Titel: Redox proteomic analysis reveals oxidative modifications of proteins by increased levels of intracellular reactive oxygen species during hypoxia adaptation of the human-pathogenic mould Aspergillus fumigatus
Art: Abstractautor
Session: Workshop 02
Eukaryotic Pathogens incl. DMykG Lecture (FG EK)

Referent: Olaf Kniemeyer (Jena)

Abstract - Text

Question: The filamentous fungus Aspergillus fumigatus is a ubiquitously distributed, opportunistic pathogen, which causes diseases ranging from allergic responses, to chronic, but also life-threatening, invasive infections. Patients at risk of developing an invasive aspergillosis comprise those with blood malignancies, allogenic bone marrow as well as solid organ transplants, and chronic lung diseases. At the site of infection, A. fumigatus faces abrupt declines in oxygen concentrations. An increasing number of studies have demonstrated that elevated levels of reactive oxygen species (ROS) are formed under hypoxic growth conditions. Hypoxic ROS is able to introduce protein thiol modifications that may either impair protein functions or modulate hypoxia adaptation by activating hypoxic genes. Here, we aimed to study ROS formation in A. fumigatus under low oxygen levels and to identify proteins that undergo redox-dependent post-translational modifications under these conditions.

Methods: We characterized the proteomic response of A. fumigatus to a short period of hypoxia by LC-MS/MS-based detection of relative changes in protein abundances and redox-dependent cysteine modifications. The study was supplemented by further characterisation of a redox-sensitive protein using molecular biology techniques.

Results: The oxidant-sensing probe dichlorodihydrofluorescein diacetate revealed increased amounts of intracellular ROS in A. fumigatus exposed to hypoxia. Moreover, nuclear accumulation of the oxidative stress response regulator AfYap1 was observed under hypoxic growth conditions. The highest number (n=18) of proteins with significant increased levels of oxidatively modified thiols groups was detected after one hour of hypoxic cultivations. These included proteins with important roles in maintaining redox balance and protein folding, such as the thioredoxin Asp f 29 and the protein disulphide-isomerase PdiA. Our data also showed that the mitochondrial respiratory complex IV assembly protein Coa6 was significantly oxidised by hypoxic ROS. Deletion of the corresponding gene resulted in a complete absence of hypoxic growth in A. fumigatus.

Conclusions: Our redox proteomics study defined important cellular pathways that are targets of hypoxia-induced ROS. We further showed that Coa6 is an essential factor of respiration under hypoxia in A. fumigatus.