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Fluconazole-induced actin cytoskeleton remodeling requires phosphatidylinositol 3-phosphate 5-kinase in the pathogenic yeast Candida glabrata.

ABSTRACT: Known azole antifungal resistance mechanisms include mitochondrial dysfunction and overexpression of the sterol biosynthetic target enzyme and multidrug efflux pumps. Here, we identify, through a genetic screen, the vacuolar membrane-resident phosphatidylinositol 3-phosphate 5-kinase (CgFab1) to be a novel determinant of azole tolerance. We demonstrate for the first time that fluconazole promotes actin cytoskeleton reorganization in the emerging, inherently less azole-susceptible fungal pathogen Candida glabrata, and genetic or chemical perturbation of actin structures results in intracellular sterol accumulation and azole susceptibility. Further, CgFAB1 disruption impaired vacuole homeostasis and actin organization, and the F-actin-stabilizing compound jasplakinolide rescued azole toxicity in cytoskeleton defective-mutants including the Cgfab1? mutant. In vitro assays revealed that the actin depolymerization factor CgCof1 binds to multiple lipids including phosphatidylinositol 3,5-bisphosphate. Consistently, CgCof1 distribution along with the actin filament-capping protein CgCap2 was altered upon both CgFAB1 disruption and fluconazole exposure. Altogether, these data implicate CgFab1 in azole tolerance through actin network remodeling. Finally, we also show that actin polymerization inhibition rendered fluconazole fully and partially fungicidal in azole-susceptible and azole-resistant C. glabrata clinical isolates, respectively, thereby, underscoring the role of fluconazole-effectuated actin remodeling in azole resistance.

SUBMITTER: Bhakt P

PROVIDER: S-EPMC6221164 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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Fluconazole-induced actin cytoskeleton remodeling requires phosphatidylinositol 3-phosphate 5-kinase in the pathogenic yeast Candida glabrata.

Bhakt Priyanka P Shivarathri Raju R Choudhary Deepak Kumar DK Borah Sapan S Kaur Rupinder R

Molecular microbiology 20181003 3

Known azole antifungal resistance mechanisms include mitochondrial dysfunction and overexpression of the sterol biosynthetic target enzyme and multidrug efflux pumps. Here, we identify, through a genetic screen, the vacuolar membrane-resident phosphatidylinositol 3-phosphate 5-kinase (CgFab1) to be a novel determinant of azole tolerance. We demonstrate for the first time that fluconazole promotes actin cytoskeleton reorganization in the emerging, inherently less azole-susceptible fungal pathogen ...[more]

PMID: 30137648

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Project description:The high prevalence of fluconazole resistance among clinical isolates of Candida glabrata has greatly hampered the utility of fluconazole for the treatment of invasive candidiasis. Fluconazole resistance in this yeast is almost exclusively due to activating mutations in the transcription factor Pdr1, which result in upregulation of the ABC transporter genes CDR1, PDH1, and SNQ2 and therefore increased fluconazole efflux. However, the regulation of Pdr1 is poorly understood. In order to identify genes that interact with the Pdr1 transcriptional pathway and influence the susceptibility of C. glabrata to fluconazole, we screened a collection of deletion mutants for those exhibiting increased resistance to fluconazole. Deletion of the gene coding for a protein homologous to the Saccharomyces cerevisiae J protein Jjj1 resulted in decreased fluconazole susceptibility. We used the SAT1 flipper method to generate independent deletion mutants for JJJ1 in an SDD clinical isolate. Expression of both CDR1 and PDR1 was increased in the absence of JJJ1. In the absence of CDR1 or PDR1, deletion of JJJ1 has only a modest effect on fluconazole susceptibility. Transcriptional profiling using transcriptome sequencing (RNA-seq) revealed upregulation of genes of the Pdr1 regulon in the absence of JJJ1. Jjj1 appears to be a negative regulator of fluconazole resistance in C. glabrata and acts primarily through upregulation of the ABC transporter gene CDR1 via activation of the Pdr1 transcriptional pathway. IMPORTANCECandida glabrata is the second most common species of Candida recovered from patients with invasive candidiasis. The increasing number of infections due to C. glabrata, combined with its high rates of resistance to the commonly used, well-tolerated azole class of antifungal agents, has limited the use of this antifungal class. This has led to the preferential use of echinocandins as empirical treatment for serious Candida infections. The primary mechanism of resistance found in clinical isolates is the presence of an activating mutation in the gene encoding the transcription factor Pdr1 that results in upregulation of one or more of the efflux pumps Cdr1, Pdh1, and Snq2. By developing a better understanding of this mechanism of resistance to the azoles, it will be possible to develop strategies for reclaiming the utility of the azole antifungals against this important fungal pathogen.

Dataset Information

Fluconazole-induced actin cytoskeleton remodeling requires phosphatidylinositol 3-phosphate 5-kinase in the pathogenic yeast Candida glabrata.

Publications

Fluconazole-induced actin cytoskeleton remodeling requires phosphatidylinositol 3-phosphate 5-kinase in the pathogenic yeast Candida glabrata.

Similar Datasets

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