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Functional Investigation of Iron-Responsive Microsomal Proteins, including MirC, in Aspergillus fumigatus
Eoin D Mulvihill1, Nicola M Moloney1, Rebecca A Owens1
1Department of Biology, Maynooth University Kildare, Ireland.
Abstract:
The functionality of many microsome-associated proteins which exhibit altered abundance in response to iron limitation in Aspergillus fumigatus is unknown. Here, we generate and characterize eight gene deletion strains, and of most significance reveal that MirC (AFUA_2G05730) contributes to the maintenance of intracellular siderophore [ferricrocin (FC)] levels, augments conidiation, confers protection against oxidative stress, exhibits an intracellular localization and contributes to fungal virulence in the Galleria mellonella animal model system. FC levels were unaffected following deletion of all other genes encoding microsome-associated proteins. MirC does not appear to play a role in either siderophore export from, or uptake into, A. fumigatus. Label-free quantitative proteomic analysis unexpectedly revealed increased abundance of siderophore biosynthetic enzymes. In addition, increased expression of hapX (7.2 and 13.8-fold at 48 and 72 h, respectively; p < 0.001) was observed in ΔmirC compared to wild-type under iron-replete conditions by qRT-PCR. This was complemented by significantly elevated extracellular triacetylfusarinine C (TAFC; p < 0.01) and fusarinine C (FSC; p < 0.05) siderophore secretion. We conclude that MirC plays an important role in FC biosynthesis and contributes to the maintenance of iron homeostasis in A. fumigatus.
Insights
The study identifies MirC as a key protein in Aspergillus fumigatus, crucial for maintaining intracellular siderophore levels, fungal growth, and virulence. MirC is essential for iron homeostasis and ferricrocin biosynthesis.
Area of Science:
- Mycology
- Molecular Biology
- Biochemistry
Background:
- The function of microsome-associated proteins in *Aspergillus fumigatus* under iron limitation is largely unknown.
- Iron homeostasis is critical for fungal growth and virulence.
Purpose of the Study:
- To investigate the role of microsome-associated proteins in *Aspergillus fumigatus* iron metabolism.
- To characterize the function of the MirC protein in fungal physiology and virulence.
Main Methods:
- Generation and characterization of eight gene deletion strains in *A. fumigatus*.
- Quantitative real-time PCR (qRT-PCR) for gene expression analysis.
- Label-free quantitative proteomic analysis.
- Virulence assays using the *Galleria mellonella* model system.
Main Results:
- MirC deletion significantly impacts intracellular ferricrocin levels, conidiation, and oxidative stress resistance.
- MirC is essential for fungal virulence in the *Galleria mellonella* model.
- Proteomic analysis revealed increased siderophore biosynthetic enzymes and elevated TAFC and FSC secretion in Δ*mirC* strains.
- MirC does not appear to be involved in siderophore export or uptake.
Conclusions:
- MirC plays a significant role in ferricrocin biosynthesis and iron homeostasis in *Aspergillus fumigatus*.
- MirC contributes to fungal virulence, oxidative stress tolerance, and conidiation.
- The study highlights MirC as a potential target for antifungal strategies.