MECHANICS OF HANTAVIRAL NUCLEOCAPSID PROTEIN MEDIATED TRANSLATION INITIATION OF

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Eukaryotic translation begins with recruitment of eIF4F complex at mRNA cap with the engagement of 43S pre-initiation complex at mRNA 5' terminus. Another well characterized mechanism utilized by several viruses includes IRES translation initiation strategy that internally loads ribosomes on mRNA, independent of 5' cap. Hantaviruses, members of the Bunyaviridae family are emerging viruses that initiate mRNA translation by a different novel mechanism, using viral capsid protein (N) to engage the ribosome at mRNA cap, independent of eukaryotic IF4F complex. We will further characterize N mediated translation initiation mechanism and illustrate possible benefits of this novel strategy that favor virus replication in infected cells. We will identify the components of 43S pre-initiation complex that interact with N. N specifically binds the viral mRNA 5' UTR with high affinity and referentially facilitates the translation of viral mRNAs in vitro. We will identify and characterize the binding site for N on viral mRNA 5' UTR and will determine whether N preferentially facilitates the translation of viral RNAs in host cells. N is also an RNA chaperone that unwinds RNA duplexes. However, this RNA chaperone activity is not involved in N mediated mRNA translation. Secondary structures in mRNA 5' UTR are removed by eIF4A (a component of eIF4F complex) during ribosome scanning and identification of AUG codon. Since N functionally supplants eIF4F complex, we hypothesize that N translocates the loaded ribosomes from 5' cap to the AUG codon, avoiding the regular scanning of 5' leader. Multifaceted experimental approaches have been designed to test this hypothesis. We will use multiple experimental approaches to check whether N mediated translation strategy is a viral counter measure against host cell antiviral response.