University; Cleveland, OH USA.
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Levengood
authors.forename: Michele
authors.initials: M
authors.lastname: Tolbert
authors.forename: Mei-Ling
authors.initials: ML
authors.lastname: Li
authors.forename: Blanton S
authors.initials: BS
authors.lastname: Tolbert
mesh_terms: D001483:Base Sequence; D017124:Conserved Sequence; D004770:Enterovirus / Q000145:classification / Q000502:physiology*; D029821:Enterovirus A, Human / Q000502:physiology; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D010802:Phylogeny; D011485:Protein Binding; D014176:Protein Biosynthesis; D011487:Protein Conformation; D059370:RNA Folding; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D020218:Response Elements; D014779:Virus Replication
chemical_list: D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D012367:RNA, Viral; C000619656:hnRNPA1 protein, human
keywords: Enterovirus 71; Internal Ribosomal Entry Site; Protein-RNA interactions; RNA structure; hnRNP A1