Ribosomal biogenesis factor links ribosome homeostasis to host relevant stress survival in Aspergillus fumigatus

Abstract

The inhalation of Aspergillus fumigatus spores into the human pulmonary tract places conidia in direct contact with host immune cells which deploy a coordinated killing mechanism aimed at conidia clearance and preventing invasive infection. The fungus also employs cellular processes that aid its survival during these host mediated stresses, which allows the conidia to sustain growth while resisting attacks by the host immune cells. Ribosomal biogenesis plays a vital role in growth, survival, and stress adaptation in eukaryotic cells. In filamentous fungal pathogens, this process supports the transition of inhaled spores into invasive hyphae and is therefore an important aspect of fungal pathogenicity. It is, however, unclear whether ribosomal biogenesis factors contribute to stress survival beyond their canonical ribosomal roles and provision of support for vegetative growth. In this study, we investigated the role of A. fumigatus ribosomal biogenesis factor BFR2 in contribution to fungal stress survival and resistance to neutrophil killing. In agreement with its ribosomal biogenesis role, the repression of BFR2 impairs 18S ribosomal rRNA formation, and this impairment directly extends to the inhibition of germination and vegetative growth of fungal colonies. We optimized an induction/shut-off system that allows the transient expression of BFR2 in swollen conidia followed by subsequent depletion of BFR2 to investigate its role in stress response outside of its ribosomal biogenesis role. This transient induction followed by shut-off did not result in detectable reduction in ribosomal RNA abundance, global translation capacity, and biomass accumulation therefore indicating that pre-induction of BFR2 supports normal growth physiology in A. fumigatus over the duration tested. Nevertheless, the depletion of BFR2 following a transient induction increased A. fumigatus susceptibility to oxidative, replication and DNA-damaging stress while preserving tolerance to endoplasmic reticulum (ER) stress. The depletion of BFR2 under these conditions also increased in vitro neutrophil mediated killing without altering conidia uptake. In a mouse model of invasive aspergillosis, BFR2 depletion reduced conidial survival within neutrophils and pulmonary fungal burden, and in an immunocompromised mice model, the depletion of BFR2 attenuated virulence thereby extending survival by a few days. These findings identify BFR2 as a crucial link between ribosomal biogenesis and fungal stress survival during host relevant stresses. Thus, targeting ribosomal biogenesis factors that underpin stress survival represents a promising strategy for antifungal development that could target both ribosomal biogenesis function and also stress response functions.