The evolution of animal microRNA function Ryusuke Niwa and Frank J Slack MicroRNAs (miRNAs) are a large class of small RNAs that function as negative gene regulators in eukaryotes. They regulate diverse biological processes, and bioinformatics data indicate that each miRNA can control hundreds of gene targets, underscoring the potential influence of miRNAs on almost every genetic pathway. In addition to the roles in ontogeny, recent evidence has suggested the possibility that miRNAs have huge impacts on animal phylogeny. The dramatically expanding repertoire of miRNAs and their targets appears to be associated with major body-plan innovations as well as the emergence of phenotypic variation in closely related species. Research in the area of miRNA phylogenetic conservation and diversity suggests that miRNAs play important roles in animal evolution, by driving phenotypic variation during development. Addresses Department of Molecular, Cellular and Developmental Biology, KBT 936, Yale University, PO Box 208103, 266 Whitney Avenue, New Haven, CT 06520, USA Corresponding author: Slack, Frank J (frank.slack@yale.edu) Current Opinion in Genetics & Development 2007, 17:145–150 This review comes from a themed issue on Chromosomes and expression mechanisms Edited by Tom Misteli and Abby Demburg Available online 20th February 2007 0959-437X/$ – see front matter # 2007 Elsevier Ltd. All rights reserved. DOI 10.1016/j.gde.2007.02.004 Introduction A fundamental principle in molecular biology, the so- called ‘central dogma’, is that genes are generally protein- coding and genetic output is almost entirely transacted by proteins. With this view in mind, biologists have worked on the premise that evolution should be characterized by changes in genetic complexity that generate functionally novel proteins. However, an implication from studies in the past two decades is paradoxical: even though morpho- logically complex animals such as vertebrates possess tissues and organs that are lacking in phylogenetically basal animals, both lower and higher organisms encode a majority of the protein-coding gene families, such as transcription factors and signal transduction molecules [1  ,2,3  ]. Thus, the taxonomic distributions of protein- coding genes do not correspond to the dramatic increase in morphological complexity during animal evolution. Therefore, an idea arose that gene expression in the complex metazoan genomes required an additional level of regulation, such as by alternative splicing and non-coding RNAs [1  ,4]. Over the past few years, a new and surprisingly abundant class of RNA regulatory genes known as microRNAs (miRNAs) has been found to confer a novel layer of genetic regulation in cells [5–10]. miRNAs comprise a large family of 22-nucleotide single-stranded RNAs that silence gene expression by binding to target mRNAs. miRNA–mRNA binding usually involves strong base-pairing between the 5 0 end of a miRNA and its target complementary sequence in the 3 0 -untranslated region (3 0 UTR) of an mRNA, while additional base pairings can also contribute to the binding. miRNA binding appears to result in translational repres- sion and, in some cases, degradation of cognate mRNAs, causing partial or full silencing of the respective protein- coding genes. We now know that hundreds of distinct miRNA genes control a range of physiological processes in almost all eukaryotes, including development, growth, differentiation and metabolism [5–10]. miRNAs are cur- rently estimated to comprise 1–5% of animal genes, mak- ing them one of the most abundant classes of gene regulators [5]. Thus, this discovery introduces a new set of evolutionary mechanisms that has the potential to have profoundly influenced phenotypic complexity and diver- sity during animal phylogeny. In this brief review, we summarize recent advance of our knowledge about how the conservation and variety of miRNAs and miRNA targets have evolved, and the potential roles of miRNAs during animal evolution. Evolution of miRNA families and their expression let-7: a founder member of evolutionally conserved miRNAs The first example of an evolutionally conserved miRNA is let-7, which was originally identified as an essential reg- ulator of developmental timing in the nematode Caenor- habditis elegans [11]. Expression of let-7 miRNA is undetectable until the late larval stages in C. elegans, when let-7 directs larval to adult cell fate transitions in many tissues. Soon after the discovery of C. elegans let-7, it was demonstrated that let-7 belongs to a larger gene family that has been amazingly conserved across almost all groups of bilaterally symmetrical animals (bilaterians; Figure 1) [12,13]. Moreover, the temporal expression pattern of let-7 also appears to be highly conserved in all organisms examined. For example, let-7 in the fruit fly Drosophila melanogaster is absent from embryonic and larval stages and first appears at the entry of metamorphosis [12]. In mollusc and a polychaete annelids, let-7 miRNAs are detected in adults but not in larvae. Furthermore, in zebrafish and www.sciencedirect.com Current Opinion in Genetics & Development 2007, 17:145–150