Aspergillus fumigatus Transcription Factors Involved in the Caspofungin Paradoxical Effect

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Valero, Clara ; Colabardini, Ana Cristina ; Chiaratto, Jessica ; Pardeshi, Lakhansing ; de Castro, Patricia Alves ; Ferreira Filho, Jaire Alves ; Silva, Lilian Pereira ; Rocha, Marina Campos ; Malavazi, Iran ; Costa, Jonas Henrique ; Fill, Taicia ; Barros, Mario Henrique ; Wong, Sarah Sze Wah ; Aimanianda, Vishukumar ; Wong, Koon Ho ; Goldman, Gustavo H.

• Document type: Journal article
• Source: MBIO; v. 11, n. 3, p. 22-pg., 2020-05-01.

Abstract

Aspergillus fumigatus is the leading cause of pulmonary fungal diseases. Azoles have been used for many years as the main antifungal agents to treat and prevent invasive aspergillosis. However, in the last 10 years there have been several reports of azole resistance in A. fumigatus and new strategies are needed to combat invasive aspergillosis. Caspofungin is effective against other human-pathogenic fungal species, but it is fungistatic only against A. fumigatus. Resistance to caspofungin in A. fumigatus has been linked to mutations in the fksA gene that encodes the target enzyme of the drug beta-1,3-glucan synthase. However, tolerance of high caspofungin concentrations, a phenomenon known as the caspofungin paradoxical effect (CPE), is also important for subsequent adaptation and drug resistance evolution. Here, we identified and characterized the transcription factors involved in the response to CPE by screening an A. fumigatus library of 484 null transcription factors (TFs) in CPE drug concentrations. We identified 11 TFs that had reduced CPE and that encoded proteins involved in the basal modulation of the RNA polymerase II initiation sites, calcium metabolism, and cell wall remodeling. One of these TFs, FhdA, was important for mitochondrial respiratory function and iron metabolism. The Delta fhdA mutant showed decreased growth when exposed to Congo red or to high temperature. Transcriptome sequencing (RNA-seq) analysis and further experimental validation indicated that the Delta fhdA mutant showed diminished respiratory capacity, probably affecting several pathways related to the caspofungin tolerance and resistance. Our results provide the foundation to understand signaling pathways that are important for caspofungin tolerance and resistance. IMPORTANCE Aspergillus fumigatus, one of the most important human-pathogenic fungal species, is able to cause aspergillosis, a heterogeneous group of diseases that presents a wide range of clinical manifestations. Invasive pulmonary aspergillosis is the most serious pathology in terms of patient outcome and treatment, with a high mortality rate ranging from 50% to 95% primarily affecting immunocompromised patients. Azoles have been used for many years as the main antifungal agents to treat and prevent invasive aspergillosis. However, there were several reports of evolution of clinical azole resistance in the last decade. Caspofungin, a noncompetitive beta-1,3-glucan synthase inhibitor, has been used against A. fumigatus, but it is fungistatic and is recommended as second-line therapy for invasive aspergillosis. More in formation about caspofungin tolerance and resistance is necessary in order to refine antifungal strategies that target the fungal cell wall. Here, we screened a transcription factor (TF) deletion library for TFs that can mediate caspofungin tolerance and resistance. We have identified 11 TFs that are important for caspofungin sensitivity and/or for the caspofungin paradoxical effect (CPE). These TFs encode proteins involved in the basal modulation of the RNA polymerase II initiation sites, calcium metabolism or cell wall remodeling, and mitochondrial respiratory function. The study of those genes regulated by TFs identified in this work will provide a better understanding of the signaling pathways that are important for caspofungin tolerance and resistance. (AU)

• FAPESP's process: 16/21392-2 - "Characterization of transcription factors that have the basic domain" zipper "leucine (bZIP) ATF1 in Aspergillus fumigatus"
• Grantee: Lilian Pereira Silva
• Support Opportunities: Scholarships in Brazil - Doctorate
• FAPESP's process: 17/07536-4 - Molecular Mechanisms Involved in Paradoxal Growth Induced by Caspofungin in Aspergillus fumigatus
• Grantee: Ana Cristina Colabardini
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 16/12948-7 - The role of mitogen-activated protein kinase (MPKA) MpkA in regulating gliotoxin production in Aspergillus fumigatus.
• Grantee: Patrícia Alves de Castro Silva
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 16/07870-9 - The influence of mitogen activated protein kinases (MAPK) on the expression of genetic determinants important for Aspergillus fumigatus virulence
• Grantee: Gustavo Henrique Goldman
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 16/22062-6 - Molecular characterization of htlA and fhdA transcription factors of human pathogenic fungus Aspergillus fumigatus
• Grantee: Jéssica Chiaratto
• Support Opportunities: Scholarships in Brazil - Master
• FAPESP's process: 18/00715-3 - Molecular characterization of Aspergillus fumigatus transcription factors involved in calcium metabolism.
• Grantee: Clara Isabel Valero Fernández
• Support Opportunities: Scholarships in Brazil - Post-Doctoral