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Loss-of-Function ROX1 Mutations Suppress the Fluconazole Susceptibility of upc2AΔ Mutation in Candida glabrata, Implicating Additional Positive Regulators of Ergosterol Biosynthesis

Tomye L. Ollinger, Bao Vu, Daniel Murante, Josie Parker, Lucia Simonicova, Laura Doorley, Mark A. Stamnes, Steven Kelly Orcid Logo, P. David Rogers, W. Scott Moye-Rowley Orcid Logo, Damian J. Krysan Orcid Logo

mSphere, Volume: 6, Issue: 6

Swansea University Authors: Josie Parker, Steven Kelly Orcid Logo

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DOI (Published version): 10.1128/msphere.00830-21

Abstract

wo of the major classes of antifungal drugs in clinical use target ergosterol biosynthesis. Despite its importance, our understanding of the transcriptional regulation of ergosterol biosynthesis genes in pathogenic fungi is essentially limited to the role of hypoxia and sterol-stress-induced transcr...

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Published in: mSphere
ISSN: 2379-5042
Published: American Society for Microbiology 2021
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URI: https://cronfa.swan.ac.uk/Record/cronfa59074
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Despite its importance, our understanding of the transcriptional regulation of ergosterol biosynthesis genes in pathogenic fungi is essentially limited to the role of hypoxia and sterol-stress-induced transcription factors such as Upc2 and Upc2A as well as homologs of sterol response element binding (SREB) factors. To identify additional regulators of ergosterol biosynthesis in Candida glabrata, an important human fungal pathogen with reduced susceptibility to ergosterol biosynthesis inhibitors relative to other Candida spp., we used a serial passaging strategy to isolate suppressors of the fluconazole hypersusceptibility of a upc2A&#x394; deletion mutant. This led to the identification of loss-of-function mutations in two genes: ROX1, the homolog of a hypoxia gene transcriptional suppressor in Saccharomyces cerevisiae, and CST6, a transcription factor that is involved in the regulation of carbon dioxide response in C. glabrata. Here, we describe a detailed analysis of the genetic interaction of ROX1 and UPC2A. In the presence of fluconazole, loss of Rox1 function restores ERG11 expression to the upc2A&#x394; mutant and inhibits the expression of ERG3 and ERG6, leading to increased levels of ergosterol and decreased levels of the toxic sterol 14&#x3B1; methyl-ergosta-8,24(28)-dien-3&#x3B2;, 6&#x3B1;-diol, relative to the upc2A&#x394; mutant. 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spelling 2022-02-01T11:18:04.1170910 v2 59074 2022-01-04 Loss-of-Function ROX1 Mutations Suppress the Fluconazole Susceptibility of upc2AΔ Mutation in Candida glabrata, Implicating Additional Positive Regulators of Ergosterol Biosynthesis e563ed4e1c7db8d1e131fb78a5f8d0d5 Josie Parker Josie Parker true false b17cebaf09b4d737b9378a3581e3de93 0000-0001-7991-5040 Steven Kelly Steven Kelly true false 2022-01-04 FGMHL wo of the major classes of antifungal drugs in clinical use target ergosterol biosynthesis. Despite its importance, our understanding of the transcriptional regulation of ergosterol biosynthesis genes in pathogenic fungi is essentially limited to the role of hypoxia and sterol-stress-induced transcription factors such as Upc2 and Upc2A as well as homologs of sterol response element binding (SREB) factors. To identify additional regulators of ergosterol biosynthesis in Candida glabrata, an important human fungal pathogen with reduced susceptibility to ergosterol biosynthesis inhibitors relative to other Candida spp., we used a serial passaging strategy to isolate suppressors of the fluconazole hypersusceptibility of a upc2AΔ deletion mutant. This led to the identification of loss-of-function mutations in two genes: ROX1, the homolog of a hypoxia gene transcriptional suppressor in Saccharomyces cerevisiae, and CST6, a transcription factor that is involved in the regulation of carbon dioxide response in C. glabrata. Here, we describe a detailed analysis of the genetic interaction of ROX1 and UPC2A. In the presence of fluconazole, loss of Rox1 function restores ERG11 expression to the upc2AΔ mutant and inhibits the expression of ERG3 and ERG6, leading to increased levels of ergosterol and decreased levels of the toxic sterol 14α methyl-ergosta-8,24(28)-dien-3β, 6α-diol, relative to the upc2AΔ mutant. Our observations establish that Rox1 is a negative regulator of ERG gene biosynthesis and indicate that a least one additional positive transcriptional regulator of ERG gene biosynthesis must be present in C. glabrata. Journal Article mSphere 6 6 American Society for Microbiology 2379-5042 fluconazole, Candida glabrata, ergosterol, antifungal drug resistance 22 12 2021 2021-12-22 10.1128/msphere.00830-21 COLLEGE NANME Medicine, Health and Life Science - Faculty COLLEGE CODE FGMHL Swansea University This work was supported by NIH grants 5R01AI52494 (W.S.M.-R.) and 7R01AI131620 (P.D.R.). 2022-02-01T11:18:04.1170910 2022-01-04T16:49:49.8357327 Faculty of Medicine, Health and Life Sciences Swansea University Medical School - Medicine Tomye L. Ollinger 1 Bao Vu 2 Daniel Murante 3 Josie Parker 4 Lucia Simonicova 5 Laura Doorley 6 Mark A. Stamnes 7 Steven Kelly 0000-0001-7991-5040 8 P. David Rogers 9 W. Scott Moye-Rowley 0000-0002-7163-1120 10 Damian J. Krysan 0000-0001-6330-3365 11 59074__22015__aeccf051869d4e288fc8b5125c377b1f.pdf 59074.pdf 2022-01-04T16:52:22.9155572 Output 2590118 application/pdf Version of Record true © 2021 Ollinger et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. true eng https://creativecommons.org/licenses/by/4.0/
