Convergent evolution of microRNAs in the same clusters toward cooperative regulation of target genes

MicroRNAs (miRNAs) are endogenously expressed small noncoding RNAs that regulate the expression of target genes in many biological processes. One salient feature of miRNAs is that their genomic locations are significantly clustered in discrete loci. By surveying the evolutionary patterns of miRNA clustering in animal species, we found that conserved miRNAs are significantly enriched in clusters via duplication or de novo formation. We proposed a convergent evolution model to explain how clustering helps new miRNAs survive and develop functions related to the pre-existing miRNAs in that cluster. To support our hypothesis, we presented evidence that clustered miRNAs have cooperative repression effects on target genes in different human tissues. We further tested our model by studying the evolutionary trajectory of the miR-17~92 cluster. By transfecting individual miRNAs into human and fly cell lines and extensively profiling the transcriptome alternation with mRNA-Seq, we confirmed that clustered miRNAs cooperatively target overlapping sets of genes and regulate genes from the same biological pathways. Our experimental results also demonstrated the functional co-adaptation between new and old miRNAs in the same cluster. Our functional co-adaptation model provides new insights into the mechanism and functional significance of miRNA clustering.