The most hotly pursued class of non-coding RNA (ncRNA) in recent years has been the miRNAs. Although research still continues on miRNA discovery, the field has largely made the transition to studying the endogenous biological functions of miRNAs. In particular, speakers discussed computational or experimental approaches to identifying the specific targets of miRNA regulation, as well as genetic approaches to identifying mutant phenotypes associated with altered miRNA activity.
David Bartel (Whitehead Institute, Massachusetts Institute of Technology, Cambridge, USA) presented TargetScanS, his group's animal miRNA-target-finding algorithm. It predicts potential targets of miRNAs by searching for 3' untranslated region (UTR) sequences with highly and specifically conserved Watson-Crick matches to positions 2-7 or 2-8 (the 'seed') of the query miRNA; increased confidence is assigned to target sites with an adenosine at position 1 of the miRNA interaction site. Bartel predicted that at least one third of the genes in the human genome are miRNA targets. Lee Lim (Rosetta Inpharmatics, Seattle, USA) challenged the dogma that the main function of animal miRNAs is to inhibit productive translation without causing mRNA degradation. Lim found that miR-1 and miR-124 in humans can indeed downregulate the transcript levels of approximately 100-200 genes via 3' UTR seed matches. He speculated that mir-1 and mir-124, which are restricted to muscle/heart and the nervous system, respectively, may have rather global effects on gene expression that help maintain the identity of these tissues.
Although many animal miRNA targets are regulated by pairing of the miRNA to the 3' UTR of the target mRNA, Tom Tuschl (Rockefeller University, New York, USA) showed that several virus-encoded miRNAs are completely antisense to viral mRNAs and are likely to be targeting them for degradation. But miRNAs from any individual virus exhibit little sequence similarity to miRNAs from animals, plants, or other viruses, leading Tuschl to postulate that viral miRNAs may have independently evolved multiple times.
Jim Carrington (Oregon State University, Corvallis, USA) added another twist to the story by describing an unexpectedly complex regulatory pathway involving small RNAs in plants. Certain plant miRNAs guide the cleavage of particular ncRNAs, thereby defining new 5' ends to these transcripts. The miRNA-defined 5' end sets the frame for processive cleavage of the (presumably double-stranded) transcript at 21-nucleotide intervals by the RNase Dicer, which generates multiple siRNA molecules. At least some of these function as trans-acting siRNAs that are complementary to, and mediate cleavage of, mRNA targets different from those of the original miRNAs.
The first miRNA discovered, lin-4 in Caenorhabditis elegans, was found by virtue of the severe cell-lineage defect in mutant nematodes. Victor Ambros (Dartmouth College, Hanover, USA), whose lab discovered the lin-4 RNA, continued the discussion of the biological roles of miRNAs by describing several worm and fly miRNA knockouts. He suggested that many animal miRNAs may work redundantly in genetic circuits, as only the triple knockout of the let-7-related miRNAs mir-48, mir-84 and mir-241 revealed a heterochronic cell-lineage phenotype (in which the normal timing of developmental events is perturbed). Ambros also described how expression of mir-1 in mesodermal derivatives has been conserved between invertebrates and vertebrates, and that Drosophila lacking mir-1 display incredible defects in muscle morphogenesis, such as alterations in myoblast fusion and distortions in muscle morphology. Ronald Plasterk (Hubrecht Laboratory, Utrecht, The Netherlands) described the use of locked nucleic acid (LNA) probes to reveal beautiful and diverse expression patterns of zebrafish miRNAs in developing tissues and differentiated organs. This technique has great potential to guide biological studies of miRNA function by indicating tissue-specific sites of regulation by individual miRNAs.