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Dissertation
ADVANCING PROTEOMICS APPROACHES TO STUDY STRESS RESPONSE AND METABOLISM IN OLEAGINOUS BASIDIOMYCETE YEASTS
Washington State University
Doctor of Philosophy (PhD), Washington State University
12/2024
DOI:
https://doi.org/10.7273/000007226
Abstract
Basidiomycetes are vital to environmental decomposition of plant litter. Oleaginous basidiomycete yeasts are particularly attractive for biorefineries given their capabilities to utilize diverse biomass-derived carbon sources, to store carbon as lipids, and to synthesize biofuels, chemicals, and proteins. However, ascomycete yeasts are frequently chassis for metabolic engineering and models for studying the oleaginous phenotype. For basidiomycete yeasts, the full extent of their metabolic capabilities and intricacies of metabolic regulation are understudied.
This dissertation describes original research concerning nutrient-induced stress response and metabolic regulation in two oleaginous basidiomycete yeasts: Rhodotorula toruloides and Cutaneotrichosporon oleaginosum. Both these yeasts accumulate lipids as a result of nitrogen limitation, which is the primary bioprocessing strategy used for oil production. Their ability to accumulate lipids and the quality of the resulting oil is affected by the type and availability of carbonaceous nutrients. As building blocks of life, limiting access to these nutrients requires a cellular responses to adapt and survive. The resulting phenotypic changes are largely due to alterations in protein expression and regulation. In this work, advanced proteomics approaches are harnessed to provide insights into how cellular responses are orchestrated at the molecular level.
This work demonstrates the first example of redox proteomics applied to an oleaginous yeast to study a protein modification called cysteine oxidation, which is an important type of cell signal and regulatory mechanism in response to nitrogen limitation. Antioxidants are indispensable for combatting protein oxidation during nutrient-limited stress but also protect fungi against reactive compounds they produce to deconstruct lignin, which is the most carbon-dense component of plant cell walls. Unlike previous research endeavors that focused on filamentous fungi and bacteria, the lignin-degrading capacity of a basidomycete yeast was studied herein. Whole-cell 2D 1H-13C NMR and fluorescence microscopy were used to analyze lignin structural changes and lignin transport, whereas novel secretomics and cellular proteomics methods were employed to highlight enzymes putatively involved lignin degradation. Due to the recalcitrance of lignin, the biochemical phenotype of the yeast cultivated with alkali-purified lignin was nonetheless reminiscent of carbon limitation. Taken together, this dissertation furthers our fundamental understanding of biotechnologically-relevant basidiomycete yeasts and provides original targets for genetic engineering.
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Details
- Title
- ADVANCING PROTEOMICS APPROACHES TO STUDY STRESS RESPONSE AND METABOLISM IN OLEAGINOUS BASIDIOMYCETE YEASTS
- Creators
- Austin Gluth
- Contributors
- Bin Yang (Co-Chair)Wei-Jun Qian (Co-Chair)Scott Baker (Committee Member)Shulin Chen (Committee Member)John Miller (Committee Member)
- Awarding Institution
- Washington State University
- Academic Unit
- Department of Biological Systems Engineering
- Theses and Dissertations
- Doctor of Philosophy (PhD), Washington State University
- Publisher
- Washington State University
- Number of pages
- 310
- Identifiers
- 99901195537901842
- Language
- English
- Resource Type
- Dissertation