Nywening, A. | Search bioRxiv

bioRxiv2023

The Aspergillus fumigatus maiA gene contributes to cell wall homeostasis and fungal virulence.

In this study, two distinct in vitro infection models of Aspergillus fumigatus, using murine macrophages (RAW264.7) and human lung epithelial cells (A549), were employed to identify the genes important for fungal adaptation during infection. Transcriptomic analyses of co-incubated Aspergillus uncovered 140 fungal genes up-regulated in common between both models that, when compared with a previously published in vivo transcriptomic study, allowed the identification of 13 genes consistently up-regulated in all three infection conditions. Among them, the maiA gene, responsible for a critical step in the L-phenylalanine degradation pathway, was identified. Disruption of maiA resulted in a mutant strain unable to complete the Phe degradation pathway, leading to an excessive production of pyomelanin when this amino acid served as the sole carbon source. Moreover, the disruption mutant exhibited noticeable cell wall abnormalities, with reduced levels of {beta}-glucans within the cell wall. the maiA-1 mutant strain induced reduced inflammation in primary macrophages and displayed significantly lower virulence in a neutropenic mouse model of infection. This is the first study linking the A. fumigatus maiA gene to fungal cell wall homeostasis and virulence.

microbiology↗

bioRxiv2023

The sterol C-24 methyltransferase encoding gene, erg6, is essential for viability of Aspergillus species.

Ergosterol is a critical component of fungal plasma membranes. Although many currently available antifungal compounds target the ergosterol biosynthesis pathway for antifungal effect, current knowledge regarding ergosterol synthesis remains incomplete for filamentous fungal pathogens like Aspergillus fumigatus. Here, we show for the first time that the lipid droplet-associated sterol C-24 methyltransferase, Erg6, is essential for A. fumigatus viability. We further show that this essentiality extends to additional Aspergillus species, including A. lentulus, A. terreus, and A. nidulans. Neither the overexpression of a putative erg6 paralog, smt1, nor the exogenous addition of ergosterol could rescue erg6 deficiency. Importantly, Erg6 downregulation results in a dramatic decrease in ergosterol and accumulation in lanosterol and is further characterized by diminished sterol-rich plasma membrane domains (SRDs) at hyphal tips. Unexpectedly, erg6 repressed strains demonstrate wild-type susceptibility against the ergosterol-active triazole and polyene antifungals. Finally, repressing erg6 expression reduced fungal burden accumulation in a murine model of invasive aspergillosis. Taken together, our studies suggest that Erg6, which shows little homology to mammalian proteins, is potentially an attractive antifungal drug target for therapy of Aspergillus infections. IMPORTANCEA. fumigatus is the most common pathogen that causes invasive aspergillosis, a life-threatening fungal infection with more than 300,000 cases reported annually. Available antifungals to treat Aspergillus-related infection are limited to three drug classes targeting the plasma membrane (ergosterol) or the cell wall, each of which suffer from either host toxicity or rising resistance levels. As ergosta-type sterols are absent in mammalian cells but are essential for fungal viability, the ergosterol biosynthesis pathway remains an enticing target for the development of new antifungals. Although ergosterol biosynthesis has been well studied in model yeast, only a few genes have been genetically characterized in A. fumigatus. Here, we characterize Erg6, one of the fungus-specific sterol biosynthesis genes, as an essential gene in Aspergillus species. We further provide in vivo evidence of the importance of Erg6 for establishment of invasive aspergillosis. Given the importance of Erg6 in other fungal systems for growth, stress resistance, and virulence, our study suggests that development of Erg6 inhibitors may be a promising strategy for developing novel broad-spectrum antifungals.

microbiology↗