Biotinylated surfome profiling identifies potential biomarkers for diagnosis and therapy ofAspergillus fumigatusinfection
Authors: Jia Lei-Jie , Krüger Thomas , Blango Matthew G. , Kniemeyer Olaf , Brakhage Axel A. 
Published: April 2020

Aspergillus fumigatusis one of the most common airborne fungi capable of causing invasive mycoses in immunocompromised patients and allergic diseases in susceptible individuals. In both cases, fungal surface proteins mediate the first contact with the human immune system to evade immune responses or to induce hypersensitivity. Several methods have been established to study the surface proteome (surfome) ofA. fumigatus, like trypsin shaving, glucanase treatment, or formic acid extraction. Biotinylation coupled with LC-MS/MS identification of peptides is a particularly efficient method to identify the surface exposed regions of proteins that potentially mediate interaction with the host. After biotinylation of surface proteins during spore germination, we detected 314 different surface proteins, including several well-known proteins like RodA, CcpA, and DppV, as well as several allergens, heat shock proteins (HSPs), and previously undescribed surface proteins. Using immunofluorescence microscopy, we confirmed the surface localization of three HSPs, which may have moonlighting functions. Collectively, our study generated a comprehensive data set of theA. fumigatussurfome, which complements already existingA. fumigatussurface proteome data and allows us to propose a common core set ofA. fumigatussurface proteins. In addition, our study uncovers the surface-exposed regions of many proteins on the surface of spores or hyphae. These surface exposed regions are candidates for direct interaction with host cells and may represent antigenic epitopes that either induce protective immune responses or mediate immune evasion. Thus, the comprehensive datasets provided and compiled here represent reasonable immunotherapy and diagnostic targets for future investigations.

HIGHLIGHTS

Surface protein biotinylation coupled with LC-MS/MS analysis provides a comprehensive dataset of theA. fumigatussurface proteome.

314 differentA. fumigatusproteins (including immunoreactive proteins, and virulence factors) with surface exposed regions were detected.

Surface localization of three Hsp70 chaperones was confirmed by protein tagging coupled with immunofluorescence.

By comparison with other surfome datasets, a core surfome ofA. fumigatuswas defined, which provides possible biomarkers for diagnosis or therapy.

SIGNIFICANCE

Aspergillus fumigatusis the most important airborne human pathogenic mold, capable of causing both life-threatening invasive pulmonary aspergillosis in immunocompromised patients and allergic infections in atopic individuals. Despite its obvious medical relevance, timely diagnosis and efficient antifungal treatment ofA. fumigatusinfection remains a major challenge. Proteins on the surface of conidia (asexually produced spores) and mycelium directly mediate host-pathogen interaction and also may serve as targets for diagnosis and immunotherapy. However, the similarity of protein sequences betweenA. fumigatusand other organisms, and sometimes even the human host, makes selection of targets for immunological-based studies difficult. Here, using surface protein biotinylation coupled with LC-MS/MS analysis, we identified hundreds ofA. fumigatussurface proteins with exposed regions, further defining putative targets for possible diagnostic and immunotherapeutic design.