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21 Publications visible to you, out of a total of 21
Human Anti-fungal Th17 Immunity and Pathology Rely on Cross-Reactivity against Candida albicans.

Abstract (Expand)

Th17 cells provide protection at barrier tissues but may also contribute to immune pathology. The relevance and induction mechanisms of pathologic Th17 responses in humans are poorly understood. Here, … we identify the mucocutaneous pathobiont Candida albicans as the major direct inducer of human anti-fungal Th17 cells. Th17 cells directed against other fungi are induced by cross-reactivity to C. albicans. Intestinal inflammation expands total C. albicans and cross-reactive Th17 cells. Strikingly, Th17 cells cross-reactive to the airborne fungus Aspergillus fumigatus are selectively activated and expanded in patients with airway inflammation, especially during acute allergic bronchopulmonary aspergillosis. This indicates a direct link between protective intestinal Th17 responses against C. albicans and lung inflammation caused by airborne fungi. We identify heterologous immunity to a single, ubiquitous member of the microbiota as a central mechanism for systemic induction of human anti-fungal Th17 responses and as a potential risk factor for pulmonary inflammatory diseases.

Authors: P. Bacher, T. Hohnstein, E. Beerbaum, M. Rocker, M. G. Blango, S. Kaufmann, J. Rohmel, P. Eschenhagen, C. Grehn, K. Seidel, V. Rickerts, L. Lozza, U. Stervbo, M. Nienen, N. Babel, J. Milleck, M. Assenmacher, O. A. Cornely, M. Ziegler, H. Wisplinghoff, G. Heine, M. Worm, B. Siegmund, J. Maul, P. Creutz, C. Tabeling, C. Ruwwe-Glosenkamp, L. E. Sander, C. Knosalla, S. Brunke, B. Hube, O. Kniemeyer, A. A. Brakhage, C. Schwarz, A. Scheffold

Date Published: 7th Mar 2019

Publication Type: Not specified

Identification of Proteomic Markers in Head and Neck Cancer Using MALDI-MS Imaging, LC-MS/MS, and Immunohistochemistry.

Abstract (Expand)

PURPOSE: The heterogeneity of squamous cell carcinoma tissue greatly complicates diagnosis and individualized therapy. Therefore, characterizing the heterogeneity of tissue spatially and identifying … appropriate biomarkers is crucial. MALDI-MS imaging (MSI) is capable of analyzing spatially resolved tissue biopsies on a molecular level. EXPERIMENTAL DESIGN: MALDI-MSI is used on snap frozen and formalin-fixed and paraffin-embedded (FFPE) tissue samples from patients with head and neck cancer (HNC) to analyze m/z values localized in tumor and nontumor regions. Peptide identification is performed using LC-MS/MS and immunohistochemistry (IHC). RESULTS: In both FFPE and frozen tissue specimens, eight characteristic masses of the tumor's epithelial region are found. Using LC-MS/MS, the peaks are identified as vimentin, keratin type II, nucleolin, heat shock protein 90, prelamin-A/C, junction plakoglobin, and PGAM1. Lastly, vimentin, nucleolin, and PGAM1 are verified with IHC. CONCLUSIONS AND CLINICAL RELEVANCE: The combination of MALDI-MSI, LC-MS/MS, and subsequent IHC furnishes a tool suitable for characterizing the molecular heterogeneity of tissue. It is also suited for use in identifying new representative biomarkers to enable a more individualized therapy.

Authors: F. Hoffmann, C. Umbreit, T. Kruger, D. Pelzel, G. Ernst, O. Kniemeyer, O. Guntinas-Lichius, A. Berndt, F. von Eggeling

Date Published: 10th Nov 2018

Publication Type: Not specified

Module-detection approaches for the integration of multilevel omics data highlight the comprehensive response of Aspergillus fumigatus to caspofungin.

Abstract (Expand)

