From factors to mechanisms: translation and translational control in eukaryotes Thomas Preiss* and Matthias W Hentze

Transcription

1 515 From factors to mechanisms: translation and translational control in eukaryotes Thomas Preiss* and Matthias W Hentze Biochemical and genetic studies are revealing a network of interactions between eukaryotic translation initiation factors, further refining or redefining perceptions of their function. The notion of translated mrna as a closed-loop has gained support from the identification of physical and functional interactions between the two mrna ends and their associated factors. Translational control mechanisms are beginning to unravel in sufficient detail to pinpoint the affected step in the initiation pathway. Addresses Gene Expression Programme, European Molecular Biology Laboratory, Meyerhofstrasse 1, D Heidelberg, Germany * preiss@embl-heidelberg.de hentze@embl-heidelberg.de Current Opinion in Genetics & Development 1999, 9: X/99/$ see front matter 1999 Elsevier Science Ltd. All rights reserved. Abbreviations ACE adenylation control element BRE Bruno response element CPE cytoplasmic polyadenylation element CPEB CPE-binding protein eif4f eukaryotic initiation factor 4F IRE iron-responsive element IRP iron regulatory protein PABP poly(a)-binding protein PAIP PABP-interacting protein SLBP stem-loop binding protein TIF translation initiation factor tpa tissue-type plasminogen activator UTR untranslated region XFGFR1 Xenopus FGF receptor-1 Introduction The translation of eukaryotic mrnas is a highly competitive and tightly regulated step in gene expression. Control is most commonly exerted on it at the rate-limiting initiation phase. Factors involved in translation initiation have been known for some time and their biochemical activities were used to build the salient model for cap-dependent initiation of translation [1]. According to this model, the 5 cap structure (m 7 GpppN) attracts the eukaryotic initiation factor 4F (eif4f) complex to the mrna (Figure 1). eif4f is a heteromultimeric complex composed of the cap-binding protein eif4e, the RNAdependent ATPase eif4a, and the modular factor eif4g. The small (40S) ribosomal subunit binds to the 5 end of an mrna as a 43S complex including eif3, a multisubunit factor, and the ternary complex of eif2 with GTP and Met-tRNA i. eif4g can contact eif3 whereas eif4a, stimulated by eif4b, is thought to unwind secondary structure in the 5 UTR. The resulting 48S complex then advances through the initiation cycle. A lateral movement of the 43S complex along the mrna, termed scanning, is the most plausible explanation for a faithful recognition of the (usually) first AUG triplet as the start codon. Codon anticodon base-pairing with MettRNA i triggers eif2-bound GTP hydrolysis, catalysed by eif5; it has been thought that this causes dissociation of initiation factors and large (60S) subunit joining to form the 80S ribosome. The scope of our review is to summarise the recent progress in establishing a network of interactions between initiation factors and the mrna substrate. We discuss physiological examples of specific translational control with an emphasis on the interface between regulatory mechanisms and the initiation pathway. As space is limiting, the focus of this review could not be extended to include interesting developments in the area of signal transduction to translation initiation factors or progress in viral mrna translation. Initiation factor function Circularisation of mrna The 3 poly(a) tail of mrnas also participates in translation initiation [2] and acts synergistically with the cap structure [3]. In the mid nineties, a picture of the underlying mechanism began to emerge: an interaction between the poly(a)-binding protein Pab1p and both eif4g homologues was discovered in yeast [4,5]. It was shown that this interaction is required for poly(a)-dependent recruitment of 43S complexes to the mrna [5 7], leading to the hypothesis that simultaneous binding of eif4e and Pab1p to eif4g and concomitant circularisation of the mrna template may form the biochemical basis of translational synergy between cap structure and poly(a) tail (Figure 1a). Functional and physical evidence for such a circular conformation of the mrna has now been obtained. In a yeast cell-free translation system, poly(a)-tail-promoted translation lacks an inherent directionality towards the mrna 5 end and can occur internally. The cap structure serves to anchor ribosome recruitment to the 5 end, probably via Pab1p/eIF4G/eIF4E interactions. Translational synergy between the mrna end modifications originates from a superior efficiency of this combined mode to attract initiation complexes, which becomes apparent when mrnas are competing for limiting initiation factors as is usually the case [8,9 ]. Using recombinant yeast factors eif4e, eif4g and Pab1p and a synthetic capped and polyadenylated RNA, circular complexes were directly observed by atomic force microscopy [10 ]. Poly(A)-binding protein (PABP) binds to eif4g in plant and mammalian systems too. Wheat germ PABP interacts with both cognate eif4f isoforms and eif4b, and a binding determinant was found in the amino-terminal region of