title Loss-of-Function ROX1 Mutations Suppress the Fluconazole Susceptibility of upc2AΔ Mutation in Candida glabrata, Implicating Additional Positive Regulators of Ergosterol Biosynthesis
spellingShingle Loss-of-Function ROX1 Mutations Suppress the Fluconazole Susceptibility of upc2AΔ Mutation in Candida glabrata, Implicating Additional Positive Regulators of Ergosterol Biosynthesis
Josie Parker
Steven Kelly
title_short Loss-of-Function ROX1 Mutations Suppress the Fluconazole Susceptibility of upc2AΔ Mutation in Candida glabrata, Implicating Additional Positive Regulators of Ergosterol Biosynthesis
title_full Loss-of-Function ROX1 Mutations Suppress the Fluconazole Susceptibility of upc2AΔ Mutation in Candida glabrata, Implicating Additional Positive Regulators of Ergosterol Biosynthesis
title_fullStr Loss-of-Function ROX1 Mutations Suppress the Fluconazole Susceptibility of upc2AΔ Mutation in Candida glabrata, Implicating Additional Positive Regulators of Ergosterol Biosynthesis
title_full_unstemmed Loss-of-Function ROX1 Mutations Suppress the Fluconazole Susceptibility of upc2AΔ Mutation in Candida glabrata, Implicating Additional Positive Regulators of Ergosterol Biosynthesis
title_sort Loss-of-Function ROX1 Mutations Suppress the Fluconazole Susceptibility of upc2AΔ Mutation in Candida glabrata, Implicating Additional Positive Regulators of Ergosterol Biosynthesis
author_id_str_mv e563ed4e1c7db8d1e131fb78a5f8d0d5
b17cebaf09b4d737b9378a3581e3de93
author_id_fullname_str_mv e563ed4e1c7db8d1e131fb78a5f8d0d5_***_Josie Parker
b17cebaf09b4d737b9378a3581e3de93_***_Steven Kelly
author Josie Parker
Steven Kelly
author2 Tomye L. Ollinger
Bao Vu
Daniel Murante
Josie Parker
Lucia Simonicova
Laura Doorley
Mark A. Stamnes
Steven Kelly
P. David Rogers
W. Scott Moye-Rowley
Damian J. Krysan
format Journal article
container_title mSphere
container_volume 6
container_issue 6
publishDate 2021
institution Swansea University
issn 2379-5042
doi_str_mv 10.1128/msphere.00830-21
publisher American Society for Microbiology
college_str Faculty of Medicine, Health and Life Sciences
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hierarchy_top_id facultyofmedicinehealthandlifesciences
hierarchy_top_title Faculty of Medicine, Health and Life Sciences
hierarchy_parent_id facultyofmedicinehealthandlifesciences
hierarchy_parent_title Faculty of Medicine, Health and Life Sciences
department_str Swansea University Medical School - Medicine{{{_:::_}}}Faculty of Medicine, Health and Life Sciences{{{_:::_}}}Swansea University Medical School - Medicine
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description wo of the major classes of antifungal drugs in clinical use target ergosterol biosynthesis. Despite its importance, our understanding of the transcriptional regulation of ergosterol biosynthesis genes in pathogenic fungi is essentially limited to the role of hypoxia and sterol-stress-induced transcription factors such as Upc2 and Upc2A as well as homologs of sterol response element binding (SREB) factors. To identify additional regulators of ergosterol biosynthesis in Candida glabrata, an important human fungal pathogen with reduced susceptibility to ergosterol biosynthesis inhibitors relative to other Candida spp., we used a serial passaging strategy to isolate suppressors of the fluconazole hypersusceptibility of a upc2AΔ deletion mutant. This led to the identification of loss-of-function mutations in two genes: ROX1, the homolog of a hypoxia gene transcriptional suppressor in Saccharomyces cerevisiae, and CST6, a transcription factor that is involved in the regulation of carbon dioxide response in C. glabrata. Here, we describe a detailed analysis of the genetic interaction of ROX1 and UPC2A. In the presence of fluconazole, loss of Rox1 function restores ERG11 expression to the upc2AΔ mutant and inhibits the expression of ERG3 and ERG6, leading to increased levels of ergosterol and decreased levels of the toxic sterol 14α methyl-ergosta-8,24(28)-dien-3β, 6α-diol, relative to the upc2AΔ mutant. Our observations establish that Rox1 is a negative regulator of ERG gene biosynthesis and indicate that a least one additional positive transcriptional regulator of ERG gene biosynthesis must be present in C. glabrata.
published_date 2021-12-22T04:16:06Z
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