The Aspergillus fumigatus transcription factor AceA is involved not only in Cu but also in Zn detoxification through regulating transporters CrpA and ZrcA.

Copper (Cu) and zinc (Zn) are essential nutrients for both pathogens and hosts; however, their excess accumulation is toxic for all cells. The Aspergillus transcription factor AceA has been identified having function for Cu detoxification through up-regulation of the expression of the P-type ATPase, CrpA. Here, we demonstrate that Aspergillus fumigatus CrpA is involved in both Cu and Zn detoxification and a putative metallothionein AfCrdA plays a major function in Cu detoxification, and a putative transporter AfZrcA has a dominant role in Zn detoxification, but all three members are transcriptionally dependent on AfAceA. Moreover, the Cys, RGHR, and KGRP motifs in the conserved N-terminus of AfAceA are essential, but not sufficient for AfAceA-mediated Cu and Zn tolerance. Our findings suggest that fungal pathogens have developed very precise systems with overlapping machinery to respond to the two different metal stressors. Meanwhile, there is separate specific machinery for Zn detoxification in response to high environmental Zn. Importantly, virulence testing demonstrated that the conserved Cu and Zn detoxification-related Cys residues in AfAceA have key roles in pathogenesis. Therefore, these findings will broaden the current understanding of the adaption of saprophytic fungi to Cu and Zn stress and their survival in hosts and other environmental niches.

Precise Expression of Afmed15 Is Crucial for Asexual Development, Virulence, and Survival of Aspergillus fumigatus.

The rise of drug resistance in fungal pathogens is becoming a serious problem owing to the limited number of antifungal drugs available. Identifying and targeting factors essential for virulence or development unique to fungal pathogens is one approach to develop novel treatments for fungal infections. In this study, we present the identification and functional characterization of a novel developmental regulator in Aspergillus fumigatus, AfMed15, which contained a conserved Med15_fungal domain, as determined by screening of a mutant library that contained more than 2,000 hygromycin-resistant A. fumigatus transformants. Downregulating the expression of Afmed15 abolished the conidiation and decreased the fungal virulence in an insect model. Strikingly, the overexpression of Afmed15 caused fungal death accompanied by intensive autophagy. RNA sequencing of an Afmed15 overexpression strain revealed that altered gene expression patterns were associated with carbon metabolism, energy metabolism, and translation. Interestingly, the addition of metal ions could partially rescue fungal death caused by the overexpression of Afmed15, indicating that disordered ion homeostasis is a potential reason for the fungal death caused by the overexpression of Afmed15 Considering that the precise expression of Afmed15 is crucial for fungal development, virulence, and survival and that no ortholog was found in humans, Afmed15 is an ideal target for antifungal-drug development.IMPORTANCE The identification and characterization of regulators essential for virulence or development constitute one approach for antifungal drug development. In this study, we screened and functionally characterized Afmed15, a novel developmental regulator in A. fumigatus We demonstrate that the precise transcriptional expression of Afmed15 is crucial for fungal asexual development, virulence, and survival. Downregulating the expression of Afmed15 abolished the conidiation and decreased the fungal virulence in an insect model. In contrast, the overexpression of Afmed15 caused fungal death accompanied by intensive autophagy. Our study provides a foundation for further studies to identify compounds perturbing the expression of Afmed15 that may be used for the prevention of invasive A. fumigatus infections.

The Scaffold Proteins Paxillin B and α-Actinin Regulate Septation in Aspergillus nidulans via Control of Actin Ring Contraction.

Cytokinesis, as the final step of cell division, plays an important role in fungal growth and proliferation. In the filamentous fungus Aspergillus nidulans, defective cytokinesis is able to induce abnormal multinuclear or nonnucleated cells and then result in reduced hyphal growth and abolished sporulation. Previous studies have reported that a conserved contractile actin ring (CAR) protein complex and the septation initiation network (SIN) signaling kinase cascade are required for cytokinesis and septation; however, little is known about the role(s) of scaffold proteins involved in these two important cellular processes. In this study, we show that a septum-localized scaffold protein paxillin B (PaxB) is essential for cytokinesis/septation in A. nidulans The septation defects observed in a paxB deletion strain resemble those caused by the absence of another identified scaffold protein, α-actinin (AcnA). Deletion of α-actinin (AcnA) leads to undetectable PaxB at the septation site, whereas deletion of paxB does not affect the localization of α-actinin at septa. However, deletion of either α-actinin (acnA) or paxB causes the actin ring to disappear at septation sites during cytokinesis. Notably, overexpression of α-actinin acnA partially rescues the septum defects of the paxB mutant but not vice versa, suggesting AcnA may play a dominant role over that of PaxB for cytokinesis and septation. In addition, PaxB and α-actinin affect the septal dynamic localization of MobA, a conserved component of the SIN pathway, suggesting they may affect the SIN protein complex function at septa. Protein pull-down assays combined with liquid chromatography-mass spectrometry identification indicate that α-actinin AcnA and PaxB likely do not directly interact, but presumably belong to an actin cytoskeleton protein network that is required for the assembly and contraction of the CAR. Taken together, findings in this study provide novel insights into the roles of conserved scaffold proteins during fungal septation in A. nidulans.