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dc.contributor.authorWelch, Ellen Marie
dc.date2022-08-11T08:08:41.000
dc.date.accessioned2022-08-23T16:04:01Z
dc.date.available2022-08-23T16:04:01Z
dc.date.issued1999-07-16
dc.date.submitted2007-04-04
dc.identifier.urihttp://hdl.handle.net/20.500.14038/31601
dc.descriptionIn the process of seeking author's permission to provide full text.
dc.description.abstractmRNA decay is an important cellular process that regulates gene expression and is tightly linked to the process of translation. Many studies have illustrated the link between mRNA turnover and translation, indicating that mRNA decay is a cytoplasmic event. In order to investigate further the link between translation and turnover, seven mutants in translation initiation factors were analyzed for their effect on mRNA decay, including: i) three mutant alleles of the PRT1 gene (prt1-1, prt1-26 and prt1-63), which encodes a subunit of elF3; ii) sui1-1, which encodes the smallest subunit of elF3; iii) sui2-1, which encodes elF2; iv) GCN2c, which encodes the elF2 kinase, and v) cdc33-42, a mutant in the cap binding protein elF4E. The results demonstrate that the prt1-1 mutation results in stabilization of nonsense containing mRNAs without affecting the half-lives of most other mRNAs, a phenotype similar to a upf1Δ strain. To identify substrates for the nonsense-mediated mRNA decay pathway, mRNA differential display analysis was performed using RNA prepared from prt1-1, PRT1, upf1Δ and UPF1 strains. Although the abundance of the HHF2 mRNA is increased in the mutant strains the half-life is unaffected. However, the mRNA half-life of the transcriptional regulator SPT10 was increased in the mutant strains indicating the SPT10 transcript is a substrate for the nonsense-mediated mRNA decay pathway. Further characterization of the SPT10 transcript showed that it is a substrate for this pathway because the initiator AUG is present in a poor translation initiation context which results in aberrant translation initiation. Finally, several other mRNAs, predicted to be substrates for the pathway based on the leaky scanning model, were subsequently shown to decay through this pathway. These transcripts included the REV7, STE50, and UBP7 mRNAs. The results from these experiments lay the groundwork for addressing the potential regulatory role of the nonsense-mediated mRNA decay pathway.
dc.language.isoen_US
dc.rightsCopyright is held by the author, with all rights reserved.
dc.subjectSaccharomyces cerevisiae
dc.subjectRNA, Messenger
dc.subjectAcademic Dissertations
dc.subjectLife Sciences
dc.subjectMedicine and Health Sciences
dc.titleThe Role of Translation Initiation in Nonsense-Mediated mRNA Decay in the Yeast Saccharomyces Cerevisiae: a Dissertation
dc.typeDoctoral Dissertation
dc.identifier.legacycoverpagehttps://escholarship.umassmed.edu/gsbs_diss/289
dc.legacy.embargo2017-04-24T00:00:00-07:00
dc.identifier.contextkey288600
html.description.abstract<p>mRNA decay is an important cellular process that regulates gene expression and is tightly linked to the process of translation. Many studies have illustrated the link between mRNA turnover and translation, indicating that mRNA decay is a cytoplasmic event. In order to investigate further the link between translation and turnover, seven mutants in translation initiation factors were analyzed for their effect on mRNA decay, including: i) three mutant alleles of the <em>PRT1</em> gene (<em>prt1-1, prt1-26</em> and <em>prt1-63</em>), which encodes a subunit of elF3; ii) <em>sui1-1</em>, which encodes the smallest subunit of elF3; iii) <em>sui2-1</em>, which encodes elF2; iv) <em>GCN2<sup>c</sup></em>, which encodes the elF2 kinase, and v) <em>cdc33-42</em>, a mutant in the cap binding protein elF4E. The results demonstrate that the <em>prt1-1</em> mutation results in stabilization of nonsense containing mRNAs without affecting the half-lives of most other mRNAs, a phenotype similar to a <em>upf1Δ</em> strain.</p> <p>To identify substrates for the nonsense-mediated mRNA decay pathway, mRNA differential display analysis was performed using RNA prepared from <em>prt1-1, PRT1, upf1Δ</em> and <em>UPF1</em> strains. Although the abundance of the <em>HHF2</em> mRNA is increased in the mutant strains the half-life is unaffected. However, the mRNA half-life of the transcriptional regulator <em>SPT10</em> was increased in the mutant strains indicating the <em>SPT10</em> transcript is a substrate for the nonsense-mediated mRNA decay pathway. Further characterization of the <em>SPT10</em> transcript showed that it is a substrate for this pathway because the initiator AUG is present in a poor translation initiation context which results in aberrant translation initiation. Finally, several other mRNAs, predicted to be substrates for the pathway based on the leaky scanning model, were subsequently shown to decay through this pathway. These transcripts included the <em>REV7, STE50,</em> and <em>UBP7</em> mRNAs. The results from these experiments lay the groundwork for addressing the potential regulatory role of the nonsense-mediated mRNA decay pathway.</p>
dc.identifier.submissionpathgsbs_diss/289
dc.contributor.departmentGraduate School of Biomedical Sciences
dc.contributor.departmentDepartment of Molecular Genetics and Microbiology


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