Chitin synthesis in Fusarium graminearum and its inhibition by edifenphos (Hinosan)

Abstract

Membrane-bound chitin synthase from FuscUium graminearum A315 had a pH optimum of 6.5, an apparent temperature optimum of 30 "C and an apparent K,,, for UDP-N-acetylglucosamine (UDP-GlcNAc) of 2.5 mM.Digitonin-solubilized chitin synthase had a lower (about 25% of the original activity) and more variable activity than the membrane-bound chitin synthase from which it was prepared. The activity of solubilized chitin synthase was stabilized by incorporating Allosamidin (a chitinase inhibitor) into the reaction mixture, and an approximately 3.6-fold increase in activity was observed when the reaction mixture contained dimyristoylphosphatidylcholine. Concentrations of the organophosphorus fungicide edifenphos (Hinosan) up to 25 p~ had no effect on the in vitro activity of membrane-bound chitin synthase, but between 25 and 145 p~, chitin synthase activity decreased with increase in fungicide concentration. Edifenphos caused non-competitive inhibition of chitin synthase activity with an apparent Ki of 50 PM. Membrane-bound chitin synthase preparations from cultures of F. graminearurn grown in 250 pM-edifenphos had a much lower in vitro activity than preparations from cultures grown in the absence of the fungicide. Pre-growth of F. grdnearum in 250 p~ (but not 80 p~) edifenphos also caused a reduction in incorporation of [3H]GlcNAc into chitin in uivo (washed biomass was incubated in [3H]GlcNAc-containing medium lacking fungicide).

Inhibition of phosphatidylcholine and chitin biosynthesis in Pyricularia oryzae, Botrytis fabae and Fusarium graminearum by edifenphos

“…It has been reported that PC biosynthesis is inhibited by edifenphos and iprobenfos (Kodama et al, 1979(Kodama et al, , 1980Binks et al, 1990), and we have demonstrated that reduced levels of PC in a choline-requiring strain of Aspergillus nidulans (choA) were correlated with reductions in (a) the rate of in vivo chitin synthase, (b) colony radial growth rate, and (c) hyphal growth unit length . When the effects of edifenphos on F. graminearum, B. fabae and P. oryzae were examined, PC levels were found to be reduced but at markedly different fungicide concentrations (Table 1) this respect, P. oryzae was the most sensitive edifenphos followed by B. fabae with F. graminearum least sensitive, suggesting that the sensitivities of colony radial growth rates of the three fungi edifenphos may be due to a differential sensitivity inhibition of PC biosynthesis.…”

“…In F. graminearurn, we have previously shown that inhibition in colony radial growth rate by edifenphos is initially due to direct non-competitive inhibition of chitin synthase with no reduction in PC; only at edifenphos concentrations of about 250 p~ and above is a reduction in PC observed with additional inhibition of chitin synthase (Binks et al, 1990, Table 1). In the moderately sensitive fungus B. fabae, reductions in colony radial growth rate appear to be due to a combination of both direct inhibition of chitin synthase activity and an indirect inhibition of this enzyme due to a reduction in the PC content of the membrane ( Table 2).…”

“…However, conflicting views have emerged as to whether these fungicides act directly on chitin synthase or indirectly by the inhibition of PC biosynthesis (Maeda et al, 1970;De Waard et al, 1974;Brillinger, 1979;Kodama et al, 1979Kodama et al, , 1980Craig & Peberdy, 1983 a, b). Chitin synthase preparations from a number of fungi have been shown to be activated by PC (Duran & Cabib, 1978;Vermeulen & Wessels, 1983;Montgomery & Gooday, 1985;Binks et al, 1990). Previously, we have shown that edifenphos inhibits chitin synthase from F. graminearum in a non-competitive manner (Binks et al, 1990) and also inhibits PC biosynthesis, inducing a highly branched phenotype, without affecting specific growth rate (Wiebe et al, 1990).…”

“…Chitin synthase preparations from a number of fungi have been shown to be activated by PC (Duran & Cabib, 1978;Vermeulen & Wessels, 1983;Montgomery & Gooday, 1985;Binks et al, 1990). Previously, we have shown that edifenphos inhibits chitin synthase from F. graminearum in a non-competitive manner (Binks et al, 1990) and also inhibits PC biosynthesis, inducing a highly branched phenotype, without affecting specific growth rate (Wiebe et al, 1990). In this study, in vitro chitin synthase preparations from the three fungi were found to be equally sensitive to edifenphos (Fig.…”

Compounds active against cell walls of medically important fungi

“…In a recent study, the agricultural fungicide edifenphos, which is known to affect membrane phospholipid composition, was found to cause noncompetitive inhibition of chitin synthetase from Fusarium graminearum within specific concentration ranges (21).…”