title: SHAPE analysis of the RNA secondary structure of the Mouse Hepatitis Virus 5' untranslated region and N-terminal nsp1 coding sequences.
abstract: SHAPE technology was used to analyze RNA secondary structure of the 5' most 474 nts of the MHV-A59 genome encompassing the minimal 5' cis-acting region required for defective interfering RNA replication. The structures generated were in agreement with previous characterizations of SL1 through SL4 and two recently predicted secondary structure elements, S5 and SL5A. SHAPE provided biochemical support for four additional stem-loops not previously functionally investigated in MHV. Secondary structure predictions for 5' regions of MHV-A59, BCoV and SARS-CoV were similar despite high sequence divergence. The pattern of SHAPE reactivity of in virio genomic RNA, ex virio genomic RNA, and in vitro synthesized RNA was similar, suggesting that binding of N protein or other proteins to virion RNA fails to protect the RNA from reaction with lipid permeable SHAPE reagent. Reverse genetic experiments suggested that SL5C and SL6 within the nsp1 coding sequence are not required for viral replication.
authors.affiliation: Department of Microbial Pathogenesis and Immunology, Texas A&M University, College of Medicine, 407 Reynolds Medical Building, 1114 TAMU, College Station, TX 77843-1114, USA.
authors.forename: Dong
authors.initials: D
authors.lastname: Yang
authors.affiliation: Department of Microbial Pathogenesis and Immunology, Texas A&M University, College of Medicine, 407 Reynolds Medical Building, 1114 TAMU, College Station, TX 77843-1114, USA.
authors.forename: Pinghua
authors.initials: P
authors.lastname: Liu
authors.affiliation: Department of Microbial Pathogenesis and Immunology, Texas A&M University, College of Medicine, 407 Reynolds Medical Building, 1114 TAMU, College Station, TX 77843-1114, USA.
authors.forename: Elyse V
authors.initials: EV
authors.lastname: Wudeck
authors.affiliation: Department of Chemistry, Indiana University, Bloomington, IN 47405-7102, USA.
authors.forename: David P
authors.initials: DP
authors.lastname: Giedroc
authors.affiliation: Department of Microbial Pathogenesis and Immunology, Texas A&M University, College of Medicine, 407 Reynolds Medical Building, 1114 TAMU, College Station, TX 77843-1114, USA. Electronic address: jleibowitz@tamu.edu.
authors.forename: Julian L
authors.initials: JL
authors.lastname: Leibowitz
mesh_terms: D020121:5' Untranslated Regions / Q000235:genetics*; D000818:Animals; D015967:Gene Expression Regulation, Viral / Q000502:physiology*; D051379:Mice; D006517:Murine hepatitis virus / Q000235:genetics / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D012367:RNA, Viral / Q000737:chemistry* / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D020121:5' Untranslated Regions; C505227:NSP1 protein, murine hepatitis virus; D012367:RNA, Viral; D017361:Viral Nonstructural Proteins
keywords: 5′ cis-acting region; Betacoronavirus; Mouse Hepatitis Virus; Murine coronavirus; RNA secondary structure; SHAPE analysis