Physiological effects of fenpropimorph on wild-type Saccharomyces cerevisiae and fenpropimorph-resistant mutants

Abstract

Fenpropimorph-resistant mutants of Saccharomyces cerevisiae were isolated by a gradient selection procedure. The mutants were cross-resistant to other morpholines (fenpropidin, dodemorph, tridemorph) and 15-azasterol, but were susceptible to azoles (miconazole, clotrimazole, ketoconazole) and nystatin. In the absence of fenpropimorph, the major sterol produced by the mutants and the parental strain was ergosterol. In the presence of fenpropimorph, ignosterol (ergosta-8,14-dien-3j-ol) was the major sterol produced by the mutants and the parental strain. The resistance to fenpropimorph involves two recessive genes, each of which allows a semiresistance, when they are isolated apart from one another. Strain JR4 (erg3 ergll), which produces 14-methylfecosterol [14kx-methyl-ergosta-8,24(28)-dien-33-ol] as the major sterol in the presence or absence of fenpropimorph, was also found to be resistant to the drug. The growth inhibitory effect of fenpropimorph on wild-type cells appears to be linked to the production of ignosterol. The uptake of exogenous sterol by wild-type cells was greatly enhanced in the presence of fenpropimorph. The growth inhibition caused by fenpropimorph could only be overcome with bulk levels of exogenous C-5,6-unsaturated sterols.The morpholine fenpropimorph, 4-{3-[4-(1,1-dimethylethyl) phenyl]-2-methylpropyl}-2,6(cis)-dimethyl-morpholine, has been successfully used in agriculture against fungi that cause powdery mildew of cereals, cereal rusts, and leaf blotch of barley (3). This antifungal agent inhibits two enzymes of the ergosterol biosynthetic pathway. Experiments conducted in vivo and in vitro have revealed that the A'7 isomerase of Saccharomyces cerevisiae is inhibited at lower concentrations of fenpropimorph than is the C-14-reductase, while higher concentrations inhibit both enzymes (1, 13). As a consequence, exposure of susceptible fungal cultures to fenpropimorph leads to the accumulation of sterolic intermediates such as ergosta-8,14,24(28)-trienol, ergosta-8,14,22-trienol, ergosta-8,24(28)-dienol (fecosterol), ergosta-8,14-dienol (ignosterol), and ergosta-8-enol (1, 14).We have shown that sterols mediate at least four physiological functions in S. cerevisiae (17,19). One of these, the "sparking function," is essential and has an absolute requirement for C-5,6-unsaturated sterols. In wild-type S. cerevisiae, this requirement is satisfied by ergosterol. A recent study suggested that the growth inhibitory effect of fenpropimorph is not due to the accumulation of sterol intermediates but is related to the depletion of ergosterol (12). Although no field pathogen or laboratory strain has been reported with a significantly decreased susceptibility to fenpropimorph (1), we reasoned that some mutants resistant to the antifungal agent might have an altered sparking function such that a C-5,6 unsaturation is not required. Resistant mutants could help us probe the biochemical events in the sparking function.In this study, we report the isolation of new fenpropimorph-resistant strains of S. cer...

A conserved membrane-binding domain targets proteins to organelle contact sites

“…The genes encoding Osh1p and Nvj2p have a co-fitness interaction (Hillenmeyer et al, 2008) and genetically interact with the ergosterol biosynthesis gene ERG11 (Kohl et al, 2010). Mutations in ERG11 also cause fenpropimorph resistance (Lorenz and Parks, 1991). Together, these findings suggest that Osh1p and Nvj2p participate in a sterol trafficking or signaling pathway at the NVJ.…”

Cloning by Metabolic Interference in Yeast and Enzymatic Characterization of Arabidopsis thaliana Sterol Δ7-Reductase

“…This is known as the "sparking effect," which is probably related to a cell cycle control mechanism in wild type strains (20). This requirement is abolished in cells harboring fen1 and/or fen2 gene mutations (21,22) suggesting to use strain FY1679-28C, a naturally occurring fen1 mutant, as a host for the ⌬7-red screening. We also chose a plant, A. thaliana as cDNA source because of the high ⌬7-red activity present in plant microsomes 2 and its low relative genome complexity (23,24).…”

A sterol C-14 reductase encoded by FgERG24B is responsible for the intrinsic resistance of Fusarium graminearum to amine fungicides

“…The sterol D 8 -D 7 isomerase encoded by FgERG2 has been shown as a second target of amines in several fungal species (Baloch et al, 1984;Baloch & Mercer, 1987;Debieu et al, 1992Debieu et al, , 1998Debieu et al, , 2000Lorenz & Parks, 1991;Schneegurt & Henry, 1992;Ziogas et al, 1991). Searching the F. graminearum genome database showed that the fungus has a single FgERG2 gene (Da Silva Ferreira et al, 2005).…”

“…Inhibition of the sterol C-14 reductase and sterol D 8 -D 7 isomerase leads to deficiency of ergosterol, but accumulation of fecosterol or D 8 -sterols, which are toxic to fungal cells (Lorenz & Parks, 1991;Marcireau et al, 1990;Steel et al, 1989).…”

“…Biochemical studies on the mode of action of amine fungicides have shown that they inhibit ergosterol biosynthesis at the step subsequent to C-14 demethylation and/or the step of D 8 -D 7 isomerization in several fungal species (Baloch et al, 1984;Baloch & Mercer, 1987;Debieu et al, 1992Debieu et al, , 1998Debieu et al, , 2000Lorenz & Parks, 1991;Schneegurt & Henry, 1992;Ziogas et al, 1991). Inhibition of the sterol C-14 reductase and sterol D 8 -D 7 isomerase leads to deficiency of ergosterol, but accumulation of fecosterol or D 8 -sterols, which are toxic to fungal cells (Lorenz & Parks, 1991;Marcireau et al, 1990;Steel et al, 1989).…”