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. 2014 Mar 13;6(5):906-15.
doi: 10.1016/j.celrep.2014.01.037. Epub 2014 Feb 20.

RNA-DNA differences are generated in human cells within seconds after RNA exits polymerase II

Isabel X Wang  1 , Leighton J Core  2 , Hojoong Kwak  3 , Lauren Brady  4 , Alan Bruzel  5 , Lee McDaniel  6 , Allison L Richards  7 , Ming Wu  8 , Christopher Grunseich  9 , John T Lis  10 , Vivian G Cheung  11
Affiliations

RNA-DNA differences are generated in human cells within seconds after RNA exits polymerase II

Isabel X Wang et al. Cell Rep. .

Abstract

RNA sequences are expected to be identical to their corresponding DNA sequences. Here, we found all 12 types of RNA-DNA sequence differences (RDDs) in nascent RNA. Our results show that RDDs begin to occur in RNA chains ~55 nt from the RNA polymerase II (Pol II) active site. These RDDs occur so soon after transcription that they are incompatible with known deaminase-mediated RNA-editing mechanisms. Moreover, the 55 nt delay in appearance indicates that they do not arise during RNA synthesis by Pol II or as a direct consequence of modified base incorporation. Preliminary data suggest that RDD and R-loop formations may be coupled. These findings identify sequence substitution as an early step in cotranscriptional RNA processing.

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Figures

Figure 1
Figure 1
GRO-seq and PRO-seq analysis. (A) Schematic of GRO-seq and PROseq. (B) Comparison between GRO-seq and PRO-seq. Sense and antisense transcripts associated with transcription start sites (TSS) are shown for GRO-seq and PRO-seq samples. The slight shift of the PRO-seq promoter-proximal peak downstream relative to the GRO-seq peak is because the PRO-seq reads that were less than 35 nucleotides were not mapped in the analysis, and because GRO-seq maps 5’ ends and PRO-seq maps 3’ ends of nascent RNAs. (C) mRNA-seq, chromatin-bound nascent RNA-seq, GRO-seq and PRO-seq results for two representative genes, UVRAG and CAPZB. For genes with proximal Pol II pausing such as UVRAG, there are more reads mapping to the 5’ ends of genes in both GRO-seq and PRO-seq samples. Schematic gene structure is aligned to mRNA-seq results, with boxes representing exons, lines representing introns and arrowheads showing direction of transcription. Coverage is calculated using bin size of ~ 1500 bp and 600 bp, respectively. (D) Scatter plot of gene expression levels from GRO-seq and mRNA-seq (FPKM>0.1). Results from GM12750 (shown) and GM12004 are similar (r=0.45 for both samples). Heatmap indicates frequency of different expression levels.
Figure 2
Figure 2
Analysis steps to identify RNA-DNA sequence differences. See also Table S1–S3.
Figure 3
Figure 3
RNA-DNA differences in very nascent transcripts. Distributions of RDD types (A) in GRO-seq samples of two individuals, (B) in PRO-seq. RDD types were ordered as in (A) and C-to-T RDDs for the PRO-seq sample.
Figure 4
Figure 4
Locations of RDD sites within sequencing reads. (A) Locations of RDD sites along GRO-seq reads. Only reads that have defined 3’ ends (reads that contain 3’ adapter sequences) were included in our analysis. (B) Locations of RDD sites along PRO-seq reads. Schematic diagrams indicate the locations of the different segments of GRO-seq (A) and PRO-seq (B) transcripts along the sequence reads. (C) Locations of RDD sites along mRNA-seq reads. (D) R-loop forming sequences are enriched in regions immediately adjacent to RDD sites. Average R-loop scores for 2 kb of regions up and downstream of RDD sites are shown. RDD sites have significantly higher R-loop scores (P<0.001, t-test) than random control sites.
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References

    1. Alon S, Mor E, Vigneault F, Church G, Locatelli F, Galeano F, Gallo A, Shomron N, Eisenberg E. Systematic identification of edited microRNAs in the human brain. Genome Res. 2012;22:1533–1540. - PMC - PubMed
    1. Ardehali MB, Lis JT. Tracking rates of transcription and splicing in vivo. Nature structural & molecular biology. 2009;16:1123–1124. - PubMed
    1. Athanasiadis A, Rich A, Maas S. Widespread A-to-I RNA editing of Alu-containing mRNAs in the human transcriptome. PLoS Biol. 2004;2:e391. - PMC - PubMed
    1. Bahn J, Lee J, Li G, Greer C, Peng G, Xiao X. Accurate identification of A-to-I RNA editing in human by transcriptome sequencing. Genome Res. 2011;29:29. - PMC - PubMed
    1. Bar-Yaacov D, Avital G, Levin L, Richards A, Hachen N, Rebolledo Jaramillo B, Nekrutenko A, Zarivach R, Mishmar D. RNA-DNA differences in human mitochondria restore ancestral form of 16S ribosomal RNA. Genome Res. 2013 - PMC - PubMed

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