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dc.contributor.authorSTORM, KATHLEEN ANNE.
dc.creatorSTORM, KATHLEEN ANNE.en_US
dc.date.accessioned2011-10-31T18:52:29Z
dc.date.available2011-10-31T18:52:29Z
dc.date.issued1984en_US
dc.identifier.urihttp://hdl.handle.net/10150/187819
dc.description.abstractIt is well documented that shortly after infection with poliovirus, host protein synthesis is inhibited. This defect is at the level of initiation and is thought to be caused by a virally-induced alteration of the initiation factor CBPII. This factor is needed for binding and recognition of capped host mRNA to the 40S ribosomal subunit. Consequently, it has been postulated that poliovirus RNA itself is able to initiate protein synthesis via a "cap-independent" mechanism. In order to define the concept of a "cap-independent" mechanism more fully, the importance of the 5' terminus and 5' noncoding region were investigated. Consequently, restriction fragments from the poliovirus clones, pVR104 and pVR105, were selected which hybridized in four regions of the poliovirus genome: (a) a site within the coding sequence; (b) the area covering the AUG at the beginning of the NCVPOO reading frame; (c) a region in the middle of the 5' untranslated region of the genome; (d) the 5' terminal region of the genome. Hybrid molecules were used to program a micrococcal nuclease treated, PV1-infected cell-free system. Message function was measured by the synthesis of protein products, analyzed by SDS-PAGE, formation of polyribosomes and initiation complexes. As expected, the hybrid formed within the coding region allowed the synthesis of the predicted truncated protein. The hybrid covering the AUG codon at position N743 prevented translation. Interestingly, the hybrid in the untranslated 5' region and the hybrid formed at the 5' terminal sequence, inhibited translation and prevented formation of an initiation complex and polyribosomes. This suggested that poliovirus mRNA, despite its lack of a 5' cap, interacts with ribosomes in an manner similar to capped eucaryotic mRNAs and in agreement with a proposed scanning mechanism. Virus specific RNA binding proteins were also characterized in a preliminary investigation to determine which viral proteins exhibit RNA binding capability and through this interaction direct the functional role that the RNA plays within the cell. It was determined that four virus specific proteins do exhibit RNA binding properties. Due to the known functions of these viral proteins, it is unlikely that any of these proteins are responsible for the translational advantage of PV1 RNA in infected cells.
dc.language.isoenen_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.subjectPoliovirus.en_US
dc.titleDETERMINATION OF THE MECHANISM OF INITIATION OF POLIOVIRUS PROTEIN SYNTHESIS.en_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
dc.identifier.oclc693363586en_US
thesis.degree.grantorUniversity of Arizonaen_US
thesis.degree.leveldoctoralen_US
dc.identifier.proquest8501919en_US
thesis.degree.disciplineMolecular and Cellular Biologyen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.namePh.D.en_US
dc.description.noteThis item was digitized from a paper original and/or a microfilm copy. If you need higher-resolution images for any content in this item, please contact us at repository@u.library.arizona.edu.
dc.description.admin-noteOriginal file replaced with corrected file July 2023.
refterms.dateFOA2018-06-20T07:14:24Z
html.description.abstractIt is well documented that shortly after infection with poliovirus, host protein synthesis is inhibited. This defect is at the level of initiation and is thought to be caused by a virally-induced alteration of the initiation factor CBPII. This factor is needed for binding and recognition of capped host mRNA to the 40S ribosomal subunit. Consequently, it has been postulated that poliovirus RNA itself is able to initiate protein synthesis via a "cap-independent" mechanism. In order to define the concept of a "cap-independent" mechanism more fully, the importance of the 5' terminus and 5' noncoding region were investigated. Consequently, restriction fragments from the poliovirus clones, pVR104 and pVR105, were selected which hybridized in four regions of the poliovirus genome: (a) a site within the coding sequence; (b) the area covering the AUG at the beginning of the NCVPOO reading frame; (c) a region in the middle of the 5' untranslated region of the genome; (d) the 5' terminal region of the genome. Hybrid molecules were used to program a micrococcal nuclease treated, PV1-infected cell-free system. Message function was measured by the synthesis of protein products, analyzed by SDS-PAGE, formation of polyribosomes and initiation complexes. As expected, the hybrid formed within the coding region allowed the synthesis of the predicted truncated protein. The hybrid covering the AUG codon at position N743 prevented translation. Interestingly, the hybrid in the untranslated 5' region and the hybrid formed at the 5' terminal sequence, inhibited translation and prevented formation of an initiation complex and polyribosomes. This suggested that poliovirus mRNA, despite its lack of a 5' cap, interacts with ribosomes in an manner similar to capped eucaryotic mRNAs and in agreement with a proposed scanning mechanism. Virus specific RNA binding proteins were also characterized in a preliminary investigation to determine which viral proteins exhibit RNA binding capability and through this interaction direct the functional role that the RNA plays within the cell. It was determined that four virus specific proteins do exhibit RNA binding properties. Due to the known functions of these viral proteins, it is unlikely that any of these proteins are responsible for the translational advantage of PV1 RNA in infected cells.


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