pre-miRNA profiles obtained through application of locked nucleic acids and deep sequencing reveals complex 5′/3′ arm variation including concomitant cleavage and polyuridylation patterns

• Published in: Nucleic acids research Vol. 40; no. 4; pp. 1424 - 1437
• Main Authors: Burroughs, A. Maxwell, Kawano, Mitsuoki, Ando, Yoshinari, Daub, Carsten O., Hayashizaki, Yoshihide
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 01.02.2012
• Subjects: Computational Biology; Data processing; HeLa Cells; High-Throughput Nucleotide Sequencing; Humans; Landscape; MicroRNAs - chemistry; MicroRNAs - metabolism; miRNA; nucleic acids; Nucleotide Motifs; Nucleotide sequence; Oligodeoxyribonucleotides - chemistry; Oligonucleotides - chemistry; Poly U - analysis; RNA; RNA Precursors - chemistry; RNA Precursors - metabolism; RNA Processing, Post-Transcriptional; RNA-Binding Proteins - metabolism; Sequence Analysis, RNA; Uridine - analysis; Uridine - metabolism
• ISSN: 0305-1048; 1362-4962; 1362-4962
• DOI: 10.1093/nar/gkr903

Recent research hints at an underappreciated complexity in pre-miRNA processing and regulation. Global profiling of pre-miRNA and its potential to increase understanding of the pre-miRNA landscape is impeded by overlap with highly expressed classes of other non coding (nc) RNA. Here, we present a data set excluding these RNA before sequencing through locked nucleic acids (LNA), greatly increasing pre-miRNA sequence counts with no discernable effect on pre-miRNA or mature miRNA sequencing. Analysis of profiles generated in total, nuclear and cytoplasmic cell fractions reveals that pre-miRNAs are subject to a wide range of regulatory processes involving loci-specific 3'- and 5'-end variation entailing complex cleavage patterns with co-occurring polyuridylation. Additionally, examination of nuclear-enriched flanking sequences of pre-miRNA, particularly those derived from polycistronic miRNA transcripts, provides insight into miRNA and miRNA-offset (moRNA) production, specifically identifying novel classes of RNA potentially functioning as moRNA precursors. Our findings point to particularly intricate regulation of the let-7 family in many ways reminiscent of DICER1-independent, pre-mir-451-like processing, introduce novel and unify known forms of pre-miRNA regulation and processing, and shed new light on overlooked products of miRNA processing pathways.