Biology subjects

Brazil, S.

Publications and source records attributed to Brazil, S..

3 recordsLinked to original sources

bioRxiv2025

Rho2-dependent cell wall remodeling boosts the fungistatic activity of manogepix

Fosmanogepix is a new antifungal agent currently undergoing phase 3 clinical trials. Its active moiety, manogepix, inhibits Gwt1 which is an enzyme essential for the assembly and attachment of glycosylphosphatidylinositol anchors to cell wall proteins. Manogepix has a strong fungistatic activity against most human pathogenic fungal species. Here we characterized the activity of manogepix against the major human pathogenic mold Aspergillus fumigatus. We show that manogepix susceptibility is linked to the expression of gwt1, the gene encoding Gwt1, thereby demonstrating that overexpression of gwt1 can confer resistance. In agreement with a previous study conducted with a different fungal pathogen, we observed an increase of cell well chitin after manogepix treatment. Using a luciferase-based reporter assay, we show that manogepix activates the cell wall integrity stress response pathway. However, manogepix only occasionally causes cell lysis. Mutants that lack the cell wall stress sensor Wsc1 or the Rho GTPase Rho4, which is important for septum formation, are slightly more susceptible to manogepix. In contrast, mutants that lack the Rho GTPase Rho2 or the cell wall stress sensor MidA, both of which are key for A. fumigatus to survive cell wall stress caused by granulocytes, heat and other cell wall perturbing chemical compounds, grow better than wild-type when exposed to manogepix. We show that both mutants, especially {Delta}rho2 but also {Delta}midA, fail to upregulate cell wall chitin to wild-type-like levels in response to manogepix. These results implicate a role for the Rho2-dependent stress response in the fungistatic activity of manogepix against A. fumigatus.

microbiology↗

bioRxiv2025

Rho2 regulates granulocyte-triggered stress adaptation and cell wall remodeling in Aspergillus fumigatus

The airborne opportunistic fungal pathogen Aspergillus fumigatus poses a deadly threat to immunocompromised patients. Neutrophil granulocytes play a key role in the defense against invasive infections caused by this pathogen. The mechanisms by which Aspergillus defends itself against attacks by the immune system are only partially understood. Here we show that human granulocytes activate the cell wall integrity (CWI) pathway of A. fumigatus and that key components of the CWI such as the cell wall stress sensor MidA and the Rho GTPases Rho2 and Rho4 are important for the survival of Aspergillus hyphae under granulocyte attacks. A more detailed investigation of the role of Rho2 revealed that a mutant lacking rho2 is less virulent in a Galleria mellonella infection model. Overexpression of Rho2 increases the resistance of A. fumigatus hyphae to killing by granulocytes. While a mutant lacking Rho2 has a normal cell wall composition, overexpression or constitutive activation of Rho2 leads to an altered cell wall composition and impairs growths of the pathogen. The fungicidal effect of constitutive activation of Rho2 signaling, which correlates with the formation of cell wall chitin bulges, depends on the CWI MAP kinase MpkA. However, Rho2 itself does not appear to be a direct activator of the CWI MAP kinase module. Our results support a model where Rho2 in A. fumigatus actively counteracts granulocyte attacks by upregulating cell wall biosynthesis, thereby strengthening the cell wall and aiding the fungus in surviving the stress condition.

microbiology↗

bioRxiv2024

Toxic eburicol accumulation drives the antifungal activity of azoles against Aspergillus fumigatus

Azole antifungals inhibit the sterol C14-demethylase (CYP51/Erg11), a key enzyme in the ergosterol biosynthesis pathway. They have fungistatic effects against yeasts but fungicidal effects against molds. The molecular basis for this difference remained unknown. The sequence of enzymatic steps required for ergosterol biosynthesis is different in yeasts and molds. Here we show that the azole-induced synthesis of fungicidal cell wall carbohydrate patches in the pathogenic mold Aspergillus fumigatus strictly correlates with the accumulation of the CYP51 substrate eburicol. A lack of other essential ergosterol biosynthesis enzymes, such as sterol C24-methyltransferase (Erg6A), squalene synthase (Erg9) or squalene epoxidase (Erg1) does not result in comparable cell wall alterations. Partial repression of Erg6A, which converts lanosterol into eburicol, increases azole resistance. The sterol C5-desaturase (ERG3)-dependent conversion of eburicol into 14-methylergosta-8,24(28)-dien-3{beta},6-diol, the "toxic diol" responsible for the antifungal effects of azoles in yeasts, is not required for the fungicidal effects in A. fumigatus. In contrast to yeast, where a lack of ERG3 functionality causes azole resistance, A. fumigatus lacking ERG3 becomes more azole susceptible. Mutants lacking mitochondrial complex III functionality, which are less susceptible to the fungicidal effects of azoles, but get strongly inhibited in growth, convert eburicol much more efficiently into the supposedly "toxic diol". Our results support a mechanistic model where the mode of action of azoles against the pathogenic mold A. fumigatus, other than in yeast, relies on the accumulation of eburicol which exerts fungicidal effects by triggering the formation of cell wall carbohydrate patches.

microbiology↗