2 516 Differentiation and gene regulation Figure 1 Initiation factor interactions on circular mrnas. The mrna substrate is shown with the 5 and 3 ends in close proximity. Translation initiation is divided operationally into three phases: (a) binding of the 40S ribosomal subunit with associated initiation factors near the 5 end, (b) scanning along the 5 UTR, and (c) start codon recognition and 60S subunit joining. A putative recycling of translational components on the circularised mrna is also indicated. The 'enlargements' highlight interactions of initiation factors on or near the 40S ribosomal subunit. Factors shown in green contain RNAbinding motifs and/or have reported RNAbinding properties. Names of canonical initiation factors are given omitting the usual 'eif'-prefix. In (c), the core subunits of eif3 are shown in orange. The nomenclature used here for yeast eif3 subunits is based on their apparent molecular mass by SDS-PAGE (i.e. [25,28]); p110 is synonymous to Tif32p, p93 to Nip1p, p90 to Prt1p, p39 to Tif34p, and p33 to Tif35p. Lines with two arrowheads indicate additional interactions for which available information does not permit the assessment of evolutionary conservation. (See text for further details and references.) iso-eif4g. In a purified system composed of cap analogue, (iso-)eif4f, PABP and polyribo(a), all binding interactions were found to positively reinforce each other [11,12]. The interaction between the mammalian proteins involves a previously unrecognised amino-terminal region of eif4g, as demonstrated by two independent studies. First, eif4g was found by the two hybrid assay to interact with the rotaviral NSP3 protein which itself is associated with the very 3 end of the (nonadenylated) rotaviral mrnas [13 ]. NSP3 and PABP association with eif4f were found to be mutually exclusive in Rotavirus infected cells. Second, the presence of additional sequence at the amino-terminus of the highly similar eif4gii [14], prompted the cloning of additional 5 cdna sequence of eif4gi. Co-precipitation assays in vivo and in vitro demonstrated an interaction of PABP with the amino-terminal extension of both eif4gs [15 ], which maps to the region that binds NSP3. These findings uncover a new viral strategy to usurp the cellular translation machinery: NSP3 evicts PABP from the eif4f complex and acts as its functional replacement specifically for viral mrnas.

3 Translation and translational control Preiss and Hentze 517 RNA circularisation is thus emerging as a central feature of eukaryotic translation initiation. The functional significance of circularisation may be to increase translational fidelity by ensuring that only properly processed and exported mrna molecules are translated [10 ]. Translational synergy could arise as a consequence of cooperative binding events along the path to 43S recruitment. This could, for instance, affect the formation of the eif4f/pabp bridging complex or the binding of the 40S subunit itself. Another interesting and not mutually exclusive possibility is that components of the translation machinery (i.e. the ribosome) may be recycled on the same mrna molecule after termination, aided by the proximity of the termini. This inherently attractive concept divides the translational life of an mrna into the first and subsequent rounds of translation which may differ in their initiation factor requirement. This issue was addressed by employing poliovirus protease 2A as a tool [16 ], which cleaves eif4gi and II carboxy-terminally from the eif4e/pabp binding regions to block translation [17]. Under conditions of near complete cleavage of eif4gi and II, cellular mrnas continued to be translated for hours whereas, for instance, a newly transcribed reporter mrna was very poorly translated [16 ]. This points to a differential requirement for physically intact eif4g during first and subsequent rounds of translation. The 2A-cleavage of eif4g does not separate the eif4e and PABP binding regions and additional interlocking interactions may exist which help to bridge between the mrna ends [10 ] and terminating ribosomes