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Total RNA sequencing reveals nascent transcription and widespread co-transcriptional splicing in the human brain

Nature Structural & Molecular Biology volume 18, pages 1435–1440 (2011)Cite this article

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Abstract

Transcriptome sequencing allows for analysis of mature RNAs at base pair resolution. Here we show that RNA-seq can also be used for studying nascent RNAs undergoing transcription. We sequenced total RNA from human brain and liver and found a large fraction of reads (up to 40%) within introns. Intronic RNAs were abundant in brain tissue, particularly for genes involved in axonal growth and synaptic transmission. Moreover, we detected significant differences in intronic RNA levels between fetal and adult brains. We show that the pattern of intronic sequence read coverage is explained by nascent transcription in combination with co-transcriptional splicing. Further analysis of co-transcriptional splicing indicates a correlation between slowly removed introns and alternative splicing. Our data show that sequencing of total RNA provides unique insight into the transcriptional processes in the cell, with particular importance for normal brain development.

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Figure 1: A large proportion of RNA-seq reads map to intronic regions.
Figure 2: Nascent transcription and co-transcriptional splicing.
Figure 3: A gradient of RNA levels within introns.
Figure 4: Exonic and intronic RNA levels for NRXN1.
Figure 5: Experimental validation of co-transcriptional splicing in consecutive exons.
Figure 6: Levels of co-transcriptional splicing within genes.

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Acknowledgements

We thank B. Röken at the Kolmården Zoo for sharing the chimpanzee tissue sample. We also acknowledge the staff members at the Uppsala Genome Center, who conducted the SOLiD sequencing. Financial support for this project was obtained from the Swedish Foundation for Strategic Research (L.F.), the Marcus Borgström Foundation (L.C. and L.F.) and the Göran Gustafsson Foundation (L.F.).

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  1. Lucia Cavelier and Lars Feuk: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Immunology, Genetics and Pathology, Science for Life Laboratory Uppsala, Rudbeck Laboratory, Uppsala University, Uppsala, Sweden

    Adam Ameur, Ammar Zaghlool, Jonatan Halvardson, Ulf Gyllensten, Lucia Cavelier & Lars Feuk

  2. Science for Life Laboratory, Karolinska Institutet Science Park, Stockholm, Sweden

    Anna Wetterbom

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Contributions

L.C. and L.F. conceived and designed the study. A.A., U.G., L.C. and L.F. coordinated experiments and analysis. A.A., J.H. and A.W. conducted the bioinformatics analysis. A.Z. and L.C. did the sample preparation and experimental analysis. All authors participated in discussions of different parts of the study. A.A., A.Z., L.C. and L.F. wrote the manuscript. All authors read and approved the manuscript.

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Correspondence to Lucia Cavelier or Lars Feuk.

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The authors declare no competing financial interests.

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Supplementary Figures 1–7, Supplementary Tables 1–10 and Supplementary Methods (PDF 2491 kb)

Supplementary Data 1

Lists of genes with high levels of intronic RNA. (XLS 199 kb)

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Ameur, A., Zaghlool, A., Halvardson, J. et al. Total RNA sequencing reveals nascent transcription and widespread co-transcriptional splicing in the human brain. Nat Struct Mol Biol 18, 1435–1440 (2011). https://doi.org/10.1038/nsmb.2143

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