Clin Genet 2008: 74: 296–306 # 2008 The Authors Printed in Singapore. All rights reserved Journal compilation # 2008 Blackwell Munksgaard CLINICAL GENETICS doi: 10.1111/j.1399-0004.2008.01076.x Review MicroRNAs in development and disease Erson AE, Petty EM. MicroRNAs in development and disease. AE Ersona and EM Pettyb,c # Clin Genet 2008: 74: 296–306. Blackwell Munksgaard, 2008 aDepartment of Biological Sciences, Middle East Technical University (METU), Since the discovery of microRNAs (miRNAs) in Caenorhabditis elegans, Ankara, Turkey, and bDepartment of mounting evidence illustrates the important regulatory roles for miRNAs Internal Medicine and cDepartment of in various developmental, differentiation, cell proliferation, and Human Genetics, University of Michigan, apoptosis pathways of diverse organisms. We are just beginning to MI, USA elucidate novel aspects of RNA mediated gene regulation and to understand how heavily various molecular pathways rely on miRNAs for their normal function. miRNAs are small non-protein-coding transcripts that regulate gene expression post-transcriptionally by targeting Key words: cancer – development – messenger RNAs (mRNAs). While individual miRNAs have been microRNA – viral infection specifically linked to critical developmental pathways, the deregulated Corresponding author: Elizabeth M Petty, expression of many miRNAs also has been shown to have functional Department of Internal Medicine, significance for multiple human diseases, such as cancer. We continue to University of Michigan, 5220 MSRB III, discover novel functional roles for miRNAs at a rapid pace. Here, we 1150 West Medical Center Drive, Ann summarize some of the key recent findings on miRNAs, their mode of Arbor, MI 48109-0640, USA. action, and their roles in both normal development and in human Tel.: 1(734) 763-2532; pathology. A better understanding of how miRNAs operate during the fax: 1(734) 647-7979; normal life of a cell as well as in the pathogenesis of disease when e-mail: [email protected] deregulated will provide new avenues for diagnostic, prognostic, and Received 2 May 2008, revised and therapeutic applications. accepted for publication 3 July 2008 When the Human Genome Project was com- been widely investigated in various eukaryotic or- pleted, the number of genes in our genome was ganisms (1–3). By 2002, the miRNA registry (ver- estimated to be around 20–25 000, far fewer than sion 1) had 218 miRNA entries for primates, the initially predicted 100 000. All known or pre- rodents, birds, fish, worms, flies, plants and vi- dicted protein-coding genes are thought to be ex- ruses. This number dramatically increased over pressed only from a small percentage (i.e. 1.5–2%) 6 years to 6396 as of April 2008 (version 11) (4– of the whole genome. These figures are poor in- 6). This noteworthy discovery rate during a 6-year dicators of the actual genetic complexity in hu- time period along with the increasing evidence for mans. In fact, limiting the definition of Ôgene’ to the regulatory roles of many miRNAs in various describe only protein-coding units is not compre- cellular processes, such as development and differ- hensive enough either, to define genetic complex- entiation, are clear indicators of the considerable ity. As of today, the majority of the human complexity of genetic information processing in genome is hypothesized to code for many non- cells. Current estimations predict that about protein coding structural and regulatory RNAs, 30% of all the human genes are regulated by miR- including microRNAs (miRNAs), that play criti- NAs (7) and that a single miRNA can potentially cal and vital roles in generating genomic complex- target around 200 different transcripts that may ity and diversity within Homo sapiens and, also, function in different pathways in the cell (8). between species. The nomenclature for miRNAs has yet to be miRNAs are small non-protein-coding tran- fully standardized in the literature, but, in general, scripts of 16–29 nucleotide-long RNAs that regu- miRNA genes are designated by the italicized pre- late gene expression post-transcriptionally by fix Ômir’. The unitalicized prefix ÔmiR’ is used to targeting messenger RNAs (mRNAs). Since the indicate the mature form of the miRNA. The pre- initial discovery in Caenorhabditis elegans in fixes Ômir’ or ÔmiR’ are followed in most cases by 1993, miRNA-dependent gene regulation has a dash and then a number, such as miR-127. A set 296 MiRNAs in development and disease of guidelines for miRNA annotation is suggested are also likely to be transcribed by RNA polymer- by Ambros et al. (9). ase III (11). Given the large number of recent papers in the The general miRNA biogenesis pathway (Fig. 1) scientific literature describing the functional roles involves a primary miRNA (pri-miRNA) tran- of miRNAs in biological processes such as devel- script (several hundred base pairs to several kilo- opment, differentiation, cell proliferation, and