Graduate Studies

 

First Advisor

Audrey L. Atkin

Second Advisor

Kenneth W. Nickerson

Degree Name

Doctor of Philosophy (Ph.D.)

Department

Biological Sciences

Date of this Version

8-2024

Document Type

Dissertation

Citation

A dissertation presented to the faculty of the Graduate College of the University of Nebraska in partial fulfillment of requirements for the degree of Doctor of Philosophy

Major: Biological Sciences (Genetics, Cellular, and Molecular Biology)

Under the supervision of Professors Audrey L. Atkin and Kenneth W. Nickerson

Lincoln, Nebraska, August 2024

Comments

Copyright 2024, Daniel John Gutzmann. Used by permission

Abstract

Candida albicans is a polymorphic fungus and opportunistic commensal found primarily in the gastrointestinal and skin of healthy individuals. Several barriers prevent C. albicans from causing disease including a healthy immune system and microbiome. When these barriers become comprised, C. albicans can transition to a pathogen and disseminate through the intestinal mucosa leading to life-threatening bloodstream and invasive infections with mortality rates of up to 64%. Morphogenic plasticity is key to this transition and impacts virulence, adaptation to different host environments, and evasion of host immune responses. One regulator of morphogenesis is farnesol. Farnesol is a secondary metabolite of ergosterol biosynthesis produced by C. albicans throughout growth. Farnesol has a significant role in C. albicans physiology by mediating interactions with the competing microbiota, host immune system, and as an autoregulatory molecule influencing morphogenesis. The mechanisms that influence farnesol synthesis, secretion, regulation, and turnover are poorly understood. To address this gap, we first developed a high-throughput quantitative assay for farnesol accumulation and localization and found that farnesol production is temporal and peaks during early stationary phase. Using this assay, we screened a transcription regulator knockout collection to identify the regulatory network that controls farnesol synthesis and localization by identifying mutants with differences in either their farnesol accumulation, localization or decay. These mutants were involved in processes linked to farnesol’s physiological roles including cell cycle progression, white-opaque switching, yeast-mycelial dimorphism, and response to stress. To elucidate the genes involved in farnesol synthesis, we performed an RNA-seq experiment on two farnesol underproducing mutants. In comparing the differentially down-regulated genes in those mutants with those genes up-regulated in farnesol over accumulating mutants, we identified CWH8 as a candidate gene. We generated cwh8 null mutants and found that these mutants were defective in farnesol synthesis. In addition, we have made significant contributions to our understanding secretion and turnover of farnesol by elucidating the role of serum albumin and finding that farnesol salvage is absent in C. albicans. Collectively, this dissertation contributes significantly to our understanding of how farnesol is synthesized, regulated, and metabolized which is vital for interpreting farnesol’s complex physiological roles in C. albicans.

Advisors: Audrey L. Atkin and Kenneth W. Nickerson

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