BACKGROUND: Omics data provide deep insights into overall biological processes of organisms. However, integration of data from different molecular levels such as transcriptomics and proteomics, still … remains challenging. Analyzing lists of differentially abundant molecules from diverse molecular levels often results in a small overlap mainly due to different regulatory mechanisms, temporal scales, and/or inherent properties of measurement methods. Module-detecting algorithms identifying sets of closely related proteins from protein-protein interaction networks (PPINs) are promising approaches for a better data integration. RESULTS: Here, we made use of transcriptome, proteome and secretome data from the human pathogenic fungus Aspergillus fumigatus challenged with the antifungal drug caspofungin. Caspofungin targets the fungal cell wall which leads to a compensatory stress response. We analyzed the omics data using two different approaches: First, we applied a simple, classical approach by comparing lists of differentially expressed genes (DEGs), differentially synthesized proteins (DSyPs) and differentially secreted proteins (DSePs); second, we used a recently published module-detecting approach, ModuleDiscoverer, to identify regulatory modules from PPINs in conjunction with the experimental data. Our results demonstrate that regulatory modules show a notably higher overlap between the different molecular levels and time points than the classical approach. The additional structural information provided by regulatory modules allows for topological analyses. As a result, we detected a significant association of omics data with distinct biological processes such as regulation of kinase activity, transport mechanisms or amino acid metabolism. We also found a previously unreported increased production of the secondary metabolite fumagillin by A. fumigatus upon exposure to caspofungin. Furthermore, a topology-based analysis of potential key factors contributing to drug-caused side effects identified the highly conserved protein polyubiquitin as a central regulator. Interestingly, polyubiquitin UbiD neither belonged to the groups of DEGs, DSyPs nor DSePs but most likely strongly influenced their levels. CONCLUSION: Module-detecting approaches support the effective integration of multilevel omics data and provide a deep insight into complex biological relationships connecting these levels. They facilitate the identification of potential key players in the organism's stress response which cannot be detected by commonly used approaches comparing lists of differentially abundant molecules.

Authors: T. Conrad, O. Kniemeyer, S. G. Henkel, T. Kruger, D. J. Mattern, V. Valiante, R. Guthke, I. D. Jacobsen, A. A. Brakhage, S. Vlaic, J. Linde

Date Published: 20th Oct 2018

Publication Type: Not specified

Proteome Analysis Reveals the Conidial Surface Protein CcpA Essential for Virulence of the Pathogenic Fungus Aspergillus fumigatus.

Abstract (Expand)

Aspergillus fumigatus is a common airborne fungal pathogen of humans and a significant source of mortality in immunocompromised individuals. Here, we provide the most extensive cell wall proteome … profiling to date of A. fumigatus resting conidia, the fungal morphotype pertinent to first contact with the host. Using liquid chromatography-tandem mass spectrometry (LC-MS/MS), we identified proteins within the conidial cell wall by hydrogen-fluoride (HF)-pyridine extraction and proteins exposed on the surface using a trypsin-shaving approach. One protein, designated conidial cell wall protein A (CcpA), was identified by both methods and was found to be nearly as abundant as hydrophobic rodlet layer-forming protein RodA. CcpA, an amphiphilic protein, like RodA, peaks in expression during sporulation on resting conidia. Despite high cell wall abundance, the cell surface structure of DeltaccpA resting conidia appeared normal. However, trypsin shaving of DeltaccpA conidia revealed novel surface-exposed proteins not detected on conidia of the wild-type strain. Interestingly, the presence of swollen DeltaccpA conidia led to higher activation of neutrophils and dendritic cells than was seen with wild-type conidia and caused significantly less damage to epithelial cells in vitro In addition, virulence was highly attenuated when cortisone-treated, immunosuppressed mice were infected with DeltaccpA conidia. CcpA-specific memory T cell responses were detectable in healthy human donors naturally exposed to A. fumigatus conidia, suggesting a role for CcpA as a structural protein impacting conidial immunogenicity rather than possessing a protein-intrinsic immunosuppressive effect. Together, these data suggest that CcpA serves as a conidial stealth protein by altering the conidial surface structure to minimize innate immune recognition.IMPORTANCE The mammalian immune system relies on recognition of pathogen surface antigens for targeting and clearance. In the absence of immune evasion strategies, pathogen clearance is rapid. In the case of Aspergillus fumigatus, the successful fungus must avoid phagocytosis in the lung to establish invasive infection. In healthy individuals, fungal spores are cleared by immune cells; however, in immunocompromised patients, clearance mechanisms are impaired. Here, using proteome analyses, we identified CcpA as an important fungal spore protein involved in pathogenesis. A. fumigatus lacking CcpA was more susceptible to immune recognition and prompt eradication and, consequently, exhibited drastically attenuated virulence. In infection studies, CcpA was required for virulence in infected immunocompromised mice, suggesting that it could be used as a possible immunotherapeutic or diagnostic target in the future. In summary, our report adds a protein to the list of those known to be critical to the complex fungal spore surface environment and, more importantly, identifies a protein important for conidial immunogenicity during infection.