once the first translation complex has been assembled. Interestingly, an eif4g-related protein called PAIP, for PABP-interacting protein, has been discovered which binds eif4a but not eif4e [18 ]. PAIP may provide an additional contact between the poly(a) tail and the initiation machinery. Ribosomal scanning The scanning model for initiation in eukaryotes can explain the general adherence of eukaryotic mrnas to the first start codon rule, and is compatible with pseudocircular mrna substrates. Its popularity notwithstanding, direct evidence for scanning or insight into its mechanism has proven hard to obtain [19]. A reconstitution approach using highly pure or recombinant components and a toe-printing assay (i.e. the arrest of primer extension by reverse transcriptase at the leading edge of mrna-associated complexes), may now begin to shed light onto this question [20 ]. Addition of 40S subunits, ATP, eif2, 3, 4A, 4B, and 4F led to formation of a cap-proximal complex I (leading edge nucleotide from the 5 end), which could not be chased towards the initiator codon. Inclusion of eif1 and 1A, however, led to formation of an authentic 48S complex, centred over the AUG (complex II, leading edge nucleotides 3 of the AUG). eif1 and 1A act synergistically in this assay and, when added to complex I formed in their absence, require a cycle of dissociation/reassociation to assemble into complex II. Thus, although complex I is not a direct precursor of complex II, eif1 and 1A are the first initiation factors intimately linked to the positioning of the small ribosomal subunit at the translation initiation codon (Figure 1b). They might positively affect the processivity of scanning and/or form part of a clamp that closes over the mrnabinding cleft of the 40S subunit [21,22 ]. Interaction networks The yeast SUI (suppressor of initiator codon mutations) mutants which allow initiation at a UUG codon are affected in either eif1, 2, or 5 [23]. eif1 itself was found to interact with the Nip1p subunit of yeast eif3 (known as p110 in human) [22,24 ], which also binds eif5 [25 ]. In turn, eif5 interacts with eif2 [26], thus outlining an arrangement of factors involved in the final step of scanning : the recognition of the initiation codon (Figure 1). Completion of the cdna cloning of all 10 subunits of mammalian eif3 [27] allowed an exhaustive search for identifiable homologues in the yeast genome. Five yeast genes PRT1, TIF32, NIP1, TIF34, and TIF35 were found, and all five encoded proteins are part of a functional eif3 complex [25 ]; thus, they are considered to constitute a conserved core of eif3 (Figure 1c). The composition of yeast eif3 preparations varies with the purification procedure [28,29] and three additional polypeptides encoded by the genes SUI1 (a yeast homologue of eif1), GCD10, and TIF31 have been implicated as either more loosely bound eif3 subunits or as factors interacting with the complex. A complex involving eif3 with its several RNA-binding subunits is an additional candidate for the above mentioned clamp that could embed the mrna in the 40S-binding cleft [19,22 ]. Information on which of the eif3 subunits interact with eif4g is eagerly awaited. eif4e is phosphorylated at a conserved Ser 209 in response to external stimuli such as growth factors, hormones and mitogens, resulting in enhanced cap-binding [30,31]. On the basis of crystal structure data, phosphorylated Ser 209 can form a salt bridge with Lys 159, which may serve as a another type of clamp over the bound mrna [32]. A MAP kinase activated protein kinase, Mnk1 was found independently, by two-hybrid studies [33 ] and by co-precipitation assays in vivo and in vitro [34 ], to bind the carboxyl-terminus of mammalian eif4gi&ii as well as the eif4g-related protein p97 (or NAT-1/DAP-5). eif4g can thus provide a docking site for efficient and specific phosphorylation of eif4e assembled into eif4f, whereas p97, which cannot bind eif4e, could play a regulatory role by sequestering Mnk1. It came as a major surprise that the bacterial initiation factor 2 (IF2) was found to be conserved throughout evolution, with homologues identified in archae, yeasts, mammals, zebrafish, and maize [35,36 ]. S. cerevisiae strains lacking the FUN12 gene, which encodes yeast (y)if2, show a drastic shift from polysomes to inactive

