Browsing by Subject "RNAi"

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Extracellular vesicles isolated from dsRNA-sprayed barley plants exhibit no growth inhibition or gene silencing in Fusarium graminearum
(2022) Schlemmer, Timo; Lischka, Richard; Wegner, Linus; Ehlers, Katrin; Biedenkopf, Dagmar; Koch, Aline; Schlemmer, Timo; Institute of Phytomedicine, University of Hohenheim, Stuttgart, Germany; Lischka, Richard; Centre for BioSystems, Land Use and Nutrition, Institute of Phytopathology, Justus Liebig University, Giessen, Germany; Wegner, Linus; Intitute of Botany, Justus Liebig University, Giessen, Germany; Ehlers, Katrin; Intitute of Botany, Justus Liebig University, Giessen, Germany; Biedenkopf, Dagmar; Centre for BioSystems, Land Use and Nutrition, Institute of Phytopathology, Justus Liebig University, Giessen, Germany; Koch, Aline; Institute of Phytomedicine, University of Hohenheim, Stuttgart, Germany
Numerous reports have shown that incorporating a double-stranded RNA (dsRNA)-expressing transgene into plants or applying dsRNA by spraying it onto their leaves successfully protects them against invading pathogens exploiting the mechanism of RNA interference (RNAi). How dsRNAs or siRNAs are transferred between donor host cells and recipient fungal cells is largely unknown. It is speculated that plant extracellular vesicles (EVs) function as RNA shuttles between plants and their pathogens. Recently, we found that EVs isolated from host-induced gene silencing (HIGS) or spray-induced gene silencing (SIGS) plants contained dsRNA-derived siRNAs. In this study, we evaluated whether isolated EVs from dsRNA-sprayed barley ( Hordeum vulgare ) plants affected the growth of the phytopathogenic ascomycete Fusarium graminearum . Encouraged by our previous finding that dropping barley-derived EVs on F. graminearum cultures caused fungal stress phenotypes, we conducted an in vitro growth experiment in microtiter plates where we co-cultivated F. graminearum with plant EVs isolated from dsRNA-sprayed barley leaves. We observed that co-cultivation of F. graminearum macroconidia with barley EVs did not affect fungal growth. Furthermore, plant EVs containing SIGS-derived siRNA appeared not to affect F. graminearum growth and showed no gene silencing activity on F. graminearum CYP51 genes. Based on our findings, we concluded that either the amount of SIGS-derived siRNA was insufficient to induce target gene silencing in F. graminearum, indicating that the role of EVs in SIGS is minor, or that F. graminearum uptake of plant EVs from liquid cultures was inefficient or impossible.
Successful silencing of the mycotoxin synthesis gene TRI5 in fusarium culmorum and observation of reduced virulence in VIGS and SIGS experiments
(2022) Tretiakova, Polina; Voegele, Ralf Thomas; Soloviev, Alexander; Link, Tobias Immanuel
Crops constantly experience various biotic stresses during their life cycle, and Fusarium spp. remain one of the most serious groups of pathogens affecting plants. The ability to manipulate the expression of certain microorganism genes via RNAi creates the opportunity for new-generation dsRNA-based preparations to control a large number of diseases. In this study, we applied virus-induced gene silencing (VIGS), and spray-induced gene silencing (SIGS) to silence the trichothecene-producing gene TRI5 in F. culmorum as a means to reduce its aggressiveness on spring wheat. Treatment of the fungus with dsTRI5RNA in vitro reduced deoxynivalenol (DON) and 3-acetyldeoxynivalenol (3-A-DON) accumulations by 53–85% and 61–87%, respectively, and reduced TRI5 expression by 84–97%. VIGS decreased the proportion of infected wheat spikelets by 73%, but upregulation was observed for TRI5. SIGS on wheat leaves and ears using certain dsTRI5RNA amounts negatively impacted F. culmorum growth. However, when performing in vivo analyses of TRI5 mRNA levels, the upregulation of the gene was determined in the variants where fungal colonization was restricted, suggesting a compensatory reaction of the pathogen to RNAi.