Authors: V. Voltersen, M. G. Blango, S. Herrmann, F. Schmidt, T. Heinekamp, M. Strassburger, T. Kruger, P. Bacher, J. Lother, E. Weiss, K. Hunniger, H. Liu, P. Hortschansky, A. Scheffold, J. Loffler, S. Krappmann, S. Nietzsche, O. Kurzai, H. Einsele, O. Kniemeyer, S. G. Filler, U. Reichard, A. A. Brakhage

Date Published: 2nd Oct 2018

Publication Type: Not specified

Smelling the difference: Transcriptome, proteome and volatilome changes after mating

Abstract (Expand)

Mushrooms, such as Schizophyllum commune, have a specific odor. Whether this is linked to mating, prerequisite for mushroom formation, or also found in monokaryotic, unmated strains, was investigated … with a comprehensive study on the transcriptome and proteome of this model organism. Mating interactions were investigated using a complete, cytosolic proteome map for unmated, monokaryotic, as well as for mated, dikaryotic mycelia. The regulations of the proteome were compared to transcriptional changes upon mating and to changes in smell by volatilome studies. We could show a good overlap between proteome and transcriptome data, but extensive posttranslational regulation was identified for more than 80% of transcripts. This suggests down-stream regulation upon interaction of mating partners and formation of the dikaryon that is competent to form fruiting bodies. The volatilome was shown to respond to mating by a broader spectrum of volatiles and increased emission of the mushroom smell molecules 3-octanone and 1-octen-3-ol, as well as ethanol and beta-bisabolol in the dikaryon. Putatively involved biosynthetic proteins like alcohol dehydrogenases, Ppo-like oxygenases, or sesquiterpene synthases showed correlating transcriptional regulation depending on either mono- or dikaryotic stages.

Authors: D. Freihorst, M. Brunsch, S. Wirth, K. Krause, , , M. Kunert, W. Boland, E. Kothe

Date Published: 7th Sep 2016

Publication Type: Not specified

Immunoproteomic Analysis of Antibody Responses to Extracellular Proteins of Candida albicans Revealing the Importance of Glycosylation for Antigen Recognition.

Abstract (Expand)

During infection, the human pathogenic fungus Candida albicans undergoes a yeast-to-hypha transition, secretes numerous proteins for invasion of host tissues, and modulates the host's immune response. … Little is known about the interplay of C. albicans secreted proteins and the host adaptive immune system. Here, we applied a combined 2D gel- and LC-MS/MS-based approach for the characterization of C. albicans extracellular proteins during the yeast-to-hypha transition, which led to a comprehensive C. albicans secretome map. The serological responses to C. albicans extracellular proteins were investigated by a 2D-immunoblotting approach combined with MS for protein identification. On the basis of the screening of sera from candidemia and three groups of noncandidemia patients, a core set of 19 immunodominant antibodies against secreted proteins of C. albicans was identified, seven of which represent potential diagnostic markers for candidemia (Xog1, Lip4, Asc1, Met6, Tsa1, Tpi1, and Prx1). Intriguingly, some secreted, strongly glycosylated protein antigens showed high cross-reactivity with sera from noncandidemia control groups. Enzymatic deglycosylation of proteins secreted from hyphae significantly impaired sera antibody recognition. Furthermore, deglycosylation of the recombinantly produced, secreted aspartyl protease Sap6 confirmed a significant contribution of glycan epitopes to the recognition of Sap6 by antibodies in patient's sera.

Authors: T. Luo, T. Kruger, U. Knupfer, L. Kasper, N. Wielsch, B. Hube, A. Kortgen, M. Bauer, E. J. Giamarellos-Bourboulis, G. Dimopoulos, A. A. Brakhage, O. Kniemeyer

Date Published: 5th Aug 2016

Publication Type: Not specified

The hypoxia-induced dehydrogenase HorA is required for coenzyme Q10 biosynthesis, azole sensitivity and virulence of Aspergillus fumigatus

Abstract (Expand)

Aspergillus fumigatus is the predominant airborne pathogenic fungus causing invasive aspergillosis in immunocompromised patients. During infection A. fumigatus has to adapt to oxygen-limiting conditions … in inflammatory or necrotic tissue. Previously, we identified a mitochondrial protein to be highly up-regulated during hypoxic adaptation. Here, this protein was found to represent the novel oxidoreductase HorA. In Saccharomyces cerevisiae a homologue was shown to play a role in biosynthesis of coenzyme Q. Consistently, reduced coenzyme Q content in the generated DeltahorA mutant indicated a respective function in A. fumigatus. Since coenzyme Q is involved in cellular respiration and maintaining cellular redox homeostasis, the strain DeltahorA displayed an impaired response to both oxidative and reductive stress, a delay in germination and an accumulation of NADH. Moreover, an increased resistance against antifungal drugs was observed. All phenotypes were completely reversed by the addition of the synthetic electron carrier menadione. The deletion strain DeltahorA showed significantly attenuated virulence in two murine infection models of invasive pulmonary aspergillosis. Therefore, the biosynthesis of coenzyme Q and, particularly, the fungal-specific protein HorA play a crucial role in virulence of A. fumigatus. Due to its absence in mammals, HorA might represent a novel therapeutic target against fungal infections. This article is protected by copyright. All rights reserved.

