Multiple interfaces control activity of the Candida glabrata Pdr1 transcription factor mediating azole drug resistance

ABSTRACTThe widespread emergence of antifungal drug resistance poses a severe clinical problem. Though predicted to play a role in this phenomenon, the drug:H+antiporters (DHA) of the major facilitator superfamily have largely escaped characterization in pathogenic yeasts. This work describes the first DHA from the pathogenic yeastCandida glabratareported to be involved in antifungal drug resistance, theC. glabrata QDR2(CgQDR2) gene (ORFCAGL0G08624g). The expression ofCgQDR2inC. glabratawas found to confer resistance to the antifungal drugs miconazole, tioconazole, clotrimazole, and ketoconazole. By use of a green fluorescent protein (GFP) fusion, the CgQdr2 protein was found to be targeted to the plasma membrane inC. glabrata. In agreement with these observations,CgQDR2expression was found to decrease the intracellular accumulation of radiolabeled clotrimazole inC. glabrataand to play a role in the extrusion of this antifungal from preloaded cells. Interestingly, the functional heterologous expression ofCgQDR2in the model yeastSaccharomyces cerevisiaefurther confirmed the role of this gene as a multidrug resistance determinant: its expression was able to complement the susceptibility phenotype exhibited by itsS. cerevisiaehomologue,QDR2, in the presence of imidazoles and of the antimalarial and antiarrhythmic drug quinidine. In contrast to the findings reported for Qdr2, CgQdr2 expression does not contribute to the ability of yeast to grow under K+-limiting conditions. Interestingly,CgQDR2transcript levels were seen to be upregulated inC. glabratacells challenged with clotrimazole or quinidine. This upregulation was found to depend directly on the transcription factor CgPdr1, the major regulator of multidrug resistance in this pathogenic yeast, which has also been found to be a determinant of quinidine and clotrimazole resistance inC. glabrata.

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Temporal transcriptional response of Candida glabrata during macrophage infection reveals a multifaceted transcriptional regulator CgXbp1 important for macrophage response and drug resistance

10.1101/2021.09.28.462173 ◽

2021 ◽

Author(s):

Maruti Nandan Rai ◽

Chirag Parsania ◽

Rikky Rai ◽

Niranjan Shirgaonkar ◽

Kaeling Tan ◽

...

Keyword(s):

Drug Resistance ◽

Transcription Factor ◽

Rna Polymerase Ii ◽

Candida Glabrata ◽

Drug Tolerance ◽

High Temporal Resolution ◽

Macrophage Infection ◽

Transcriptional Responses ◽

Macrophage Response ◽

Genome Wide

AbstractCandida glabrata can thrive inside macrophages and tolerate high levels of azole antifungals. These innate abilities render infections by this human pathogen a clinical challenge. How C. glabrata reacts inside macrophages and what is the molecular basis of its drug tolerance are not well understood. Here, we mapped genome-wide RNA polymerase II (RNAPII) occupancy in C. glabrata to delineate its transcriptional responses during macrophage infection in high temporal resolution. RNAPII profiles revealed dynamic C. glabrata responses to macrophage with genes of specialized pathways activated chronologically at different times of infection. We identified an uncharacterized transcription factor (CgXbp1) important for the chronological macrophage response, survival in macrophages, and virulence. Genome-wide mapping of CgXbp1 direct targets further revealed its multi-faceted functions, regulating not only virulence-related genes but also genes associated with drug resistance. Finally, we showed that CgXbp1 indeed also affects azole resistance. Overall, this work presents a powerful approach for examining host-pathogen interaction and uncovers a novel transcription factor important for C. glabrata’s survival in macrophages and drug tolerance.

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Faculty Opinions recommendation of Prevalent mutator genotype identified in fungal pathogen Candida glabrata promotes multi-drug resistance.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.726252186.793516529 ◽

2016 ◽

Author(s):

Leah Cowen ◽

Michelle Leach

Keyword(s):

Drug Resistance ◽

Fungal Pathogen ◽

Candida Glabrata ◽

Multi Drug Resistance

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New Insights into the Pleiotropic Drug Resistance Network from Genome-Wide Characterization of the YRR1 Transcription Factor Regulation System

Molecular and Cellular Biology ◽

10.1128/mcb.22.8.2642-2649.2002 ◽

2002 ◽

Vol 22 (8) ◽

pp. 2642-2649 ◽

Cited By ~ 68

Author(s):

Stéphane Le Crom ◽

Frédéric Devaux ◽

Philippe Marc ◽

Xiaoting Zhang ◽

W. Scott Moye-Rowley ◽

...

