On 7th September, 2020 Krisztina Tóthné Szabó defended her PhD thesis entitled “Investigation of a Fungus Specific Protein Phosphatase”

2020, September 11 - 10:00

C. albicans is an opportunistic human pathogen that contains a novel fungus specific Ser/Thr protein phosphatase, called CaPpz1. This enzyme has several important physiological roles such as the regulation of cation homeostasis, cell wall biosynthesis, morphological changes, oxidative stress response, and virulence of the pathogen. In the present study we used modern omics technologies to investigate its further functions and the consequences of phosphatase deletion under oxidative stress.

First, we analyzed the adaptation mechanisms of the phosphatase deletion mutant (KO) compared with WT strain by proteomics. After 2D gel electrophoresis we identified 25 proteins with mass spectrometry, whose phosphorylation levels and/or protein levels altered in the KO strain. The functional analyses of the affected proteins confirmed the involvement of CaPpz1 in protein synthesis, morphology as well as in oxidative stress response. In addition, we proved a novel role for CaPpz1 in biofilm formation.

Second, we applied transcriptomics to investigate how the ppz1 KO and WT strains responded to a short oxidative stress induced by tert-butyl hydroperoxide (tBOOH). We demonstrated that tBOOH treatment reduced the growth rate and blocked the proliferation of the fungal cells without affecting their viability or vitality significantly. Furthermore, upon oxidative stress and/or phosphatase deletion the cells responded by elevated antioxidant enzyme activities and oxidized glutathione concentration. Finally, we carried out a detailed RNA sequencing study. We found that the lack of CaPpz1, or the oxidative treatment alone, have only a small or moderate effect, but together they generate a robust change in the transcriptome, that indicates a strong positive interaction between the two experimental conditions. Based on gene ontology enrichment analysis we selected 44 genes for further RT-qPCR validations. We confirmed the functions of CaPpz1 in transmembrane transport and oxidation-reduction processes. The expression of genes coding for cell surface proteins and cytosolic ribosomal proteins were downregulated by tBOOH, while the amounts of mRNAs associated with transport processes, oxidoreductase activity, and RNA processing were upregulated. All of these changes were enhanced in the KO strain. From these results we conclude that in C. albicans CaPpz1 plays a protective role against oxidative damage. Our data suggest that the specific inhibition of this phosphatase combined with a proper oxidative treatment may be applied as a possible approach to a topical antifungal therapy.


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