Comprehensive analysis of RNA-Seq data reveals extensive RNA editing in a human transcriptome

Abstract

A n A ly s i s RNA editing is a post-transcriptional event that recodes hereditary information. Here we describe a comprehensive profile of the RNA editome of a male Han Chinese individual based on analysis of ~767 million sequencing reads from poly(A) + , poly(A) − and small RNA samples. We developed a computational pipeline that carefully controls for false positives while calling RNA editing events from genome and whole-transcriptome data of the same individual. We identified 22,688 RNA editing events in noncoding genes and introns, untranslated regions and coding sequences of protein-coding genes. Most changes (~93%) converted A to I(G), consistent with known editing mechanisms based on adenosine deaminase acting on RNA (ADAR). We also found evidence of other types of nucleotide changes; however, these were validated at lower rates. We found 44 editing sites in microRNAs (miRNAs), suggesting a potential link between RNA editing and miRNAmediated regulation. Our approach facilitates large-scale studies to profile and compare editomes across a wide range of samples.RNA editing is an integral step in generating the diversity and plasticity of cellular RNA signatures. Most editing events convert A to I(G) (adenosine to inosine, which is translated as guanosine), and are catalyzed by the double-stranded RNA-specific ADAR family of proteins. On the basis of its overabundance in repetitive Alu elements and the brain transcriptome 1-3 , RNA editing has been viewed as a key determinant in primate evolution and the development of higher brain functions 4 . Many outstanding questions on the extent and consequences of RNA editing in humans remain unanswered, despite extensive documentation of edited sites through bioinformatics approaches 5-9 and the reported roles of editing in altering genetic messages and other post-transcriptional events such as RNA splicing and miRNA regulation 2,10-12 . Global and unequivocal identification of RNA editing targets represents a critical first step in further understanding this post-transcriptional modification. This calls for complete information on whole-genome and transcriptome sequences from the same individual, so as to exclude polymorphisms and mutations among populations, as well as experimental approaches with the necessary high-throughput sequencing and base resolution 13,14 . Whole-transcriptome deep-sequencing technologies (e.g., RNASeq) [15][16][17] , with their capacity to simultaneously assay the entire transcriptome, represent an excellent choice of tool in this regard. Recent studies reporting the use of target-specific RNA-Seq 4,18 , the combination of DNA capture and parallel sequencing 19 , and mRNA-Seq 20,21 to find human RNA editing sites attest to the notion that this strategy is advantageous in addressing many outstanding questions of the editing phenomenon and its implications on the transcriptome.In this study, we used RNA-Seq to identify post-transcriptional editing events. Our unbiased and in-depth approach revealed many editing sites in transcripts corresp...

ADAR-mediated RNA editing in non-coding RNA sequences

“…Recent analysis indicated that 20% of the editing sites in the 3′-UTRs may alter miRNA target sites [23]. These results suggest RNA editing may regulate gene expression by editing miRNA target sites in 3′-UTRs.…”

“…Approximately 93% of these editing sites convert adenosine to inosine, consistent with known A-to-I RNA editing [23]. Inosine is recognized as guanosine in protein-RNA interfaces and base pairing.…”

“…Mammalian glutamate receptors and serotonin receptors and Drosophila sodium channels are examples of regions with such selective editing [25]. However, most of the pre-mRNA editing sites are in non-coding regions such as UTRs and introns [23,64]. These editing sites are usually non-selectively modified by ADARs.…”

“…However, this study only monitored five edited 3′-UTRs by fusing a red fluorescent protein gene in C. elegans [70]. In fact, many RNA editing events have been identified in UTRs, especially in mammals [23,64,74,75]. However, the function of these RNA editing sites remains unknown.…”

“…Therefore, it is possible that dsRNA binding proteins, such as ADARs, interact with the structural RNAs [23,64,74,75]. ADARs can convert adenosine to inosine in AC mismatches and create IC pairs to stabilize the double-stranded structures.…”