The control of translation and mRNA degradation is an important part of the regulation of gene expression. It is now clear that small RNA molecules are common and effective modulators of gene expression in many eukaryotic cells. These small RNAs that control gene expression can be either endogenous or exogenous micro RNAs (miRNAs) and short interfering RNAs (siRNAs) and can affect mRNA degradation and translation, as well as chromatin structure, thereby having impacts on transcription rates. In this review, we discuss possible mechanisms by which miRNAs control translation and mRNA degradation. An emerging theme is that miRNAs, and siRNAs to some extent, target mRNAs to the general eukaryotic machinery for mRNA degradation and translation control.

Small RNAs, including small interfering RNAs (siRNAs) and microRNAs (miRNAs) can silence target genes through several different effector mechanisms1. Whereas siRNA-directed mRNA cleavage is increasingly understood, the mechanisms by which miRNAs repress protein synthesis are obscure. Recent studies have revealed the existence of specific cytoplasmic foci, referred to herein as processing bodies (P-bodies), which contain untranslated mRNAs and can serve as sites of mRNA degradation2-7. Here we demonstrate that Argonaute proteins -the signature components of the RNA interference (RNAi) effector complex, RISC -localize to mammalian P-bodies. Moreover, reporter mRNAs that are targeted for translational repression by endogenous or exogenous miRNAs become concentrated in P-bodies in a miRNA-dependent manner. These results provide a link between miRNA function and mammalian P-bodies and suggest that translation repression by RISC delivers mRNAs to P-bodies, either as a cause or as a consequence of inhibiting protein synthesis.RNAi was initially characterized as a post-transcriptional gene silencing mechanism in which the experimental introduction of long double-stranded RNAs (dsRNAs) induces sequencespecific destruction of homologous mRNAs (reviewed in ref. 1). RNAi pathways are initiated when dsRNAs are processed by Dicer into siRNAs of 21-26 nucleotides. siRNAs are incorporated into the effector complex RISC. In RISC, the siRNA is bound by an Argonaute protein, which uses the sequence of the siRNA to select and cleave complementary substrates (reviewed in ref. 8).RISC can also silence gene expression by preventing protein synthesis. Genetic studies of Caenorhabditis elegans that are mutant for Dicer forged the initial link between a previously known class of small regulatory RNAs, the stRNAs, and the RNAi pathway9-13. Subsequent studies showed that stRNAs are archetypes of a large class of regulatory RNAs, known as miRNAs (reviewed in ref. 14). Although miRNA and siRNA pathways can be biochemically compartmentalized, both types of RNAs enter RISC, bind to Argonaute proteins and identify their silencing targets in conceptually similar ways. They differ, at least in animals, in that miRNAs most often pair imperfectly with their targets and are thus unable to direct Argonautemediated cleavage15. Instead, miRNAs repress protein synthesis in a cleavage-independent fashion8,14,15.3 Correspondence should be addressed to G.J.H. (e-mail: hannon@cshl.org) and R.P. (e-mail: rrparker@u.arizona.edu).Note added in proof: After this study had been accepted for publication, Sen and Blau30 reported similar observations of the localization of human Ago2 to cytoplasmic P-bodies.Note: Supplementary Information is available on the Nature Cell Biology website. The mechanism by which miRNAs repress translation of their target mRNAs is unknown. Conceivably, RISC could prevent protein synthesis from miRNA targets in one of several ways. RISC could affect translation, per se, by altering rates of initiation or elongation by the riboso...

Recent experiments have defined cytoplasmic foci, referred to as processing bodies (P-bodies), wherein mRNA decay factors are concentrated and where mRNA decay can occur. However, the physical nature of P-bodies, their relationship to translation, and possible roles of P-bodies in cellular responses remain unclear. We describe four properties of yeast P-bodies that indicate that P-bodies are dynamic structures that contain nontranslating mRNAs and function during cellular responses to stress. First, in vivo and in vitro analysis indicates that P-bodies are dependent on RNA for their formation. Second, the number and size of P-bodies vary in response to glucose deprivation, osmotic stress, exposure to ultraviolet light, and the stage of cell growth. Third, P-bodies vary with the status of the cellular translation machinery. Inhibition of translation initiation by mutations, or cellular stress, results in increased P-bodies. In contrast, inhibition of translation elongation, thereby trapping the mRNA in polysomes, leads to dissociation of P-bodies. Fourth, multiple translation factors and ribosomal proteins are lacking from P-bodies. These results suggest additional biological roles of P-bodies in addition to being sites of mRNA degradation.

The major pathways of mRNA turnover in eukaryotes initiate with shortening of the poly(A) tail. We demonstrate by several criteria that CCR4 and CAF1 encode critical components of the major cytoplasmic deadenylase in yeast. First, both Ccr4p and Caf1p are required for normal mRNA deadenylation in vivo. Second, both proteins localize to the cytoplasm. Third, purification of Caf1p copurifies with a Ccr4p-dependent poly(A)-specific exonuclease activity. We also provide evidence that the Pan2p/Pan3p nuclease complex encodes the predominant alternative deadenylase. These results, and previous work on Pan2p/Pan3p, define the mRNA deadenylases in yeast. The strong conservation of Ccr4p, Caf1p, Pan2p, and Pan3p indicates that they will function as deadenylases in other eukaryotes. Interestingly, because Ccr4p and Caf1p interact with transcription factors, these results suggest an unexpected link between mRNA synthesis and turnover.

The major pathways of mRNA turnover in eukaryotic cells are initiated by shortening of the poly(A) tail. Recent work has identified Ccr4p and Pop2p as components of the major cytoplasmic deadenylase in yeast. We now demonstrate that CCR4 encodes the catalytic subunit of the deadenylase and that Pop2p is dispensable for catalysis. In addition, we demonstrate that at least some of the Ccr4p/Pop2p‐associated Not proteins are cytoplasmic, and lesions in some of the NOT genes can lead to defects in mRNA deadenylation rates. The Ccr4p deadenylase is inhibited in vitro by addition of the poly(A) binding protein (Pab1p), suggesting that dissociation of Pab1p from the poly(A) tail may be rate limiting for deadenylation in vivo. In addition, the rapid deadenylation of the COX17 mRNA, which is controlled by a member of the Pumilio family of deadenylation activators Puf3p, requires an active Ccr4p/Pop2p/Not deadenylase. These results define the Ccr4p/Pop2p/Not complex as the cytoplasmic deadenylase in yeast and identify positive and negative regulators of this enzyme complex.

No abstract

No abstract