Keyword(s):

Drug Resistance ◽

Transcription Factor ◽

Binding Site ◽

Time Course ◽

Target Genes ◽

Regulation System ◽

Dependent Manner ◽

Pleiotropic Drug Resistance ◽

Genome Wide

ABSTRACT Yrr1p is a recently described Zn2Cys6 transcription factor involved in the pleiotropic drug resistance (PDR) phenomenon. It is controlled in a Pdr1p-dependent manner and is autoregulated. We describe here a new genome-wide approach to characterization of the set of genes directly regulated by Yrr1p. We found that the time-course production of an artificial chimera protein containing the DNA-binding domain of Yrr1p activated the 15 genes that are also up-regulated by a gain-of-function mutant of Yrr1p. Gel mobility shift assays showed that the promoters of the genes AZR1, FLR1, SNG1, YLL056C, YLR346C, and YPL088W interacted with Yrr1p. The putative consensus Yrr1p binding site deduced from these experiments, (T/A)CCG(C/T)(G/T)(G/T)(A/T)(A/T), is strikingly similar to the PDR element binding site sequence recognized by Pdr1p and Pdr3p. The minor differences between these sequences are consistent with Yrr1p and Pdr1p and Pdr3p having different sets of target genes. According to these data, some target genes are directly regulated by Pdr1p and Pdr3p or by Yrr1p, whereas some genes are indirectly regulated by the activation of Yrr1p. Some genes, such as YOR1, SNQ2, and FLR1, are clearly directly controlled by both classes of transcription factor, suggesting an important role for the corresponding membrane proteins.

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Dysfunction of Prohibitin 2 Results in Reduced Susceptibility to Multiple Antifungal Drugs via Activation of the Oxidative Stress-Responsive Transcription Factor Pap1 in Fission Yeast

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00860-18 ◽

2018 ◽

Vol 62 (11) ◽

Cited By ~ 1

Author(s):

Qiannan Liu ◽

Fan Yao ◽

Guanglie Jiang ◽

Min Xu ◽

Si Chen ◽

...

Keyword(s):

Oxidative Stress ◽

Drug Resistance ◽

Transcription Factor ◽

Fission Yeast ◽

Mitochondrial Protein ◽

Antifungal Drugs ◽

Antifungal Drug ◽

Gene Encoding ◽

Content Type ◽

Antifungal Drug Resistance

ABSTRACT The fight against resistance to antifungal drugs requires a better understanding of the underlying cellular mechanisms. In order to gain insight into the mechanisms leading to antifungal drug resistance, we performed a genetic screen on a model organism, Schizosaccharomyces pombe, to identify genes whose overexpression caused resistance to antifungal drugs, including clotrimazole and terbinafine. We identified the phb2+ gene, encoding a highly conserved mitochondrial protein, prohibitin (Phb2), as a novel determinant of reduced susceptibility to multiple antifungal drugs. Unexpectedly, deletion of the phb2+ gene also exhibited antifungal drug resistance. Overexpression of the phb2+ gene failed to cause drug resistance when the pap1+ gene, encoding an oxidative stress-responsive transcription factor, was deleted. Furthermore, pap1+ mRNA expression was significantly increased when the phb2+ gene was overexpressed or deleted. Importantly, either overexpression or deletion of the phb2+ gene stimulated the synthesis of NO and reactive oxygen species (ROS), as measured by the cell-permeant fluorescent NO probe DAF-FM DA (4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate) and the ROS probe DCFH-DA (2′,7′-dichlorodihydrofluorescein diacetate), respectively. Taken together, these results suggest that Phb2 dysfunction results in reduced susceptibility to multiple antifungal drugs by increasing NO and ROS synthesis due to dysfunctional mitochondria, thereby activating the transcription factor Pap1 in fission yeast.

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The AP-1 transcription factor JunB is essential for multiple myeloma cell proliferation and drug resistance in the bone marrow microenvironment

Leukemia ◽

10.1038/leu.2016.358 ◽

2016 ◽

Vol 31 (7) ◽

pp. 1570-1581 ◽

Cited By ~ 23

Author(s):

F Fan ◽

M H Bashari ◽

E Morelli ◽

G Tonon ◽

S Malvestiti ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Cell Proliferation ◽

Drug Resistance ◽

Transcription Factor ◽

Myeloma Cell ◽

Bone Marrow Microenvironment ◽

Multiple Myeloma Cell

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Clitocine Reversal of P-Glycoprotein Associated Multi-Drug Resistance through Down-Regulation of Transcription Factor NF-κB in R-HepG2 Cell Line

PLoS ONE ◽

10.1371/journal.pone.0040720 ◽

2012 ◽

Vol 7 (8) ◽

pp. e40720 ◽

Cited By ~ 41

Author(s):

Jianguo Sun ◽

Chilam Au Yeung ◽

Ngai Na Co ◽

Tsun Yee Tsang ◽

Esmond Yau ◽

...

