Candida glabrata is the second leading cause of hospital-acquired systemic Candida infections. The prevalence of drug resistant C. glabrata infections has increased over the past 30 years, and this has contributed to high mortality rates among immunocompromised and immunosuppressed populations. Resistance to azole-class antifungals in C. glabrata and Saccharomyces cerevisiae is mediated by Pdr1, a Zn(II)2Cys6 transcription factor that initiates the transcription of drug efflux pump genes. Many Zn(II)2Cys6 transcription factors are regulated by phosphorylation, and we hypothesized that Pdr1 activity is regulated by phosphorylation as well. To test this hypothesis, we predicted phosphosites in C. glabrata Pdr1 using the NetPhos Yeast server and selected high-probability sites based on comparison with known phosphosites in S. cerevisiae Pdr1. A pdr1Δ C. glabrata strain was complemented with Pdr1 variants containing alanine or glutamic acid at predicted phosphosites. Spot dilution assays and agar gradient diffusion assays identified 5 alanine variants and 2 glutamic acid variants that restored sensitivity to fluconazole with minimum inhibitory concentrations similar to the pdr1Δ strain. Currently, mass spectrometry phosphoproteomics is being used to confirm these locations as phosphosites and to identify additional sites for investigation. Future work will characterize how phosphorylation affects Pdr1 localization, protein-protein interactions, and protein-DNA interactions.

Mutation of predicted Pdr1 phosphosites restores fluconazole sensitivity in Candida glabrata

Where thousands of chemistry professionals meet to share ideas and advance scientific and technical knowledge. Spring 2023 abstracts, posters, and presentations will launch at the end of March.

ACS Spring 2023

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.