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Direct sequencing of Arabidopsis thaliana RNA reveals patterns of cleavage and polyadenylation

Abstract

It has recently been shown that RNA 3′-end formation plays a more widespread role in controlling gene expression than previously thought. To examine the impact of regulated 3′-end formation genome-wide, we applied direct RNA sequencing to A. thaliana. Here we show the authentic transcriptome in unprecedented detail and describe the effects of 3′-end formation on genome organization. We reveal extreme heterogeneity in RNA 3′ ends, discover previously unrecognized noncoding RNAs and propose widespread reannotation of the genome. We explain the origin of most poly(A)+ antisense RNAs and identify cis elements that control 3′-end formation in different registers. These findings are essential to understanding what the genome actually encodes, how it is organized and how regulated 3′-end formation affects these processes.

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Figure 1: Genome-wide patterns of A. thaliana RNA 3′-end formation.
Figure 2: Internal priming is rare or absent in DRS.
Figure 3: Identification of unannotated or previously undetected ncRNAs.
Figure 4: Identification of ncRNAs at sites affected by the exosome.
Figure 5: Most antisense expression derives from convergent gene pairs with overlapping 3′UTRs.
Figure 6: Cis-element analysis at cleavage sites.
Figure 7: Multifunctional cis elements within the same 3′UTR.

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Acknowledgements

We thank T. Walsh for computing support, E. Bayne and R. Lyons for comments on the manuscript, Trivalent Editing and P. Smith for proofreading and the Biotechnology and Biological Sciences Research Council (BB/H002286/1) (A.S., C.D., C.C., G.J.B. and G.G.S.), the Scottish Government (C.H. and G.G.S.) and the US National Institutes of Health (HG005230 and HG005279) (F.O. and P.M.M.) for funding.

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Contributions

G.G.S. and G.J.B. conceived and supervised the research. C.H. generated RNA samples. F.O. performed DRS. A.S. and C.C. analyzed the DRS data. C.D. and V.Z. did the molecular analyses of RNA. G.G.S. wrote the paper, and all authors read and commented on it.

Corresponding authors

Correspondence to Geoffrey J Barton or Gordon G Simpson.

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Competing interests

F.O. and P.M.M. are employees of Helicos Biosciences Corporation.

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Sherstnev, A., Duc, C., Cole, C. et al. Direct sequencing of Arabidopsis thaliana RNA reveals patterns of cleavage and polyadenylation. Nat Struct Mol Biol 19, 845–852 (2012). https://doi.org/10.1038/nsmb.2345

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