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Paracoccidoides brasiliensis 30 kDa adhesin: identification as a 14-3-3 protein, cloning and subcellular localization
Julhiany de Fatima da Silva1, Haroldo César de Oliveira, Caroline Maria Marcos
1Department of Clinical Analyses, Faculty of Pharmaceutical Sciences, São Paulo State University - University Estadual Paulista Araraquara, São Paulo, Brazil.
Abstract:
Paracoccidoides brasiliensis adhesion to lung epithelial cells is considered an essential event for the establishment of infection and different proteins participate in this process. One of these proteins is a 30 kDa adhesin, pI 4.9 that was described as a laminin ligand in previous studies, and it was more highly expressed in more virulent P. brasiliensis isolates. This protein may contribute to the virulence of this important fungal pathogen. Using Edman degradation and mass spectrometry analysis, this 30 kDa adhesin was identified as a 14-3-3 protein. These proteins are a conserved group of small acidic proteins involved in a variety of processes in eukaryotic organisms. However, the exact function of these proteins in some processes remains unknown. Thus, the goal of the present study was to characterize the role of this protein during the interaction between the fungus and its host. To achieve this goal, we cloned, expressed the 14-3-3 protein in a heterologous system and determined its subcellular localization in in vitro and in vivo infection models. Immunocytochemical analysis revealed the ubiquitous distribution of this protein in the yeast form of P. brasiliensis, with some concentration in the cytoplasm. Additionally, this 14-3-3 protein was also present in P. brasiliensis cells at the sites of infection in C57BL/6 mice intratracheally infected with P. brasiliensis yeast cells for 72 h (acute infections) and 30 days (chronic infection). An apparent increase in the levels of the 14-3-3 protein in the cell wall of the fungus was also noted during the interaction between P. brasiliensis and A549 cells, suggesting that this protein may be involved in host-parasite interactions, since inhibition assays with the protein and this antibody decreased P. brasiliensis adhesion to A549 epithelial cells. Our data may lead to a better understanding of P. brasiliensis interactions with host tissues and paracoccidioidomycosis pathogenesis.
Insights
A 14-3-3 protein from Paracoccidoides brasiliensis aids fungal adhesion to lung cells. Inhibiting this protein reduces fungal attachment, offering insights into paracoccidioidomycosis pathogenesis.
Area of Science:
- Mycology
- Infectious Diseases
- Molecular Biology
Background:
- Paracoccidoides brasiliensis adhesion to lung epithelial cells is crucial for infection establishment.
- A 30 kDa adhesin, identified as a 14-3-3 protein, is upregulated in virulent isolates and may enhance fungal virulence.
Purpose of the Study:
- To characterize the role of the P. brasiliensis 14-3-3 protein in host-fungus interactions.
- To investigate the protein's function during the pathogenic process of paracoccidioidomycosis.
Main Methods:
- Cloning and heterologous expression of the P. brasiliensis 14-3-3 protein.
- Subcellular localization studies using immunocytochemistry in vitro and in vivo infection models.
- Inhibition assays to assess the effect on fungal adhesion to epithelial cells.
Main Results:
- The 14-3-3 protein is ubiquitously distributed in P. brasiliensis yeast cells, concentrated in the cytoplasm.
- The protein is present in fungal cells at infection sites in a mouse model (acute and chronic).
- Increased 14-3-3 protein levels in the fungal cell wall were observed during interaction with A549 cells; antibody inhibition reduced fungal adhesion.
Conclusions:
- The P. brasiliensis 14-3-3 protein is involved in fungal adhesion to host epithelial cells.
- This protein likely plays a significant role in the pathogenesis of paracoccidioidomycosis.
- Targeting this adhesin could be a strategy to combat P. brasiliensis infections.