Keyword(s):

Drug Resistance ◽

Transcription Factor ◽

Cell Line ◽

Hepg2 Cell ◽

Multi Drug Resistance ◽

Regulation Of Transcription ◽

Hepg2 Cell Line ◽

Down Regulation ◽

P Glycoprotein

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Abstract 4409: Cholesterol pathway determines ovarian cancer drug resistance through transcription factor SREBP2

10.1158/1538-7445.am2017-4409 ◽

2017 ◽

Cited By ~ 1

Author(s):

Galina Karashchuk ◽

Nataliya Karashchuk ◽

Signe Caksa ◽

Tyler S. Smith ◽

Alexander S. Brodsky

Keyword(s):

Ovarian Cancer ◽

Drug Resistance ◽

Transcription Factor ◽

Cancer Drug ◽

Cancer Drug Resistance

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Upregulation of the Adhesin Gene EPA1 Mediated by PDR1 in Candida glabrata Leads to Enhanced Host Colonization

mSphere ◽

10.1128/msphere.00065-15 ◽

2016 ◽

Vol 1 (2) ◽

Cited By ~ 19

Author(s):

Luis A. Vale-Silva ◽

Beat Moeckli ◽

Riccardo Torelli ◽

Brunella Posteraro ◽

Maurizio Sanguinetti ◽

...

Keyword(s):

Drug Resistance ◽

Epithelial Cells ◽

Candida Glabrata ◽

Resistance Mechanism ◽

Cell Types ◽

Host Cells ◽

Regulation Mechanism ◽

Content Type

ABSTRACT Candida glabrata is an important fungal pathogen in human diseases and is also rapidly acquiring drug resistance. Drug resistance can be mediated by the transcriptional activator PDR1, and this results in the upregulation of multidrug transporters. Intriguingly, this resistance mechanism is associated in C. glabrata with increased virulence in animal models and also with increased adherence to specific host cell types. The C. glabrata adhesin gene EPA1 is a major contributor of virulence and adherence to host cells. Here, we show that EPA1 expression is controlled by PDR1 independently of subtelomeric silencing, a known EPA1 regulation mechanism. Thus, a relationship exists between PDR1, EPA1 expression, and adherence to host cells, which is critical for efficient virulence. Our results demonstrate that acquisition of drug resistance is beneficial for C. glabrata in fungus-host relationships. These findings further highlight the challenges of the therapeutic management of C. glabrata infections in human patients. Candida glabrata is the second most common Candida species causing disseminated infection, after C. albicans. C. glabrata is intrinsically less susceptible to the widely used azole antifungal drugs and quickly develops secondary resistance. Resistance typically relies on drug efflux with transporters regulated by the transcription factor Pdr1. Gain-of-function (GOF) mutations in PDR1 lead to a hyperactive state and thus efflux transporter upregulation. Our laboratory has characterized a collection of C. glabrata clinical isolates in which azole resistance was found to correlate with increased virulence in vivo. Contributing phenotypes were the evasion of adhesion and phagocytosis by macrophages and an increased adhesion to epithelial cells. These phenotypes were found to be dependent on PDR1 GOF mutation and/or C. glabrata strain background. In the search for the molecular effectors, we found that PDR1 hyperactivity leads to overexpression of specific cell wall adhesins of C. glabrata. Further study revealed that EPA1 regulation, in particular, explained the increase in adherence to epithelial cells. Deleting EPA1 eliminates the increase in adherence in an in vitro model of interaction with epithelial cells. In a murine model of urinary tract infection, PDR1 hyperactivity conferred increased ability to colonize the bladder and kidneys in an EPA1-dependent way. In conclusion, this study establishes a relationship between PDR1 and the regulation of cell wall adhesins, an important virulence attribute of C. glabrata. Furthermore, our data show that PDR1 hyperactivity mediates increased adherence to host epithelial tissues both in vitro and in vivo through upregulation of the adhesin gene EPA1. IMPORTANCE Candida glabrata is an important fungal pathogen in human diseases and is also rapidly acquiring drug resistance. Drug resistance can be mediated by the transcriptional activator PDR1, and this results in the upregulation of multidrug transporters. Intriguingly, this resistance mechanism is associated in C. glabrata with increased virulence in animal models and also with increased adherence to specific host cell types. The C. glabrata adhesin gene EPA1 is a major contributor of virulence and adherence to host cells. Here, we show that EPA1 expression is controlled by PDR1 independently of subtelomeric silencing, a known EPA1 regulation mechanism. Thus, a relationship exists between PDR1, EPA1 expression, and adherence to host cells, which is critical for efficient virulence. Our results demonstrate that acquisition of drug resistance is beneficial for C. glabrata in fungus-host relationships. These findings further highlight the challenges of the therapeutic management of C. glabrata infections in human patients.

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