MicroRNAs to Nanog, Oct4 and Sox2 coding regions modulate embryonic stem cell differentiation

Abstract

Most miRNA target sequences that have been studied reside in the 3′ UTR, the part of the messenger RNA that is downstream of the coding region. In this work Rigoutsos and colleagues demonstrate that the coding regions of several genes encoding transcription factors involved in the maintenance of stem cell identity, such as Nanog, Oct4, and Sox2, have miRNA target sites. Three miRNAs that are upregulated when embryonic stem cells are induced to differentiate bind these sites in various combinations, and thereby confer specific phenotypes. The coding regions of several genes that encode transcription factors involved in maintenance of stem cell identity, such as Nanog, Oct4, and Sox2, have miRNA target sites. Three miRNAs that are upregulated when embryonic stem cells are induced to differentiate bind these sites in various combinations, and thereby confer specific phenotypes. MicroRNAs (miRNAs) are short RNAs that direct messenger RNA degradation or disrupt mRNA translation in a sequence-dependent manner1,2,3,4,5,6,7. For more than a decade, attempts to study the interaction of miRNAs with their targets were confined to the 3′ untranslated regions of mRNAs1, fuelling an underlying assumption that these regions are the principal recipients of miRNA activity. Here we focus on the mouse Nanog, Oct4 (also known as Pou5f1) and Sox2 genes8,9,10,11 and demonstrate the existence of many naturally occurring miRNA targets in their amino acid coding sequence (CDS). Some of the mouse targets analysed do not contain the miRNA seed, whereas others span exon–exon junctions or are not conserved in the human and rhesus genomes. miR-134, miR-296 and miR-470, upregulated on retinoic-acid-induced differentiation of mouse embryonic stem cells, target the CDS of each transcription factor in various combinations, leading to transcriptional and morphological changes characteristic of differentiating mouse embryonic stem cells, and resulting in a new phenotype. Silent mutations at the predicted targets abolish miRNA activity, prevent the downregulation of the corresponding genes and delay the induced phenotype. Our findings demonstrate the abundance of CDS-located miRNA targets, some of which can be species-specific, and support an augmented model whereby animal miRNAs exercise their control on mRNAs through targets that can reside beyond the 3′ untranslated region.