4 518 Differentiation and gene regulation Figure 2 CPE/ACE AAUAAA A(n+x) CH 3 cap 40S 40S 60S 40S Current Opinion in Genetics & Development Iron regulatory proteins Iron regulatory proteins (IRPs) inhibit ferritin translation by binding to specific 5 UTR sites, the iron-responsive elements (IREs). Using a novel procedure to purify initiation complexes, the regulatory mechanism was analysed in the rabbit reticulocyte lysate system. The cap-proximal binding of IRP-1 to the IRE was shown to permit assembly of eif4f and eif4b on the mrna but to impede the association of eif3 and the 40S ribosomal subunit [37 ]. As one might predict, moving the repressor complex further away from the cap allows the recruitment of mammalian 43S complexes to the mrna. After temporary stalling, they then progress linearly through the IRE to the initiator codon [38]. The apparent scanning arrest is probably overcome by active displacement of IRP-1, because the passive dissociation rate is slow. Regarding the mechanism of initiation, these results also indicate that eif4f can bind to the cap without prior association with 40Sbound eif3. The data also indicate that the RNA-helicase activity of eif4a/4b alone does not suffice for IRP-1 displacement [37 ], a property that the 48S complex seems to possess [38]; it is thus uncertain whether eif4a/4b could serve as the sole motor of scanning. mrna masking and cytoplasmic adenylation control. CPE/masking elements in the 3 UTR may affect translation by direct regulation of translation, and/or control of polyadenylation status. Arrows denote processes which may lead to repression (white) and/or activation of translation (black). Cap ribose methylation stimulated by poly(a) elongation is indicated as CH3, and A(n+x) denotes a poly(a) tail undergoing dynamic changes in length. monosomes. Translation extracts prepared from such mutant strains can be rescued by addition of recombinant yif2 [35 ]. Human (h) or archeal (a) IF2 can substitute for yif2 [36 ]. eif2 and yif2 can both stimulate first peptide bond formation in a heterologous model system, but they are not functional isozymes: eif2 and yif2 cannot suppress each other s mutations. hif2 and yif2 depend on intact GTP-binding domains for full activity. Collectively, these findings force a rethinking of the previous model that eif2 is the functional analogue of IF2. They also suggest the existence of a second GTP hydrolysis step in eukaryotic translation initiation. Further study of IF2 function should reveal unexpected parallels between prokaryotic and eukaryotic protein synthesis. Translational regulation Improved understanding of the translation initiation pathway provides a much stronger basis for understanding translational control mechanisms. Translation can be regulated globally by affecting initiation factor function (e.g. through phosphorylation of eif4e, the 4E-binding proteins [31], and eif2α, or by eif4g cleavage [17]). Such global control affects many cellular mrnas and thus total protein synthesis. In addition, translational regulation of specific mrnas has been studied and is generally exerted through cis-acting elements on the mrna. Recent progress in the latter area is discussed below. Developmental control masking and polyadenylation The 3 UTR of mrnas is a common location of cis-acting elements that control their localisation and/or translation, particularly during oogenesis and early embryogenesis. At first glance, involvement of 3 UTR elements in translational regulation seems counter-intuitive. However, this appears in a different light when one considers the role of the poly(a) tail and the importance of 5 /3 end interactions. Two originally quite separate notions of translational control mechanisms in development, mrna masking and cytoplasmic polyadenylation, have begun to converge. Different possible explanations for an interdependence between masking and regulated polyadenylation have been proposed [39]: First, a 3 UTR element represses translation by maintaining a short poly(a) tail; second, polyadenylation is required to inactivate a 3 UTR repressor element/protein, or third, 3 UTR element(s) control polyadenylation and translation independently an elongated tail may then reinforce translational activity (Figure 2). The latter may act through counteracting a default deadenylation pathway that sets in during maturation [40 ] and/or to boost the translational efficiency of the mrna in the competitive cellular environment [8]. Several recent studies described below highlight the interrelation between masking and polyadenylation in different systems but do not (yet) allow one to propose a unifying model of the underlying mechanism(s). Tissue-type plasminogen activator (tpa) mrna is dormant in primary mouse oocytes and has a short poly(a) tail of ~50 adenosines. Meiotic maturation leads to translational activation and cytoplasmic polyadenylation. Both, the initial poly(a) shortening and the subsequent elongation

