Unknown,Transcriptomics,Genomics,Proteomics

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Improved annotation of C. elegans microRNAs by deep sequencing reveals structures associated with processing by Drosha and Dicer

ABSTRACT: MicroRNAs (miRNAs) are small regulatory RNAs that are essential in all studied metazoans. Research has focused on the prediction and identification of novel miRNAs, while little has been done to validate, annotate, and characterize identified miRNAs. Using Illumina sequencing, ~20 million small RNA sequences were obtained from Caenorhabditis elegans. Of the 175 miRNAs listed on the miRBase database, 106 were validated as deriving from a stem–loop precursor with hallmark characteristics of miRNAs. This result suggests that not all sequences identified as miRNAs belong in this category of small RNAs. Our large data set of validated miRNAs facilitated the determination of general sequence and structural characteristics of miRNAs and miRNA precursors. In contrast to previous observations, we did not observe a preference for the 59 nucleotide of the miRNA to be unpaired compared to the 59 nucleotide of the miRNA*, nor a preference for the miRNA to be on either the 59 or 39 arm of the miRNA precursor stem–loop. We observed that steady-state pools of miRNAs have fairly homogeneous termini, especially at their 59 end. Nearly all mature miRNA–miRNA* duplexes had two nucleotide 39 overhangs, and there was a preference for a uracil in the first and ninth position of the mature miRNA. Finally, we observed that specific nucleotides and structural distortions were overrepresented at certain positions adjacent to Drosha and Dicer cleavage sites. Our study offers a comprehensive data set of C. elegans miRNAs and their precursors that significantly decreases the uncertainty associated with the identity of these molecules in existing databases. Deep sequencing of small RNAs in wild-type (N2) mixed stage C. elegans, roughly 20 million sequencing reads were obtained

ORGANISM(S): Caenorhabditis elegans

SUBMITTER: Michael Warf

PROVIDER: E-GEOD-24704 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Improved annotation of C. elegans microRNAs by deep sequencing reveals structures associated with processing by Drosha and Dicer.

Warf M Bryan MB Johnson W Evan WE Bass Brenda L BL

RNA (New York, N.Y.) 20110209 4

MicroRNAs (miRNAs) are small regulatory RNAs that are essential in all studied metazoans. Research has focused on the prediction and identification of novel miRNAs, while little has been done to validate, annotate, and characterize identified miRNAs. Using Illumina sequencing, ∼20 million small RNA sequences were obtained from Caenorhabditis elegans. Of the 175 miRNAs listed on the miRBase database, 106 were validated as deriving from a stem-loop precursor with hallmark characteristics of miRNAs ...[more]

PMID: 21307183

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Project description:MicroRNA (miRNA) and endogenous siRNA (endo-siRNA) are two essential classes of small noncoding RNAs (sncRNAs) in eukaryotic organisms. The class of miRNA is diverse and there exist noncanonical miRNAs that bypass the canonical miRNA biogenesis pathway. In order to identify noncanonical miRNAs and endo-siRNAs responding to virus infection and study their potential function, we sequenced small-RNA species from cells lytically infected with murine gammaherpesvirus 68. In addition to 3 novel canonical miRNAs in mouse, two antisense miRNAs in virus and 25 novel noncanonical miRNAs, including miRNAs derived from tRNAs, snoRNAs and introns, in the host were identified. These noncanonical miRNAs exhibited features distinct from canonical miRNAs in the lengths and structures of miRNA hairpins as well as base pairings and first nucleotide preference. Many of the novel miRNAs are conserved in mammals. In addition to several known murine endo-siRNAs detected by the sequencing profiling, a novel locus in the mouse genome was identified to give rise to endo-siRNAs. This novel endo-siRNA locus is comprised of two tandem inverted B4 short interspersed nuclear elements (SINEs). Unexpectedly, the SINE-derived endo-siRNAs were found in a variety of sequencing data as well as virus-infected cells. Moreover, a murine miRNA was up-regulated more than 35 fold in infected than in mock-treated cells. The putative target genes of the viral and the up-regulated murine miRNAs were potentially involved in processes of gene transcription and protein phosphorylation and localized to membranes, suggesting their role in manipulating the host basal immune system during lytic infection. Our results extended the number of noncanonical miRNAs in mammals and shed new lights on their potential functions of lytic infection of MHV68. Mouse NIH 3T12 cells infectd with MHV68 (3 samples) and mock-treated (2 samples) were examined. Noncanonical microRNAs and endogenous siRNAs discovery in lytic infection of murine gammaherpesvirus MHV68 (NC_001826.2).

Dataset Information

Improved annotation of C. elegans microRNAs by deep sequencing reveals structures associated with processing by Drosha and Dicer

Publications

Improved annotation of C. elegans microRNAs by deep sequencing reveals structures associated with processing by Drosha and Dicer.

Similar Datasets

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