Transcriptional Regulation of Chitin Synthases by Calcineurin Controls Paradoxical Growth of Aspergillus fumigatus in Response to Caspofungin

Abstract

Attenuated activity of echinocandin antifungals at high concentrations, known as the "paradoxical effect," is a well-established phenomenon in Candida albicans and Aspergillus fumigatus. In the yeast C. albicans, upregulation of chitin biosynthesis via the protein kinase C (PKC), high-osmolarity glycerol response (HOG), and Ca 2؉ /calcineurin signaling pathways is an important cell wall stress response that permits growth in the presence of high concentrations of echinocandins. However, nothing is known of the molecular mechanisms regulating the mold A. fumigatus and its paradoxical response to echinocandins. Here, we show that the laboratory strain of A. fumigatus and five of seven clinical A. fumigatus isolates tested display various magnitudes of paradoxical growth in response to caspofungin. Interestingly, none of the eight strains showed paradoxical growth in the presence of micafungin or anidulafungin. Treatment of the ⌬cnaA and ⌬crzA strains, harboring gene deletions of the calcineurin A subunit and the calcineurin-dependent transcription factor, respectively, with high concentrations of caspofungin revealed that the A. fumigatus paradoxical effect is calcineurin pathway dependent. Exploring a molecular role for CnaA in the compensatory chitin biosynthetic response, we found that caspofungin treatment resulted in increased chitin synthase gene expression, leading to a calcineurin-dependent increase in chitin synthase activity. Taken together, our data suggest a mechanistic role for A. fumigatus calcineurin signaling in the chitin biosynthetic response observed during paradoxical growth in the presence of high-dose caspofungin treatment.The echinocandin antifungals inhibit 1,3-␤-D-glucan synthesis, and all three currently available echinocandins (caspofungin, anidulafungin, and micafungin) have activity against both yeasts and molds. However, in vitro and in vivo studies with this antifungal class have revealed an interesting "paradoxical effect" on growth, evidenced by a recurrent increase in the growth of the fungal organism at drug concentrations above a certain threshold (36).The paradoxical effect was first described for caspofungin treatment of Candida albicans and was not found to be due to mutation or increased expression of glucan synthase activity (30,32). Further studies showed that this paradoxical effect on growth occurred in many Candida species, appearing in both planktonic and biofilm cells (22). Early efforts to elucidate a mechanism revealed increased chitin content in the cell wall upon caspofungin treatment (31). Initial studies with the Saccharomyces cerevisiae FKS1 deletion mutant, a strain defective in the 1,3-␤-D-glucan synthase, showed that chitin biosynthesis is upregulated in response to glucan depletion (14). Furthermore, at the molecular level, several signal transduction pathways have been implicated in the regulation of the C. albicans paradoxical effect, including protein kinase C (PKC), highosmolarity glycerol response (HOG), and calcineurin signaling events (25,34,35). Alt...

Regulatory Circuitry Governing Fungal Development, Drug Resistance, and Disease

“…When these ⌬cnaA or ⌬crzA mutants are treated with caspofungin, they show even further decreased (1,3)-␤-D-glucan content (123,186). Intriguingly, while wild-type cells compensate for caspofungin-induced (1,3)-␤-D-glucan depletion by increasing chitin synthase gene expression and cell wall chitin contents, the ⌬cnaA and ⌬crzA deletion strains fail to do so (185,186), indicating that calcineurin may regulate compensatory chitin synthesis in the presence of echinocandin stress.…”

Saprotrophic competitiveness and biocontrol fitness of a genetically modified strain of the plant-growth-promoting fungus Trichoderma hamatum GD12

“…2d). This showed that sporulation in the N-acetyl-b-D-glucosaminidase mutant can be induced by exposure to caspofungin, but that the ability to produce spores is lost following release from the drug.Little is known about b-1,3-glucan synthases and sporulation in filamentous fungi, but in the human pathogen Aspergillus fumigatus, exposure to the drug results in upregulation of chitin biosynthetic genes and stimulation of chitin synthesis (Fortwendel et al, 2010). It is reasonable to speculate that a similar process occurs in DThnag : : hph during caspofungin exposure, leading to improved cell wall integrity which supports transient and partial restoration of sporulation.…”

The putative flavin carrier family FlcA-C is important forAspergillus fumigatusvirulence

“…[24][25][26][27][28] In all these fungal pathogens, calcineurin is important for growth, morphology, state transitions, cation homeostasis, stress responses, and cell membrane and cell wall integrity pathways. 1,2,[29][30][31][32][33] Aspergillus fumigatus is an opportunistic fungal pathogen and the most important species causing pulmonary fungal infections. 15,[34][35][36][37][38] Although A. fumigatus is able to cause many clinical forms, the most important is the invasive pulmonary aspergillosis (IPA), that has mortality rates as high as 80% in immunosuppressed patients.…”