Author ORCID Identifier

0000-0001-5299-0008

Date of Graduation

5-2021

Document Type

Dissertation (PhD)

Program Affiliation

Microbiology and Molecular Genetics

Degree Name

Doctor of Philosophy (PhD)

Advisor/Committee Chair

Ambro van Hoof

Committee Member

Catherine Denicourt

Committee Member

Theresa M. Koehler

Committee Member

Michael C. Lorenz

Committee Member

Kevin A. Morano

Abstract

Posttranscriptional gene regulation is essential to maintain gene expression fidelity. This is partially achieved by mRNA decay. When no longer required, mRNA is degraded by two alternative pathways. The decapping enzyme Dcp2 removes the 5` m7G cap of mRNAs, allowing Xrn1 to degrade the mRNA from the 5` end. Alternatively, mRNA is degraded from the 3` end by the RNA exosome.

While decapping by Dcp2 is a critical step in mRNA decay, its physiological function has been unclear. Null mutations of Saccharomyces cerevisiae DCP2 have been reported to be lethal in some studies but slow-growing in others. In this study, I show that Dcp2 is required for continuous growth under standard laboratory conditions. I found multiple suppressors of the growth defect of a DCP2 null mutant via experimental evolution and genome sequencing. These suppressors act by at least three independent mechanisms. They do not rescue defects in mRNA decay. Instead, they appear to partially alleviate global disturbance of the transcriptome in the dcp2 mutant, which confers growth improvement. One of the suppressors appears to suppress by affecting translation of CUY codons.

mRNAs that are generated by mistakes need to be degraded. Nonstop mRNAs are transcripts that lack an in-frame stop codon. Thus, the ribosome translating the nonstop mRNA reads through downstream of the coding region and produces aberrant nonstop proteins that are potentially toxic to cells. Cells exploit surveillance mechanisms to suppress the expression of nonstop mRNAs. The RNA exosome and its cofactors, the Ski complex and Ski7, are required for degradation of nonstop mRNAs. While Ski7 is known to have nonstop decay specific function, its exact role in nonstop decay remains unknown. Moreover, whether Ski7-mediated nonstop mRNA decay is mechanistically linked to the surveillance pathway for nonstop protein has been unclear. In this study, I determine the genetic interactions between components of the two surveillance pathways and show that the Ski7-mediated nonstop mRNA decay mechanism functions independently of nonstop protein degradation mechanism.

Keywords

DCP2, mRNA decay, decapping, KAP123, tL(GAG)G

Available for download on Thursday, May 05, 2022

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