The bZIP Transcription Factor HapX Is Post-Translationally Regulated to Control Iron Homeostasis in Aspergillus fumigatus

Abstract

The airborne fungus Aspergillus fumigatus causes opportunistic infections in humans with high mortality rates in immunocompromised patients. Previous work established that the bZIP transcription factor HapX is essential for virulence via adaptation to iron limitation by repressing iron-consuming pathways and activating iron acquisition mechanisms. Moreover, HapX was shown to be essential for transcriptional activation of vacuolar iron storage and iron-dependent pathways in response to iron availability. Here, we demonstrate that HapX has a very short half-life during iron starvation, which is further decreased in response to iron, while siderophore biosynthetic enzymes are very stable. We identified Fbx22 and SumO as HapX interactors and, in agreement, HapX post-translational modifications including ubiquitination of lysine161, sumoylation of lysine242 and phosphorylation of threonine319. All three modifications were enriched in the immediate adaptation from iron-limiting to iron-replete conditions. Interfering with these post-translational modifications, either by point mutations or by inactivation, of Fbx22 or SumO, altered HapX degradation, heme biosynthesis and iron resistance to different extents. Consistent with the need to precisely regulate HapX protein levels, overexpression of hapX caused significant growth defects under iron sufficiency. Taken together, our results indicate that post-translational regulation of HapX is important to control iron homeostasis in A. fumigatus.

Copper acquisition is essential for plant colonization and virulence in a root-infecting vascular wilt fungus

“…Furthermore, the inability of the Fo mac1 Δ mutant to grow in -Cu conditions confirms that the function of Mac1 is highly conserved across different fungi, which is in line with a previous study showing that the mac1 gene from A. fumigatus can functionally complement a mac1 Δ mutant of S. cerevisiae (Park et al, 2017). By contrast, we found that the post-translational regulation of Mac1 activity in Fo differs from that reported in other fungi where inactivation under copper sufficiency occurs mainly by protein degradation and/or cytoplasmic retention, analogous to what has been reported for the iron-responsive transcription factor HapX(Lopez-Berges et al, 2021). In S. cerevisiae , Mac1 is stable at low copper concentrations but rapidly degraded at concentrations above 10 µM CuSO 4(Zhu et al., 1998) .…”

Modular Inducible Multigene Expression System for Filamentous Fungi

“…Initially, we analyzed each promoter separately, to better establish the optimal conditions for their induction or repression. PxylP has been widely used in A. fumigatus ( 38 , 41 – 44 ), and the general conditions to obtain a high level of expression were set to 1% glucose and 1% xylose in minimal medium, while glucose alone would be suitable to efficiently downregulate PxylP -driven gene expression. We conducted a titration analysis using 1% glucose and xylose in a concentration range from 0% to 3%, and we confirmed, as previously described ( 38 ), that a concentration as low as 0.09% xylose was sufficient for induction ( Fig.…”

The Siderophore Ferricrocin Mediates Iron Acquisition in Aspergillus fumigatus

“…As the latter are known to be iron regulated at the TL level ( 6 , 26 , 27 ), the control of TLs of the RIA and SIA genes during germination appears to be mediated by developmental regulators rather than by the iron-sensing transcription factors. However, HapX was recently shown to be subject to posttranslational modification ( 34 ), and therefore, it cannot be excluded that SIA and RIA are influenced by posttranslational modification of HapX and/or SreA. Moreover, the significantly higher TLs of genes involved in biosynthesis and uptake of FC in conidia and early germination ( sit1 , sidC , sidL ) than genes involved in biosynthesis and uptake of TAFC ( mirB , sidG ) indicated a role of biosynthesis and uptake of FC.…”