α‐(1,4)‐Amylase, but not α‐ and β‐(1,3)‐glucanases, may be responsible for the impaired growth and morphogenesis of Paracoccidioides brasiliensis induced by N‐glycosylation inhibition

Abstract

The cell wall of Paracoccidioides brasiliensis, which consists of a network of polysaccharides and glycoproteins, is essential for fungal pathogenesis. We have previously reported that N-glycosylation of proteins such as N-acetyl-β-d-glucosaminidase is required for the growth and morphogenesis of P. brasiliensis. In the present study, we investigated the influence of tunycamicin (TM)-mediated inhibition of N-linked glycosylation on α- and β-(1,3)-glucanases and on α-(1,4)-amylase in P. brasiliensis yeast and mycelium cells. The addition of 15 µg/ml TM to the fungal cultures did not interfere with either α- or β-(1,3)-glucanase production and secretion. Moreover, incubation with TM did not alter α- and β-(1,3)-glucanase activity in yeast and mycelium cell extracts. In contrast, α-(1,4)-amylase activity was significantly reduced in underglycosylated yeast and mycelium extracts after exposure to TM. In spite of its importance for fungal growth and morphogenesis, N-glycosylation was not required for glucanase activities. This is surprising because these activities are directed to wall components that are crucial for fungal morphogenesis. On the other hand, N-glycans were essential for α-(1,4)-amylase activity involved in the production of malto-oligosaccharides that act as primer molecules for the biosynthesis of α-(1,3)-glucan. Our results suggest that reduced fungal α-(1,4)-amylase activity affects cell wall composition and may account for the impaired growth of underglycosylated yeast and mycelium cells. © 2013 The Authors. Yeast published by John Wiley & Sons Ltd.

Paracoccin distribution supports its role in Paracoccidioides brasiliensis growth and dimorphic transformation

“…brasiliensis following temperature shift. The features we observed were consistent with both original reports [ 13 , 28 , 34 ] and more recent descriptions [ 35 – 38 ] of the fungal morphotypes. As expected, hyphae showed a typical filamentous branching structure with growth tips [ 39 ].…”

Extracellular vesicles from Paracoccidioides pathogenic species transport polysaccharide and expose ligands for DC-SIGN receptors

“…In both species, the Amy1p seems to mediate hydrolysis of α-(1→4)-glucan polysaccharides. P. brasiliensis Amy1p produces malto-oligosaccharides that could be involved in the initiation of α-(1→3)-glucan synthesis or in the process of transglycosylation 40 41 . Despite the fact that Amy1p has not been detected in the secretome of yeast-phase P. brasiliensis 21 or H. capsulatum 42 , enzymes responsible for carbohydrate metabolism, such as cell wall α-1,3-glucan synthase mok13 (PADG_03169), endo-β-1,3-glucanase (PADG_07351), glucan 1–3-β-glucosidase (PADG_07615), and α-mannosidase (PADG_04148) are transported by EVs 21 , suggesting that EV polysaccharides and carbohydrate metabolism enzymes could have role in glucan synthesis and remodeling of the cell wall, as well as in lipid and protein glycosylation.…”

Virulence-Associated Enzymes of Cryptococcus neoformans

“…As nature's "recyclers," environmental fungi secrete a number of degradative enzymes to breakdown macromolecules and obtain nutrients in the environment (7,(66)(67)(68)(69). C. neoformans is no exception and releases a number of lipases, proteases, and DNases.…”