5 Translation and translational control Preiss and Hentze 519 require a UA-rich element, the ACE or CPE (adenylation control or cytoplasmic polyadenylation element) in the tpa 3 UTR. Cytoplasmic polyadenylation additionally requires the nuclear polyadenylation hexanucleotide [39]. Injection of ACE competitor transcripts into oocytes now shows that the short poly(a) tail in primary oocytes alone is insufficient for dormancy and that a titrateable factor needs to bind to the ACE. A candidate for this masking function is an approx. 80 kda protein which cross-links to the ACE. Removal of the factor, however, only leads to awakening of the tpa mrna in primary oocytes if at least a short poly(a) tail is present. It also induces a more rapid polyadenylation of the mrna during maturation [40 ]. Thus, the findings with tpa mrna are most consistent with the third scenario listed above. The mrna for Xenopus FGF receptor-1 (XFGFR 1) is repressed in immature oocytes as a result of the translation inhibitory motif in its 3 UTR. Oocyte maturation induces polyadenylation and activation of translation. When maturation is induced directly, by injection of maturationpromoting factor, XFGFR translation occurs at normal levels concomitant with only minimal poly(a) tail elongation on the mrna. Treatment of c-mos-depleted oocytes with progesterone, by contrast, does not induce progression through meiosis or XFGFR translation but the mrna gets efficiently polyadenylated [41 ]. Here, masking as well as unmasking can be uncoupled from polyadenylation and unmasking can occur in the presence of a short tail. Translational activation of Xenopus cyclin B1 mrna during oocyte maturation also coincides with CPE-dependent polyadenylation. Injection of excess CPE competitor RNA into immature oocytes causes translational unmasking of the endogenous mrna in the absence of poly(a) tail elongation [42 ], consistent with the third hypothesis above. By contrast, reporter mrnas bearing the cyclin B1 3 UTR with functional CPEs, a tail of 30 adenosines, but without the hexanucleotide polyadenylation motif are not activated during oocyte maturation. This latter result is more suggestive of the second explanation. The CPE-binding protein (CPEB) is essential for cytoplasmic polyadenylation. Additionally, its binding activity correlates strongly with translational repression. These findings raise the possibility that CPEB could function both positively and negatively at different stages of development. Interestingly, an EMCV internal ribosome entry site-driven reporter mrna is much less sensitive to CPE-mediated repression. Internal ribosome entry sitedriven translation, because of differential initiation factor requirement, may not depend on interactions that are necessary for the cellular cyclin B1 transcript. It has also been suggested that polyadenylation may function by stimulating cap-ribose methylation [43]. Recent studies with maternal surf clam mrnas also highlight the interplay between masking and polyadenylation. p82 was originally found to bind a U-rich masking element in the 3 UTR of ribonucleotide reductase mrna and to mediate translational repression in vitro; recently it was identified as clam CPEB [44,45 ]. The ribonucleotide reductase mrna 3 UTR contains six CPE-like motifs, including two in the masking element, which together with the hexanucleotide motif support p82-mediated polyadenylation in egg extract. Fertilisation leads to translational activation of masked mrnas and p82 is phosphorylated by a cdc2-like kinase before being degraded. The onset of spermatogenesis in the Caenorhabditis elegans hermaphrodite depends on translational repression of tra-2 mrna. Tra-2 repression is mediated via two 28 nucleotide direct repeat elements (DREs or TGEs [tra-2 and GLI elements]) in the 3 UTR [46]. A yeast three-hybrid-screen identified GLD-1, a germline-specific member of the STAR-family of proteins, as a specific TGE-binding factor. GLD-1 is a component of the complex that forms on TGEs with worm extracts. Translational repression mediated through the TGEs by GLD-1 was demonstrated in a yeast in vitro system and depends on a functional KHdomain in the recombinant protein. Ectopic expression of GLD-1 and reporter constructs in somatic cells causes specific repression in vivo [47 ]. Intriguingly, both TGE-mediated control and the STAR protein family are evolutionarily conserved. In contrast to most other cellular mrnas, metazoan histone mrnas lack a poly(a) tail. They end with a conserved stem-loop that fulfils many of the functions of the poly(a) tail. It is essential for translation [48], and interacts with SLBP (stem-loop binding protein) in mammalian