Dissertation
Functional and potentially adaptive role of polyglutamine in S. cerevisiae Med15
University of Iowa
Doctor of Philosophy (PhD), University of Iowa
Summer 2021
DOI: 10.17077/etd.005935
Abstract
Cells can respond to stress and changing environments by altering gene expression. The RNA Polymerase II Mediator is a conserved eukaryotic protein complex which mediates expression of target genes through recruitment of RNA Polymerase II to promoters of target genes. Med15, a subunit of the Mediator complex, is located at a critical position in the complex where it is responsible for interacting with DNA-bound signal-specific transcription factors. The wide array of phenotypic defects in Saccharomyces cerevisiae yeast cells in which MED15 has been deleted reflect the role of Med15 in regulating stress response genes as well as genes involved in the metabolism of diverse biomolecules. Med15 is intrinsically disordered and glutamine-rich with three polyglutamine tracts that vary in length among strains. The variation in length (number of glutamines) is due to the instability of the underlying repetitive DNA sequences. In some proteins, expansion of the polyglutamine tracts beyond a threshold length causes neurodegenerative disease. Despite the potential risk of disease-causing expansions, polyglutamine tracts are surprisingly prevalent in eukaryotic proteomes and are specifically enriched in transcriptional regulators, suggesting that polyglutamine tracts are involved in modulating gene expression.
My work focuses on understanding how the intrinsic disorder and glutamine-rich nature of Med15 influences the activity of the protein in yeast. My approach included computational analyses of MED15 sequence variation across yeast strains and non-fungal eukaryotes, an analysis of the functional consequences of natural and synthetic variation in MED15 polyglutamine tracts, and analysis of commercial alcoholic fermentation associated MED15 alleles.
I found that the three polyglutamine tracts within Med15 are highly variable in length and that while a significant amount of tract length variability is tolerated, the transcriptional regulation activity of Med15 varies with polyglutamine tract length. My studies show that many naturally occurring combinations of polyglutamine tract lengths allow Med15 to function normally, but that hybrid genes with engineered (non-natural) combinations of glutamine tracts have impaired function. Results of my studies support a role for polyglutamine tract length variation in facilitating adaptation to the grape juice fermentation environment. I found that natural MED15 alleles acquired from commercial alcoholic fermentation yeast strains with polyglutamine tract length polymorphisms exhibit improved fermentation phenotypes and enhanced expression of fermentation genes.
Finally, my work begins to describe the Med15 features that are conserved across eukaryotes, including an enrichment of disorder-promoting residues instead of conserved primary sequence. I found that non-fungal Med15 orthologs fail to complement defects in the S. cerevisiae med15 mutant and that even fungal orthologs only partially complement med15 phenotypes. This suggests that the conserved disorder does not correspond directly to conserved function. Instead, I propose that the intrinsic disorder of Med15 has permitted Med15 to diverge alongside existing and novel interacting transcription factors. My findings are consistent with a role for the glutamine-rich nature of Med15 in two important ways: allowing adaptation of closely related strains to new environments and allowing for malleability in diverging organisms.
Details
- Title: Subtitle
- Functional and potentially adaptive role of polyglutamine in S. cerevisiae Med15
- Creators
- David George Cooper
- Contributors
- Jan Fassler (Advisor)Josep Comeron (Committee Member)Albert Erives (Committee Member)Chad Grueter (Committee Member)Anna Malkova (Committee Member)
- Resource Type
- Dissertation
- Degree Awarded
- Doctor of Philosophy (PhD), University of Iowa
- Degree in
- Integrated Biology
- Date degree season
- Summer 2021
- DOI
- 10.17077/etd.005935
- Publisher
- University of Iowa
- Number of pages
- xiv, 262 pages
- Copyright
- Copyright 2021 David George Cooper
- Language
- English
- Description illustrations
- color illustrations
- Description bibliographic
- Includes bibliographical references (pages 241-262).
- Public Abstract (ETD)
Baker’s yeast is a single-celled organism that survives in diverse environments from the human body to grape skins. Yeast respond to changing environments by turning on the most useful set of genes for a given condition. Med15 is a protein that is important in promoting the expression of other genes that help yeast respond to stress or to metabolize new types of nutrients.
Yeast is an excellent model for understanding gene expression in more complex organisms. Med15 is an unusual protein with an overwhelming abundance of a single amino acid, glutamine. Some stretches of the protein consist of many glutamines in a row (polyglutamine tracts). Different strains of yeast can have polyglutamine tracts of different lengths, or number of consecutive glutamine residues.
I proposed that Med15 variants might be optimized to help the yeast best survive in specific environments. The first goal of my work was to determine if the different versions of Med15 with polyglutamine tracts of different lengths regulated transcription differently. I found that Med15 from wine yeast promote the expression of genes that help yeast more efficiently ferment grape juice. The second goal of my work was to describe how the polyglutamine tracts affect Med15 activity. I found that Med15 does not function as well without polyglutamine tracts, and some activities of the protein can be changed by making the polyglutamine tracts a different length. My work increases our understanding of how specific changes in Med15, or similar proteins, can contribute to the function of the protein
- Academic Unit
- Biology; Craniofacial Anomalies Research Center
- Record Identifier
- 9984124759402771
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