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DICER-LIKE 4 is required for RNA interference and produces the 21-nucleotide small interfering RNA component of the plant cell-to-cell silencing signal

Nature Genetics volume 37, pages 1356–1360 (2005)Cite this article

Abstract

In RNA interference1,2, the RNase-III enzyme Dicer3 processes exogenous double-stranded RNA into small interfering RNAs (siRNAs). siRNAs guide RNA-induced silencing complexes to cleave homologous transcripts, enabling gene-specific knock-down4. In plants, double-stranded RNA is processed into siRNA species of 21 nucleotides (nt) and 24 nt (ref. 5), but, unlike in nematodes6, the Dicer enzymes involved in this processing have not been identified. Additionally, in both plants and nematodes, systemic signals7,8,9,10 with RNA components convey the sequence-specific effects of RNA interference between cells. Here, we describe Arabidopsis thaliana mutants with altered silencing cell-to-cell movement beyond the vasculature. At least three SILENCING MOVEMENT DEFICIENT genes (SMD1, SMD2 and SMD3) are required for trafficking, the extent of which correlates with siRNA levels in the veins. Five alleles defective in synthesis of 21-nt, but not 24-nt, siRNAs carry mutations in Dicer-like 4 (DCL4) that are involved in biogenesis of trans-acting siRNAs11,12. We show that the biogenesis and function of trans-acting siRNA can be genetically uncoupled from a bona fide DCL4-dependent pathway that accounts for RNA interference and for production of the 21-nt siRNA component of the plant cell-to-cell silencing signal.

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Figure 1: Phenotypic and molecular characterization of class I, class II and class III mutants.
Figure 2: Phenotypic and molecular analysis of class IV mutants.
Figure 3: The five alleles defining class IV mutants correspond to loss-of-function and missense mutations in DCL4.
Figure 4: An integrated model for short-range and long-range cell-to-cell movement of RNA silencing in plants.

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Acknowledgements

We thank P. Brodersen and L. Navarro for critical reading of the manuscript, S. MacFarlane for providing TRV-PDS, M. Alioua for technical support and R. Wagner's team for greenhouse work. Research in our laboratory is supported by the Centre National de la Recherche Scientifique and an EMBO Young Investigator award attributed to O.V.

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  1. Institut de Biologie Moléculaire des Plantes du CNRS-UPR 2357, 12, rue du Général Zimmer, Strasbourg, 67084, Cedex, France

    Patrice Dunoyer, Christophe Himber & Olivier Voinnet

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  1. Patrice Dunoyer
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  2. Christophe Himber
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  3. Olivier Voinnet
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Correspondence to Olivier Voinnet.

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Supplementary information

Supplementary Fig. 1

Northern analysis of SULPHUR mRNA accumulation in the four mutant classes. (PDF 259 kb)

Supplementary Fig. 2

Quantitative RT-PCR (RT-qPCR) analyses of ta-siRNA target accumulation in the five independant alleles of class IV mutant. (PDF 916 kb)

Supplementary Note (PDF 83 kb)

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Dunoyer, P., Himber, C. & Voinnet, O. DICER-LIKE 4 is required for RNA interference and produces the 21-nucleotide small interfering RNA component of the plant cell-to-cell silencing signal. Nat Genet 37, 1356–1360 (2005). https://doi.org/10.1038/ng1675

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