Phosphatase activity on the cell wall ofFonsecaeapedrosoi

Abstract

The activity of a phosphatase was characterized in intact mycelial forms of Fonsecaea pedrosoi, a pathogenic fungus that causes chromoblastomycosis. At pH 5.5, this fungus hydrolyzed p-nitrophenylphosphate (p-NPP) to p-nitrophenol (p-NP) at a rate of 12.78 +/- 0.53 nmol p-NP per h per mg hyphal dry weight. The values of Vmax and apparent Km for p-NPP hydrolyses were measured as 17.89 +/- 0.92 nmol p-NP per h per mg hyphal dry weight and 1.57 +/- 0.26 mmol/l, respectively. This activity was inhibited at increased pH, a finding compatible with an acid phosphatase. The enzymatic activity was strongly inhibited by classical inhibitors of acid phosphatases such as sodium orthovanadate (Ki = 4.23 micromol/l), sodium molybdate (Ki = 7.53 micromol/l) and sodium fluoride (Ki = 126.78 micromol/l) in a dose-dependent manner. Levamizole (1 mmol/l) and sodium tartrate (10 mmol/l), had no effect on the enzyme activity. Cytochemical localization of the acid phosphatase showed electrondense cerium phosphate deposits on the cell wall, as visualized by transmission electron microscopy. Phosphatase activity in F. pedrosoi seems to be associated with parasitism, as sclerotic cells, which are the fungal forms mainly detected in chromoblastomycosis lesions, showed much higher activities than conidia and mycelia did. A strain of F. pedrosoi recently isolated from a human case of chromoblastomycosis also showed increased enzyme activity, suggesting that the expression of surface phosphatases may be stimulated by interaction with the host.

Ectophosphatase activity in conidial forms of Fonsecaea pedrosoi is modulated by exogenous phosphate and influences fungal adhesion to mammalian cells

“…The phosphatase activity was strongly inhibited by P i , and cultivation of F. pedrosoi conidia in the absence of exogenous P i resulted in the generation of fungal cells expressing an ectophosphatase activity 130-fold higher than that expressed by fungi cultivated in the presence of P i , which agreed with our previous biochemical observations ( Kneipp et al, 2003). The depletion of phosphate from the culture medium apparently induced the expression of a different ectophosphatase, as suggested by the differences in the affinity for the artificial substrate p-NPP and by the fact that Fe 3+ did not enhance the ectophosphatase activity from conidia cultivated in P i -depleted medium.…”

“…Acid phosphatases (EC 3.1.3.2) have been described in many yeasts (Vogel & Hinnen, 1990;VasilevaTonkova et al, 1996). These enzymes may exist as soluble or secreted forms (Jolivet et al, 1998), or remain attached to the outer surface of the inner membrane (Arnold et al, 1988) or cell wall (González et al, 1993;Bernard et al, 2002;Kneipp et al, 2003).…”

Structure, Cellular Distribution, Antigenicity, and Biological Functions of Fonsecaea pedrosoi Ceramide Monohexosides

“…We therefore investigated the influence of the monoclonal antibody to CMH on the adhesion and killing of sclerotic cells by the murine macrophage cell line J774. 16. For this purpose, we incubated sclerotic bodies, previously treated with irrelevant IgG or anti-CMH antibodies (10 g/ml), for 1 h with a monolayer of murine macrophages in ratios of 1 fungus per macrophage or 10 phagocytes per fungal cell.…”

“…Kneipp and coworkers described a higher phosphatase activity in sclerotic cells than in conidial and mycelial forms (16), which was associated with parasitism. However, it also suggests that these enzymes could be involved in F. pedrosoi dimorphism, although the details by which it happens remain to be elucidated.…”

Chromoblastomycosis

“…Proteolytic enzymes can be detrimental to various components of the host defense mechanisms, leading to immunological escape or antimicrobial resistance (204,205). Kneipp et al demonstrated that muriform cells, in comparison to conidia and mycelia, have high phosphatase activity associated with pathogenicity (206,207). Phosphorylated substrates were hydrolyzed by surface ectophosphatases.…”