Dicer-like 4 is involved in restricting the systemic movement of Zucchini yellow mosaic virus in Nicotiana benthamiana

[EN] Zucchini yellow mosaic virus (ZYMV) induces serious diseases in cucurbits. To create a tool to screen for resistance genes, we cloned a wild ZYMV isolate and inserted the visual marker Roseal to obtain recombinant clone ZYMV-Rosl. While in some plant-virus combinations Roseal induces accumulation of anthocyanins in infected tissues, ZYMV-Rosl infection of cucurbits did not lead to detectable anthocyanin accumulation. However, the recombinant virus did induce dark red pigmen-tation in infected tissues of the model plant Nicotiana ben-thamiana. In this species, ZYMV-Rosl multiplied efficiently in local inoculated tissue but only a few progeny particles estab-lished infection foci in upper leaves. We used this system to analyze the roles of Dicer-like (DCL) genes, core components of plant antiviral RNA silencing pathways, in ZYMV infection. ZYMV-Rosl local replication was not significantly affected in single DCL knockdown lines nor in double DCL2/4 and triple DCL2/3/4 knockdown lines. ZYMV-Rosl systemic accumula-tion was not affected in knockdown lines DCL1, DCL2, and DCL3. However in DCL4 and also in DCL2/4 and DCL2/3/4 knockdown lines, ZYMV-Rosl systemic accumulation dra-matically increased, which highlights the key role of DCL4 in restricting virus systemic movement. The effect of DCL4 on ZYMV systemic movement was confirmed with a wild-type version of the virus.