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Loqs and R2D2 act sequentially in the siRNA pathway in Drosophila

Abstract

In Drosophila melanogaster, the small interfering RNA (siRNA) pathway is triggered by exogenous double-stranded RNA (dsRNA) or upon viral infection. This pathway requires Dicer-2 (Dcr-2) in association with a dsRNA-binding protein (dsRBP) called R2D2. A potentially distinct siRNA pathway, which requires Dcr-2 in association with a different dsRBP, called Loquacious (Loqs), is activated by endogenous dsRNA derived from transposons, structured loci and overlapping transcripts. Here we show that different sources of dsRNA enter a common siRNA pathway that requires R2D2 and Loqs. R2D2 and loqs mutants show impaired silencing triggered by injection of exogenous dsRNA or by artificial and natural expression of endogenous dsRNA. In addition, we show that these dsRBPs function sequentially and nonredundantly in collaboration with Dcr-2. Loqs is primarily required for dsRNA processing, whereas R2D2 is essential for the subsequent loading of siRNAs into effector Ago–RISC complexes.

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Figure 1: R2D2 and Loqs are required for silencing triggered by dsRNA from endogenous or exogenous sources.
Figure 2: R2D2 and Loqs are required at different steps of the siRNA pathway.
Figure 3: R2D2 and Loqs are required in the endogenous siRNA pathway.
Figure 4: Requirements of R2D2 and Loqs for silencing of transposable elements (TEs).
Figure 5: The impact of Loqs and R2D2 on miRNAs.
Figure 6: The siRNA pathway in Drosophila.

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Acknowledgements

We thank C. Reinke, Y. Bei and Carthew laboratory members for valuable discussion; F. Valsecchi for help with bioinformatics; C. Horvath, E. Sontheimer and J. Brickner (Northwestern University) for sharing equipment and reagents; P. Zamore (University of Massachusetts Medical School) for loqsf00791; M. Siomi (Keio University School of Medicine) for ago2414 stocks; Q. Liu (University of Texas Southwestern Medical Center) for R2D21 and loqsKO stocks and G. Hannon (Cold Spring Harbor Laboratory) for anti-Loqs antibody. This work was supported by a grant from the US National Institutes of Health (GM68743) to R.W.C.

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J.T.M. and K.K. conceived and designed the experiments; J.T.M., K.K., P.-H.W., T.M.A. and N.J. performed the experiments; J.T.M., K.K. and R.W.C. analyzed the data; J.T.M. and R.W.C. wrote the paper.

Corresponding author

Correspondence to Richard W Carthew.

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Marques, J., Kim, K., Wu, PH. et al. Loqs and R2D2 act sequentially in the siRNA pathway in Drosophila. Nat Struct Mol Biol 17, 24–30 (2010). https://doi.org/10.1038/nsmb.1735

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