Understanding structured-viral 3'UTR control of protein sysnthesis
Year: 2018
Type: dissertation
Abstract: In 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.
DOI: 10.25677/cmgm-r756
Author: Hartwick, Erik William
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W3134885468