Show simple item record

dc.contributor.advisorKieft, Jeffrey S.
dc.contributor.advisorZhao, Rui
dc.contributor.authorHartwick, Erik William
dc.contributor.committeememberChurchill, Mair
dc.contributor.committeememberJohnson, Aaron M.
dc.contributor.committeememberHesselberth, Jay
dc.contributor.committeememberEvans, Thomas C.
dc.date.accessioned2018-08-31T15:37:42Z
dc.date.available2020-08-30T15:37:46Z
dc.date.submitted2018
dc.descriptionIncludes bibliographical references.
dc.descriptionSummer
dc.description.abstractIn addition to the well-known 5’ cap and 3’ poly(A) tail, it is becoming increasingly clear that signals based on structured RNA elements within a messenger 3’ untranslated region (3’ UTR) can enhance translation in eukaryotes; these are poorly understood. To address this, I used a powerful model system from the turnip yellow mosaic virus (TYMV). The TYMV viral RNAs are 5’-capped but instead of a poly(A) tail, they terminate in a highly structured 3’ UTR that contains two structural domains: an upstream pseudoknot domain (UPD) and a tRNA-like structure (TLS). These domains work together to enhance translation, making this system an intriguing model for learning fundamental rules of how a complex structured 3’ UTR affects protein synthesis. Using a combination of biochemistry, translation assays, structural biology, and biophysics, I discovered that the two RNA domains form a conformationally dynamic higher-order architecture in which interactions between the domains affect translation enhancement. Furthermore, the RNA structure appears to act as a ribosome-induced structural switch through programmed global conformational changes. Using multi-dimensional chemical mapping, mutagenesis, and functional assays, I characterized this interaction and identified novel point mutants that affect the interaction and either increase or decrease function. The structure of the entire 3’ UTR was solved by x-ray crystallography, giving novel insight into the architecture and structural details of the RNA and illuminating how a structured, dynamic, multi-domain RNA at the 3’ end can regulate translation. In addition, this investigation lends insight into how translation enhancement is achieved by this 3’ UTR, providing the first direct evidence that this structured 3’UTR operates at the level of translation initiation. Additionally, my preliminary findings suggest the TYMV 3’UTR may have a direct role in the late stages of the initiation pathway through novel interactions with ribosome complexes at the start codon. Taken together the work presented here shows how programmed conformational changes to a 3’ UTR can regulate translation initiation from the 5’ end of eukaryotic messages, providing new insight into 5’ and 3’ end communication.
dc.identifierHartwick_ucdenveramc_1639D_10551.pdf
dc.identifier.urihttps://hdl.handle.net/10968/2590
dc.languageEnglish
dc.publisherUniversity of Colorado Anschutz Medical Campus. Strauss Health Sciences Library
dc.rightsCopyright of the original work is retained by the author.
dc.rights.accessEmbargo Expires: 08/30/2020
dc.subjectRNA chemical mapping
dc.subjectStructured 3'UTR
dc.subjecttRNA-like structure
dc.subjectRNA structure
dc.subjectConformational dynamics
dc.subjectTranslation
dc.subject.meshRNA
dc.subject.meshRNA, Transfer
dc.titleUnderstanding structured-viral 3’UTR control of protein sysnthesis
dc.title.alternativeProgrammed RNA conformational dynamics and the regulation of translation rates : understanding structured-viral 3’UTR control of protein synthesis
dc.typeThesis
dcterms.embargo.expires2020-08-30
thesis.degree.disciplineStructural Biology & Biochemistry
thesis.degree.grantorUniversity of Colorado at Denver, Anschutz Medical Campus
thesis.degree.levelDoctoral
thesis.degree.nameDoctor of Philosophy (Ph.D.)


Files in this item

Thumbnail

This item appears in the following Collection(s)

Show simple item record