title: Hantavirus nucleocapsid protein has distinct m7G cap- and RNA-binding sites.
abstract: Hantaviruses, members of the Bunyaviridae family, are emerging category A pathogens that carry three negative stranded RNA molecules as their genome. Hantavirus nucleocapsid protein (N) is encoded by the smallest S segment genomic RNA (viral RNA). N specifically binds mRNA caps and requires four nucleotides adjacent to the cap for high affinity binding. We show that the N peptide has distinct cap- and RNA-binding sites that independently interact with mRNA cap and viral genomic RNA, respectively. In addition, N can simultaneously bind with both mRNA cap and vRNA. N undergoes distinct conformational changes after binding with either mRNA cap or vRNA or both mRNA cap and vRNA simultaneously. Hantavirus RNA-dependent RNA polymerase (RdRp) uses a capped RNA primer for transcription initiation. The capped RNA primer is generated from host cell mRNA by the cap-snatching mechanism and is supposed to anneal with the 3' terminus of vRNA template during transcription initiation by single G-C base pairing. We show that the capped RNA primer binds at the cap-binding site and induces a conformational change in N. The conformationally altered N with a capped primer loaded at the cap-binding site specifically binds the conserved 3' nine nucleotides of vRNA and assists the bound primer to anneal at the 3' terminus. We suggest that the cap-binding site of N, in conjunction with RdRp, plays a key role during the transcription and replication initiation of vRNA genome.
authors.affiliation: Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City, Kansas 66103, USA. mmir@kumc.edu
authors.forename: Mohammad A
authors.initials: MA
authors.lastname: Mir
authors.forename: Sheema
authors.initials: S
authors.lastname: Sheema
authors.forename: Abdul
authors.initials: A
authors.lastname: Haseeb
authors.forename: Absarul
authors.initials: A
authors.lastname: Haque
mesh_terms: D020029:Base Pairing; D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers / Q000737:chemistry; D006476:Orthohantavirus / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D019590:Nucleocapsid Proteins / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D012313:RNA / Q000737:chemistry*; D012315:RNA Caps / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D013050:Spectrometry, Fluorescence / Q000379:methods; D013696:Temperature; D014158:Transcription, Genetic
chemical_list: D017931:DNA Primers; D019590:Nucleocapsid Proteins; D012315:RNA Caps; D012333:RNA, Messenger; D012313:RNA