Doctoral Dissertations

Orcid ID

http://orcid.org/0000-0002-8100-7399

Date of Award

8-2019

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Microbiology

Major Professor

Todd Reynolds

Committee Members

Jeffrey Becker, Elizabeth Fozo, Maitreyi Das, Timonthy Sparer

Abstract

Candida albicans is an important cause of systemic infections in the immune compromised population. However, drug resistance and toxicity have put limits on antifungals efficacy. The C. albicans cell wall is considered a good therapeutic target due to its role in fungal pathogenicity. Thus, a potential method for improving antifungal drugs could be to enhance the detection of fungal cell wall antigens by host immune cells. Detection of C. albicans largely occurs through the receptor Dectin-1 that can recognize β [beta] (1,3)-glucan, an important component of fungal cell walls. However, a layer of glycosylated proteins masks the β (1,3)-glucan, hiding it from immune detection. In order to better understand possible mechanisms of unmasking β (1,3)-glucan, we must develop a deeper comprehension of the mechanism behind unmasking.The phosphatidylserine (PS) synthase enzyme (Cho1) was reported to control β (1,3)-glucan exposure. In this dissertation, I utilized classical genetics and biochemical methods to identify the potential protein(s) involved in causing this phenotype. Several fungal cell wall associated signaling pathways are overly activated when CHO1 is disrupted, including GTPase Cdc42, a central regulator of cellular polarity and morphological development, and its regulated protein kinase Cek1. When Cek1 is activated independent of CHO1, it leads to β (1,3)-glucan exposure. To understand thoroughly how Cek1 and its associated pathway govern β (1,3)-glucan exposure, I further screened the upstream signaling protein(s) causing Cek1 over-activation. A novel signaling cascade was identified where the predicted GTPase activating protein (GAP) Lrg1 represses Cek1 activity by downregulating the GTPase Cdc42 and its downstream MAPKKK, Ste11. The consequences to virulence for upregulation of Cek1 are that pathogenicity is diminished in the mouse model of systemic infection, and this correlates with increased cytokine responses from macrophages. Data from RNA-sequencing demonstrates that a number of cell wall associated genes are significantly up-regulated transcriptionally when Cek1 is hyper-activated, which might be responsible for the cell wall exposure. Thus, we propose a model that Cek1 hyperactivation causes β (1,3)-glucan exposure by upregulating cell wall proteins and leads to a more robust immune detection in vivo, promoting more effective clearance.

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