Authors: K. Kroll, E. Shekhova, D. J. Mattern, A. Thywissen, , M. Strassburger, T. Heinekamp, , ,

Date Published: 19th Mar 2016

Publication Type: Not specified

Proteomic Profiling of Serological Responses to Aspergillus fumigatus Antigens in Patients with Invasive Aspergillosis

Abstract (Expand)

Aspergillus fumigatus is the species that most commonly causes the opportunistic infection invasive aspergillosis (IA) in patients being treated for hematological malignancies. Little is known about … the A. fumigatus proteins that trigger the production of Aspergillus-specific IgG antibodies during the course of IA. To characterize the serological response to A. fumigatus protein antigens, mycelial proteins were separated by 2-D gel electrophoresis. The gels were immunoblotted with sera from patients with probable and proven IA and control patients without IA. We identified 49 different fungal proteins, which gave a positive IgG antibody signal. Most of these antigens play a role in primary metabolism and stress responses. Overall, our analysis identified 18 novel protein antigens from A. fumigatus. To determine whether these antigens can be used as diagnostic or prognostic markers or exhibit a protective activity, we employed supervised machine learning with decision trees. We identified two candidates for further analysis, the protein antigens CpcB and Shm2. Heterologously produced Shm2 induced a strongly proinflammatory response in human peripheral blood mononuclear cells after in vitro stimulation. In contrast, CpcB did not activate the immune response of PBMCs. These findings could serve as the basis for the development of an immunotherapy of IA.

Authors: J. Teutschbein, S. Simon, J. Lother, J. Springer, P. Hortschansky, C. O. Morton, , , E. Conneally, T. R. Rogers, , ,

Date Published: 15th Mar 2016

Publication Type: Not specified

Hitting the caspofungin salvage pathway of human-pathogenic fungi with the novel lasso peptide humidimycin (MDN-0010)

Abstract (Expand)

Fungal infections have increased dramatically in the last 2 decades, and fighting infectious diseases requires innovative approaches such as the combination of two drugs acting on different targets or … even targeting a salvage pathway of one of the drugs. The fungal cell wall biosynthesis is inhibited by the clinically used antifungal drug caspofungin. This antifungal activity has been found to be potentiated by humidimycin, a new natural product identified from the screening of a collection of 20,000 microbial extracts, which has no major effect when used alone. An analysis of transcriptomes and selected Aspergillus fumigatus mutants indicated that humidimycin affects the high osmolarity glycerol response pathway. By combining humidimycin and caspofungin, a strong increase in caspofungin efficacy was achieved, demonstrating that targeting different signaling pathways provides an excellent basis to develop novel anti-infective strategies.

Authors: , M. C. Monteiro, J. Martin, R. Altwasser, N. El Aouad, I. Gonzalez, , E. Mellado, S. Palomo, N. de Pedro, I. Perez-Victoria, J. R. Tormo, F. Vicente, F. Reyes, O. Genilloud,

Date Published: 8th Jun 2015

Publication Type: Not specified

The novel globin protein fungoglobin is involved in low oxygen adaptation of Aspergillus fumigatus

Abstract (Expand)

The human pathogenic fungus Aspergillus fumigatus normally lives as a soil saprophyte. Its environment includes poorly oxygenated substrates that also occur during tissue invasive growth of the fungus … in the human host. Up to now, few cellular factors have been identified that allow the fungus to efficiently adapt its energy metabolism to hypoxia. Here, we cultivated A. fumigatus in an O2 -controlled fermenter and analysed its responses to O2 limitation on a minute timescale. Transcriptome sequencing revealed several genes displaying a rapid and highly dynamic regulation. One of these genes was analysed in detail and found to encode fungoglobin, a previously uncharacterized member of the sensor globin protein family widely conserved in filamentous fungi. Besides low O2 , iron limitation also induced transcription, but regulation was not entirely dependent on the two major transcription factors involved in adaptation to iron starvation and hypoxia, HapX and SrbA respectively. The protein was identified as a functional haemoglobin, as binding of this cofactor was detected for the recombinant protein. Gene deletion in A. fumigatus confirmed that haem-binding fungoglobins are important for growth in microaerobic environments with O2 levels far lower than in hypoxic human tissue.

Authors: F. Hillmann, , N. Beckmann, M. Cyrulies, M. Strassburger, T. Heinekamp, H. Haas, , ,

Date Published: 7th Jul 2014

Publication Type: Not specified

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