β-1,3-Glucan recognition by
Acanthamoeba castellanii
as a putative mechanism of amoeba-fungal interactions
Authors: Ferreira Marina da Silva , Gonçalves Diego de Souza , Mendoza Susana Ruiz , de Oliveira Gabriel Afonso , Pontes Bruno , la Noval Claudia Rodríguez-de , Honorato Leandro , Ramos Luis Felipe Costa , Nogueira Fábio C. S. , Domont Gilberto B. , Casadevall Arturo , Nimrichter Leonardo , Peralta Jose Mauro , Guimaraes Allan J. 
Published: January 2024

ABSTRACT

In this study, we conducted an in-depth analysis to characterize potential
Acanthamoeba castellanii
(
Ac
) proteins capable of recognizing fungal β-1,3-glucans.
Ac
specifically anchors curdlan or laminarin, indicating the presence of surface β-1,3-glucan-binding molecules. Using optical tweezers, strong adhesion of laminarin- or curdlan-coated beads to
Ac
was observed, highlighting their adhesive properties compared to controls (characteristic time τ of 46.9 and 43.9 s, respectively). Furthermore,
Histoplasma capsulatum
(
Hc
) G217B, possessing a β-1,3-glucan outer layer, showed significant adhesion to
Ac
compared to a
Hc
G186 strain with an α-1,3-glucan outer layer (τ of 5.3 s vs τ 83.6 s). The addition of soluble β-1,3-glucan substantially inhibited this adhesion, indicating the involvement of β-1,3-glucan recognition. Biotinylated β-1,3-glucan-binding proteins from
Ac
exhibited higher binding to
Hc
G217B, suggesting distinct recognition mechanisms for laminarin and curdlan, akin to macrophages. These observations hinted at the β-1,3-glucan recognition pathway’s role in fungal entrance and survival within phagocytes, supported by decreased fungal viability upon laminarin or curdlan addition in both phagocytes. Proteomic analysis identified several
Ac
proteins capable of binding β-1,3-glucans, including those with lectin/glucanase superfamily domains, carbohydrate-binding domains, and glycosyl transferase and glycosyl hydrolase domains. Notably, some identified proteins were overexpressed upon curdlan/laminarin challenge and also demonstrated high affinity to β-1,3-glucans. These findings underscore the complexity of binding via β-1,3-glucan and suggest the existence of alternative fungal recognition pathways in
Ac
.

IMPORTANCE

Acanthamoeba castellanii
(
Ac
) and macrophages both exhibit the remarkable ability to phagocytose various extracellular microorganisms in their respective environments. While substantial knowledge exists on this phenomenon for macrophages, the understanding of
Ac
’s phagocytic mechanisms remains elusive. Recently, our group identified mannose-binding receptors on the surface of
Ac
that exhibit the capacity to bind/recognize fungi. However, the process was not entirely inhibited by soluble mannose, suggesting the possibility of other interactions. Herein, we describe the mechanism of β-1,3-glucan binding by
A. castellanii
and its role in fungal phagocytosis and survival within trophozoites, also using macrophages as a model for comparison, as they possess a well-established mechanism involving the Dectin-1 receptor for β-1,3-glucan recognition. These shed light on a potential parallel evolution of pathways involved in the recognition of fungal surface polysaccharides.