base pairs) that is subsequently cleaved by the apoptosis, it is clear that miRNAs are critical reg- nuclear RNase III enzyme, Drosha, and its partner ulatory factors important for normal health and DGCR8 (DiGeorge syndrome critical region gene are also highly relevant to disease processes. Here, 8) forming the 60–70 nucleotide-long precursor we present a brief outline of miRNA biogenesis miRNA (pre-miRNA) with a 3# overhang of two and action mechanisms with an emphasis on key nucleotides (12, 13). The pre-miRNA is then trans- roles of miRNAs in normal development and ported to the cytoplasm through the nuclear export human pathology. protein, exportin-5, together with Ran-guanosine triphosphate (14). Cytoplasmic RNase III, Dicer, and its RNA-binding partner TRBP, human immunodeficiency virus (HIV)-1 transactivating miRNA biogenesis responsive element (TAR) RNA-binding protein, Similar to mRNAs, miRNAs can be transcribed in cleave the pre-miRNA into a 20–25 nucleotide a time- and tissue-specific manner. Many miR- mature miRNA duplex. This double-stranded NAs are transcribed by RNA polymerase II and RNA then assembles into a multiprotein complex, have a poly(A) tail and a 5# cap (10). However, RNA-induced silencing complex (RISC), which is exceptions do exist. For instance, Borchert et al. composed of Dicer, TRBP, and Argonaute 2 demonstrated polymerase III dependent tran- (Ago2) (13, 15–17). Caudy and Hannon (18) also scription of a human chromosome 19 miR cluster showed presence of Ago2, Drosophila fragile X- (C19MC). miR-515-1, miR-517a, miR-517c and related protein, vasa intronic gene and the micro- miR-519a-1, as well as other miRNAs with coccal nuclease family member Tudor-SN (Dro- upstream tRNA, repeat or transposable elements, sophila CG7008) in the Drosophila RISC structure. Fig. 1. Overview of the microRNA (miRNA) biogenesis pathway. Up to several kb long pri-miRNA is usually transcribed by RNA polymerase II, has a 5#cap and 3#poly(A) tail. Pri-miRNAs fold into stem loop structures that are cleaved by nuclear RNase III Drosha and its RNA-binding partner DGCR8 (DiGeorge syndrome critical region gene 8) into 60–70 nucleotide long pre-miRNA, leaving a two-nucleotide 3# overhang which is then transported to the nucleus through exportin 5, Ran- GTP. Cytoplasmic RNase III, Dicer, and its RNA-binding partner TRBP cleave the pre-miRNA into a 20–25 nucleotide mature miRNA duplex. The double-stranded RNA then assembles into the RNA-induced silencing complex (RISC). One of the strands is degraded, whereas the strand harboring the mature miRNA and the RISC complex is directed to the target mRNA. At this point, different routes are known to exist for gene silencing; translational repression or mRNA degradation (see text for details). 297 Erson and Petty RISC brings the target mRNA and the mature miR-1224 and miR-1225) in mammals and 16 pri- miRNA strand (also known as the Ôguide’ strand) mate-specific mirtrons have been identified (29). together while causing removal of the Ôpassenger’ miRNAs can also be found in exons of non-coding strand (19). The selection of the guide or the pas- RNAs [e.g. miR-155 is encoded within an exon of senger strands is currently thought to be based on the non-coding RNA known as B-cell integration the thermodynamic characteristics of the strands. cluster] (30, 31). It has been generally accepted that the less stable Relatively less is known about the transcrip- strand becomes the guide strand and is the target tional regulation of intergenic or intronic (if dif- mRNA-binding miRNA while the more stable pas- ferent than the host gene) miRNAs. Some recent senger strand is removed from the complex for deg- data showed that c-myc induced transcription of radation. On the contrary, however, Ro et al. (20) a miRNA cluster, miR-17-92 (32), (33) and p53 provides evidence that both strands of the miRNA induced expression of miR-34 genes (reviewed in duplex (sister strands) can indeed be functional and 34). Not surprisingly, expression of miRNAs is possibly co-accumulate in a tissue-dependent man- also shown to be affected by epigenetic mecha- ner to target different mRNA populations. How- nisms. Saito et al. (35) demonstrated increased ever, the mechanism of how such specific expression of miR-127, along with 16 of 313 other regulation takes place is yet to be examined. human miRNAs examined, after DNA methyla- Interestingly, there seems to be additional fine tion and histone deactylase inhibitor treatments. tuning mechanisms to manage the biogenesis of After treatment, proto-oncogene BCL6, a poten- miRNAs. For example, Lin 28, a developmentally tial target of miR-127, was translationally down- regulated RNA-binding protein, has been recently regulated, reportedly because of the increased shown to selectively block let-7 miRNA process- expression of the miR-127 (also see 36 for a review ing (potentially by binding to the terminal loop of epigenetically regulated miRNAs).