Identifications of conserved 7-mers in 3'-UTRs and microRNAs in Drosophila

Abstract

Background: MicroRNAs (miRNAs) are a class of endogenous regulatory small RNAs which play an important role in posttranscriptional regulations by targeting mRNAs for cleavage or translational repression. The base-pairing between the 5'-end of miRNA and the target mRNA 3'-UTRs is essential for the miRNA:mRNA recognition. Recent studies show that many seed matches in 3'-UTRs, which are fully complementary to miRNA 5'-ends, are highly conserved. Based on these features, a two-stage strategy can be implemented to achieve the de novo identification of miRNAs by requiring the complete base-pairing between the 5'-end of miRNA candidates and the potential seed matches in 3'-UTRs.

Results: We presented a new method, which combined multiple pairwise conservation information, to identify the frequently-occurred and conserved 7-mers in 3'-UTRs. A pairwise conservation score (PCS) was introduced to describe the conservation of all 7-mers in 3'-UTRs between any two Drosophila species. Using PCSs computed from 6 pairs of flies, we developed a support vector machine (SVM) classifier ensemble, named Cons-SVM and identified 689 conserved 7-mers including 63 seed matches covering 32 out of 38 known miRNA families in the reference dataset. In the second stage, we searched for 90 nt conserved stem-loop regions containing the complementary sequences to the identified 7-mers and used the previously published miRNA prediction software to analyze these stem-loops. We predicted 47 miRNA candidates in the genome-wide screen.

Conclusion: Cons-SVM takes advantage of the independent evolutionary information from the 6 pairs of flies and shows high sensitivity in identifying seed matches in 3'-UTRs. Combining the multiple pairwise conservation information by the machine learning approach, we finally identified 47 miRNA candidates in D. melanogaster.

Figure 1

The flowchart of the method. The whole method consists of two stages: in the first stage, conserved 7-mers are identified by considering all 7-mers' conservation patterns in six pairs of flies; in the second stage, pre-miRNAs and mature miRNAs are predicted by adding seed-matching information into published miRNA prediction methods in the whole genome.

Figure 2

Comparions of the results using the three methods. The number in each block indicates the corresponding number of 7-mers in that part. The number in the parenthesis indicates the number of reference miRNA families in that block.

Figure 3

The 689 conserved 7-mers identified by Cons-SVM matching with the 59 reference miRNAs. Much more sites matched with the 1–7 nt or 2–8 nt of the mature miRNAs.

Figure 4

The nucleotide composition of the 59 reference miRNAs. The 5' first nucleotide of mature miRNAs significantly favours "U". Other sites do not show similar nucleotide bias. The logo plot is produced by WebLogo [49].