Sphingolipid C-9 Methyltransferases Are Important for Growth and Virulence but Not for Sensitivity to Antifungal Plant Defensins in Fusarium graminearum

The C-9-methylated glucosylceramides (GlcCers) are sphingolipids unique to fungi. They play important roles in fungal growth and pathogenesis, and they act as receptors for some antifungal plant defensins. We have identified two genes, FgMT1 and FgMT2, that each encode a putative sphingolipid C-9 methyltransferase (C-9-MT) in the fungal pathogen Fusarium graminearum and complement a Pichia pastoris C-9-MT-null mutant. The Delta Fgmt1 mutant produced C-9-methylated GlcCer like the wild-type strain, PH-1, whereas the Delta Fgmt2 mutant produced 65 to 75% nonmethylated and 25 to 35% methylated GlcCer. No Delta Fgmt1 Delta Fgmt2 double-knockout mutant producing only nonmethylated GlcCer could be recovered, suggesting that perhaps C-9-MTs are essential in this pathogen. This is in contrast to the nonessential nature of this enzyme in the unicellular fungus P. pastoris. The Delta Fgmt2 mutant exhibited severe growth defects and produced abnormal conidia, while the Delta Fgmt1 mutant grew like the wild-type strain, PH-1, under the conditions tested. The Delta Fgmt2 mutant also exhibited drastically reduced disease symptoms in wheat and much-delayed disease symptoms in Arabidopsis thaliana. Surprisingly, the Delta Fgmt2 mutant was less virulent on different host plants tested than the previously characterized Delta Fggcs1 mutant, which lacks GlcCer synthase activity and produces no GlcCer at all. Moreover, the Delta Fgmt1 and Delta Fgmt2 mutants, as well as the P. pastoris strain in which the C-9-MT gene was deleted, retained sensitivity to the antifungal plant defensins MsDef1 and RsAFP2, indicating that the C-9 methyl group is not a critical structural feature of the GlcCer receptor required for the antifungal action of plant defensins.