Different components of the RNAi machinery are required for conidiation, ascosporogenesis, virulence, DON production and fungal inhibition by exogenous dsRNA in the Head Blight pathogenFusarium graminearum

Abstract

AbstractGene silencing through RNA interference (RNAi) shapes many biological processes in filamentous fungi, including pathogenicity. In this study we explored the requirement of key components of fungal RNAi machinery, including DICER-like 1 and 2 (FgDCL1,FgDCL2), ARGONAUTE 1 and 2 (FgAGO1,FgAGO2), AGO-interacting proteinFgQIP (QDE2-interacting protein), RecQ helicase (FgQDE3), and four RNA-dependent RNA polymerases (FgRdRP1,FgRdRP2,FgRdRP3,FgRdRP4), in the ascomycete mycotoxin-producing fungal pathogenFusarium graminearum(Fg) for sexual and asexual multiplication, pathogenicity, and its sensitivity to double-stranded (ds)RNA. We corroborate and extend earlier findings that conidiation, ascosporogenesis and Fusarium Head Blight (FHB) symptom development require an operable RNAi machinery. The involvement of RNAi in conidiation is dependent on environmental conditions as it is detectable only under low light (< 2 µmol m−2s−1). Although both DCLs and AGOs partially share their functions, the sexual ascosporogenesis is mediated primarily byFgDCL1 andFgAGO2, whileFgDCL2 andFgAGO1 contribute to asexual conidia formation and germination.FgDCL1 andFgAGO2 also account for pathogenesis as their knock-out (KO) results in reduced FHB development. Apart from KO mutantsΔdcl2andΔago1, mutantsΔrdrp2, Δrdrp3, Δrdrp4, Δqde3andΔqipare strongly compromised for conidiation, while KO mutations in allRdPRs, QDE3andQIPstrongly affect ascosporogenesis. Analysis of trichothecenes mycotoxins in wheat kernels showed that the relative amount of deoxynivalenol (DON), calculated as [DON] per amount of fungal genomic DNA, was reduced in all spikes infected with RNAi mutants, suggesting the possibility that the fungal RNAi pathways affectFg’s DON production in wheat spikes. Moreover, gene silencing by exogenous target gene specific dsRNA (spray-induced gene silencing, SIGS) is dependent on fungal DCLs, AGOs, and QIP, but not on QDE3. Together these data show that inF. graminearumdifferent key components of the RNAi machinery are crucial in different steps of fungal development and pathogenicity.