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Competition between ADAR and RNAi pathways for an extensive class of RNA targets

Nature Structural & Molecular Biology volume 18, pages 1094–1101 (2011)Cite this article

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Abstract

Adenosine deaminases that act on RNAs (ADARs) interact with double-stranded RNAs, deaminating adenosines to inosines. Previous studies of Caenorhabditis elegans indicated an antagonistic interaction between ADAR and RNAi machineries, with ADAR defects suppressed upon additional knockout of RNAi. This suggests a pool of common RNA substrates capable of engaging both pathways. To define and characterize such substrates, we examined small RNA and mRNA populations of ADAR mutants and identified a distinct set of loci from which RNAi-dependent short RNAs are markedly upregulated. At these same loci, we observed populations of multiply edited transcripts, supporting a specific role for ADARs in preventing access to the RNAi pathway for an extensive population of dsRNAs. Characterization of these loci revealed a substantial overlap with noncoding and intergenic regions, suggesting that the landscape of ADAR targets may extend beyond previously annotated classes of transcripts.

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Figure 1: Diverse consequences of dsRNA formation.
Figure 2: Small RNA accumulation at the F07B7 histone locus in the absence of ADAR activity.
Figure 3: Characteristics of ADAR-modulated RNA loci.
Figure 4: Dependence of ARL small RNAs on the RNAi machinery.
Figure 5: A substantial class of additional ARL-associated siRNAs are evident in 5′ phosphate–independent capture and sequencing.
Figure 6: Accumulation of a second population of siRNAs in cis to ARLs.
Figure 7: A-to-G changes in mRNA show unique enrichment for ARL, inverted repeat and transposon regions.

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Acknowledgements

We thank J. Gent, S. Gu, M. Stadler, H. Zhang, K. Artiles, J. Pak, L. Gracey, A. Sidow, Z. Weng, P. Lacroute and P. Parameswaran for help and suggestions; the Caenorhabditis Genetics Center, National Bioresource Project and B. Bass (University of Utah) for strains; Stanford Graduate Fellowship (D.W.), Stanford Dean's Fellowship and Machiah Foundation (A.T.L.); and the US National Institutes of Health (R01GM37706 to A.Z.F.) for support.

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Authors and Affiliations

  1. Department of Genetics, Stanford University School of Medicine, Stanford, California, USA

    Diane Wu & Andrew Z Fire

  2. Department of Pathology, Stanford University School of Medicine, Stanford, California, USA

    Ayelet T Lamm & Andrew Z Fire

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  1. Diane Wu
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Contributions

D.W. and A.Z.F. designed experiments and wrote the paper. D.W. prepared samples, created small RNA libraries and analyzed the data. A.T.L. created mRNA libraries and participated in discussions.

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Correspondence to Andrew Z Fire.

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Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–7 and Supplementary Methods (PDF 6558 kb)

Supplementary Data 1

Sample description (XLS 29 kb)

Supplementary Data 2

ADAR-affected sRNA expression over ARLs (XLS 116 kb)

Supplementary Data 3

Annotation enrichment of ARLs (XLS 24 kb)

Supplementary Data 4

Secondary siRNA expression for all genes (XLS 633 kb)

Supplementary Data 5

Genome-wide annotation of putative A-to-I editing (XLS 559 kb)

Supplementary Data 6

Statistics and raw data from Sanger sequencing (XLSX 72 kb)

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Wu, D., Lamm, A. & Fire, A. Competition between ADAR and RNAi pathways for an extensive class of RNA targets. Nat Struct Mol Biol 18, 1094–1101 (2011). https://doi.org/10.1038/nsmb.2129

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