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Two distinct mechanisms generate endogenous siRNAs from bidirectional transcription in Drosophila melanogaster

A Corrigendum to this article was published on 01 September 2008

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Abstract

Cis-natural antisense transcripts (cis-NATs) have been speculated to be substrates for endogenous RNA interference (RNAi), but little experimental evidence for such a pathway in animals has been reported. Analysis of massive Drosophila melanogaster small RNA data sets now reveals two mechanisms that yield endogenous small interfering RNAs (siRNAs) via bidirectional transcription. First, >100 cis-NATs with overlapping 3′ exons generate 21-nt and, based on previously published small RNA data, Dicer-2 (Dcr-2)–dependent, 3′-end modified siRNAs. The processing of cis-NATs by RNA interference (RNAi) seems to be actively restricted, and the selected loci are enriched for nucleic acid–based functions and include Argonaute-2 (AGO2) itself. Second, we report that extended intervals of the thickveins and klarsicht genes generate exceptionally abundant siRNAs from both strands. These siRNA clusters derive from atypical cis-NAT arrangements involving introns and 5′ or internal exons, but their biogenesis is similarly Dcr-2– and AGO2-dependent. These newly recognized siRNA pathways broaden the scope of regulatory networks mediated by small RNAs.

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Figure 1: Distinct size distributions of different small RNA–generating loci.
Figure 2: The overlap regions of a subset of 3′ cis-NAT loci preferentially generate siRNAs.
Figure 3: Most coexpressed 3′ cis-NAT pairs do not generate siRNAs.
Figure 4: The thickveins (tkv) and klarsicht (klar) loci define a distinct class of exceptionally abundant siRNAs derived from bidirectional transcription.
Figure 5: Biogenesis and structural features of klar siRNAs.

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Change history

  • 18 June 2008

    In the version of this article initially published, the Abstract did not convey that two conclusions regarding cis-NATs, presented in Table 1, were derived from analysis of previously published data (references 20 and 21 in the paper). The third sentence of the Abstract should read: “First, >100 cis-NATs with overlapping 3′ exons generate 21-nt and, based on previously published small RNA data, Dicer-2 (Dcr-2)–dependent, 3′-end modified siRNAs.” The correction has been made to the HTML and PDF versions of the article.

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Acknowledgements

We thank E. Hodges and G. Hannon for Solexa sequencing, A. Viale and the Sloan-Kettering Genomics Core for hybridizing Affymetrix microarrays, and N. Socci for assistance with microarray analysis. K.O. was supported by a grant from the Charles Revson Foundation. E.C.L. was supported by grants from the Burroughs Wellcome Foundation, the V Foundation for Cancer Research, the Sidney Kimmel Cancer Foundation and the US National Institutes of Health (R01-GM083300 and U01-HG004261).

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Contributions

N.L. performed the small RNA size analyses in Figure 1; R.M. performed some of the northern analysis in Figure 5; S.B. performed the cis-NAT bioinformatics; and K.O. constructed the small RNA libraries and performed the remaining functional tests. The contributions of S.B. and K.O. were considered equal. All authors contributed to the preparation of the manuscript.

Corresponding author

Correspondence to Eric C Lai.

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Okamura, K., Balla, S., Martin, R. et al. Two distinct mechanisms generate endogenous siRNAs from bidirectional transcription in Drosophila melanogaster. Nat Struct Mol Biol 15, 581–590 (2008). https://doi.org/10.1038/nsmb.1438

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