Frequency and fate of microRNA editing in human brain

Abstract

Primary transcripts of certain microRNA (miRNA) genes (pri-miRNAs) are subject to RNA editing that converts adenosine to inosine (A→I RNA editing). However, the frequency of the pri-miRNA editing and the fate of edited pri-miRNAs remain largely to be determined. Examination of already known pri-miRNA editing sites indicated that adenosine residues of the UAG triplet sequence might be edited more frequently. In the present study, therefore, we conducted a large-scale survey of human pri-miRNAs containing the UAG triplet sequence. By direct sequencing of RT–PCR products corresponding to pri-miRNAs, we examined 209 pri-miRNAs and identified 43 UAG and also 43 non-UAG editing sites in 47 pri-miRNAs, which were highly edited in human brain. In vitro miRNA processing assay using recombinant Drosha-DGCR8 and Dicer-TRBP (the human immuno deficiency virus transactivating response RNA-binding protein) complexes revealed that a majority of pri-miRNA editing is likely to interfere with the miRNA processing steps. In addition, four new edited miRNAs with altered seed sequences were identified by targeted cloning and sequencing of the miRNAs that would be processed from edited pri-miRNAs. Our studies predict that ∼16% of human pri-miRNAs are subject to A→I editing and, thus, miRNA editing could have a large impact on the miRNA-mediated gene silencing.

Mammalian microRNAs: experimental evaluation of novel and previously annotated genes

“…These results indicate that mature edited miRNAs are very rare and difficult to distinguish above the background level of sequencing errors. The low frequency of editing in mature miRNAs was consistent with the findings that edited processed miRNAs are more than fourfold less common in mice relative to humans (Landgraf et al 2007), and are less common than edited miRNA precursors (Kawahara et al 2008). The latter observation might be due to rapid degradation or impaired processing, which has been shown for miR-142 (Yang et al 2006) and miR-151 (Kawahara et al 2007a).…”

“…In addition to editing in the miR-376 cluster described previously (Kawahara et al 2007b(Kawahara et al , 2008, we found another eight miRNAs that are edited within the seed of either the miRNA or the miRNA*. A-to-I editing could also affect miRNA loading, and thereby indirectly affect targeting.…”

“…Sixteen A-to-G events passed the filters and subsequent manual examination, all of which occurred only in the brain library ( Table 2). Five of these inferred editing sites were also observed in a low-throughput sequencing effort in human brain samples (Kawahara et al 2008), indicating that editing of some miRNAs is conserved between mammals. Consistent with that study, eight of 16 editing sites occurred in a UAG motif.…”

“…(Top) Sequencing data from our study are mapped to the mir-155 hairpin as in C. The mRNAs with 8mer and 7mer-A1 sites for the minor isoform were excluded from the analysis because these sites overlapped with 7mer-m8 sites for the major isoform. some miRNA precursors (Blow et al 2006;Landgraf et al 2007;Kawahara et al 2008). Because I pairs with C, such edits could change miRNA target recognition.…”

ADAR-mediated RNA editing in non-coding RNA sequences

“…These two editing sites did not affect cleavage of pri-miR-151 by Drosha-DGCR8 complex, but they inhibited cleavage of pre-miR-151 by Dicer-TRBP complex [101]. A large-scale survey of human pri-miRNA editing showed that 16% of human pri-miRNAs undergo A-to-I RNA editing, and may affect the Drosha or Dicer cleavage [102]. These findings suggest that A-to-I editing can modulate miRNA biogenesis.…”