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2015 ; 25
(9
): 1401-9
Nephropedia Template TP
Chang TC
; Pertea M
; Lee S
; Salzberg SL
; Mendell JT
Genome Res
2015[Sep]; 25
(9
): 1401-9
PMID26290535
show ga
Precise regulation of microRNA (miRNA) expression is critical for diverse
physiologic and pathophysiologic processes. Nevertheless, elucidation of the
mechanisms through which miRNA expression is regulated has been greatly hindered
by the incomplete annotation of primary miRNA (pri-miRNA) transcripts. While a
subset of miRNAs are hosted in protein-coding genes, the majority of pri-miRNAs
are transcribed as poorly characterized noncoding RNAs that are 10's to 100's of
kilobases in length and low in abundance due to efficient processing by the
endoribonuclease DROSHA, which initiates miRNA biogenesis. Accordingly, these
transcripts are poorly represented in existing RNA-seq data sets and exhibit
limited and inaccurate annotation in current transcriptome assemblies. To
overcome these challenges, we developed an experimental and computational
approach that allows genome-wide detection and mapping of pri-miRNA structures.
Deep RNA-seq in cells expressing dominant-negative DROSHA resulted in much
greater coverage of pri-miRNA transcripts compared with standard RNA-seq. A
computational pipeline was developed that produces highly accurate pri-miRNA
assemblies, as confirmed by extensive validation. This approach was applied to a
panel of human and mouse cell lines, providing pri-miRNA transcript structures
for 1291/1871 human and 888/1181 mouse miRNAs, including 594 human and 425 mouse
miRNAs that fall outside protein-coding genes. These new assemblies uncovered
unanticipated features and new potential regulatory mechanisms, including links
between pri-miRNAs and distant protein-coding genes, alternative pri-miRNA
splicing, and transcripts carrying subsets of miRNAs encoded by polycistronic
clusters. These results dramatically expand our understanding of the organization
of miRNA-encoding genes and provide a valuable resource for the study of
mammalian miRNA regulation.