The Aspergillus fumigatus Phosphoproteome Reveals Roles of High-Osmolarity Glycerol Mitogen-Activated Protein Kinases in Promoting Cell Wall Damage and Caspofungin Tolerance

Author(s):Show less -

Mattos, Eliciane Cevolani ^[1] ; Silva, Lilian Pereira ^[1] ; Valero, Clara ^[1] ; de Castro, Patricia Alves ^[1] ; dos Reis, Thaila Fernanda ^[1] ; Ribeiro, Liliane F. C. ^[2] ; Marten, Mark R. ^[2] ; Silva-Rocha, Rafael ^[3] ; Westmann, Caua ^[3] ; Tomich de Paula da Silva, Carlos Henrique ^{[1, 4]} ; Taft, Carlton Anthony ^[5] ; Al-Furaiji, Narjes ^[6] ; Bromley, Michael ^[6] ; Mortensen, Uffe H. ^[7] ; Benz, J. Philipp ^{[8, 9]} ; Brown, Neil Andrew ^[10] ; Goldmana, Gustavo H. ^{[1, 8]}

Affiliation

^[1] Univ Sao Paulo, Fac Ciencias Farmaceut Ribeirao Preto, Ribeirao Preto - Brazil

^[2] Univ Maryland Baltimore Cty, Dept Chem Biochem & Environm Engn, Baltimore, MD 21228 - USA

^[3] Univ Sao Paulo, Fac Med Ribeirao Preto, Ribeirao Preto - Brazil

^[4] Univ Sao Paulo, Dept Quim, FFCLRP, Ribeirao Preto - Brazil

^[5] CBPF, Rio De Janeiro - Brazil

^[6] Univ Manchester, Fac Biol Med & Hlth, Sch Biol Sci, Div Infect Immun & Resp Med, Manchester, Lancs - England

^[7] Tech Univ Denmark, Dept Biotechnol & Biomed, Sect Synthet Biol, Eukaryot Mol Cell Biol, Lyngby - Denmark

^[8] Tech Univ Munich, Inst Adv Study, Garching - Germany

^[9] Tech Univ Munich, Holzforsch Munchen, TUM Sch Life Sci Weihenstephan, Freising Weihenstephan - Germany

^[10] Univ Bath, Dept Biol & Biochem, Bath, Avon - England

• Document type: Journal article
• Source: MBIO; v. 11, n. 1 JAN-FEB 2020.
• Web of Science Citations: 0

Abstract

The filamentous fungus Aspergillus fumigatus can cause a distinct set of clinical disorders in humans. Invasive aspergillosis (IA) is the most common life-threatening fungal disease of immunocompromised humans. The mitogen-activated protein kinase (MAPK) signaling pathways are essential to the adaptation to the human host. Fungal cell survival is highly dependent on the organization, composition, and function of the cell wall. Here, an evaluation of the global A. fumigatus phosphoproteome under cell wall stress caused by the cell wall-damaging agent Congo red (CR) revealed 485 proteins potentially involved in the cell wall damage response. Comparative phosphoproteome analyses with the Delta sakA, Delta mpkC, and Delta sakA Delta mpkC mutant strains from the osmotic stress MAPK cascades identify their additional roles during the cell wall stress response. Our phosphoproteomics allowed the identification of novel kinases and transcription factors (TFs) involved in osmotic stress and in the cell wall integrity (CWI) pathway. Our global phosphoproteome network analysis showed an enrichment for protein kinases, RNA recognition motif domains, and the MAPK signaling pathway. In contrast to the wild-type strain, there is an overall decrease of differentially phosphorylated kinases and phosphatases in Delta sakA, Delta mpkC, and Delta sakA Delta mpkC mutants. We constructed phosphomutants for the phosphorylation sites of several proteins differentially phosphorylated in the wild-type and mutant strains. For all the phosphomutants, there is an increase in the sensitivity to cell wall-damaging agents and a reduction in the MpkA phosphorylation upon CR stress, suggesting these phosphosites could be important for the MpkA modulation and CWI pathway regulation. IMPORTANCE Aspergillus fumigatus is an opportunistic human pathogen causing allergic reactions or systemic infections, such as invasive pulmonary aspergillosis in immunocompromised patients. The mitogen-activated protein kinase (MAPK) signal-ing pathways are essential for fungal adaptation to the human host. Fungal cell survival, fungicide tolerance, and virulence are highly dependent on the organization, composition, and function of the cell wall. Upon cell wall stress, MAPKs phosphorylate multiple target proteins involved in the remodeling of the cell wall. Here, we investigate the global phosphoproteome of the Delta sakA and Delta mpkC A. fumigatus and high-osmolarity glycerol (HOG) pathway MAPK mutants upon cell wall damage. This showed the involvement of the HOG pathway and identified novel protein kinases and transcription factors, which were confirmed by fungal genetics to be involved in promoting tolerance of cell wall damage. Our results provide understanding of how fungal signal transduction networks modulate the cell wall. This may also lead to the discovery of new fungicide drug targets to impact fungal cell wall function, fungicide tolerance, and virulence. (AU)

• FAPESP's process: 18/18043-1 - Characterization of phospho-mimetic mutants of kinases in Aspergillus fumigatus by CRISPR-cas 9 gene edition
• Grantee: Eliciane Cevolani Mattos
• Support Opportunities: Scholarships abroad - Research Internship - Post-doctor
• FAPESP's process: 16/07870-9 - The influence of mitogen activated protein kinases (MAPK) on the expression of genetic determinants important for Aspergillus fumigatus virulence
• Grantee: Gustavo Henrique Goldman
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 17/19288-5 - Characterization of MAPKs dependent-phosphoproteomes of Aspergillus fumigatus under stress conditions
• Grantee: Eliciane Cevolani Mattos
• Support Opportunities: Scholarships in Brazil - Post-Doctoral