somatic cells. SLBP participates in pre-mrna processing [49] and is a component of polyribosomal histone mrnps. Xenopus oocytes possess two SLBPs. SLBP-2 is oocyte-specific, present at high levels in early oogenesis and degraded during early embryogenesis. SLBP-1 is the homologue of mammalian SLBP and displays a reciprocal temporal expression pattern [50 ]. This suggests the possibility of a masking phenomenon also for this nonadenylated mrna, namely that exchange of SLBP-2 for -1 on the 3 stem-loop could activate histone mrna translation at oocyte maturation. Developmental control mrna localisation and translation Normal progression through oogenesis and embryogenesis in Drosophila requires coupling between translational control and mrna localisation to achieve proper temporal and spatial protein expression [51]. For instance, translation of maternal oskar mrna is silenced during transport to the posterior pole of the oocyte and until Oskar protein is required. These processes require the 3 UTR of oskar mrna and do not involve noticeable changes in mrna poly(a) tail length. The Bruno protein recognises repeated conserved 3 UTR sequences, the Bruno response elements (BREs). Bruno prevents premature translation and colocalises with oskar mrna [52]. Oskar mrna is

6 520 Differentiation and gene regulation translated as two functional isoforms by alternative start codon usage on its mrna and an element between the two AUGs activates translation from both codons exclusively on localised mrna [53 ]. This region is a derepressor rather than an activator element, because it is only required when the 3 BREs are active. Two proteins, p50 and p68 (distinct from Bruno), interact with the 5 element and p50 also binds to the 3 BRE, apparently simultaneously with Bruno. These observations demonstrate a highly complex mechanism involving a growing list of factors to achieve localisation-dependent derepression of oskar mrna translation. The functional interactions between 5 UTR derepressor and 3 UTR repressor elements again reflects the recurrent theme of end-to-end communication. Conclusions and future directions Research on the translation initiation pathway is progressing towards a more complete picture of molecular interactions during its different phases. In particular, the concept of 5 and 3 end interactions opens up new possibilities to address poly(a)-tail-dependent and poly(a)-tail-independent means of translational regulation by specific mrna-binding proteins in a more informed context. Acknowledgements We thank A Hinnebusch, N Gunkel and the members of the Hentze lab for comments on the manuscript. P Riedinger is acknowledged for help with the figures. References and recommended reading Papers of particular interest, published within the annual period of review, have been highlighted as: of special interest of outstanding interest 1. Merrick WC, Hershey JWB: The pathway and mechanism of eukaryotic protein synthesis. In Translational Control. Edited by Hershey JWB, Mathews MB, Sonenberg N. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press; 1996: Jacobson A: Poly(A) metabolism and translation: the closed-loop model. In Translational Control. Edited by Hershey JWB, Mathews MB, Sonenberg N. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press; 1996: Gallie DR: The cap and poly(a) tail function synergistically to regulate mrna translational efficiency. Genes Dev 1991, 5: Tarun SZ Jr, Sachs AB: Association of the yeast poly(a) tail binding protein with translation initiation factor eif-4g. EMBO J 1996, 15: Tarun SZ Jr, Wells SE, Deardorff JA, Sachs AB: Translation initiation factor eif4g mediates in vitro poly(a) tail-dependent translation. Proc Natl Acad Sci USA 1997, 94: Tarun SZ Jr, Sachs AB: A common function for mrna 5 and 3 ends in translation initiation in yeast. Genes Dev 1995, 9: Kessler SH, Sachs AB: RNA recognition motif 2 of yeast Pab1p is required for its functional interaction with eukaryotic translation initiation factor 4G. Mol Cell Biol 1998, 18: Preiss T, Muckenthaler M, Hentze MW: Poly(A)-tail-promoted translation in yeast: implications for translational control. RNA 1998, 4: Preiss T, Hentze MW: Dual function of the messenger RNA cap structure in poly(a)-tail-promoted translation in yeast. Nature 1998, 392: We provide evidence for a functional cooperativity between the cap and poly(a) tail affecting both quality and quantity of translation products. Translational synergy is explained as a superior efficiency of jointly promoted initiation by the cap structure and the poly(a) tail under competitive conditions. 10. Wells SE, Hillner PE, Vale RD, Sachs AB: Circularization of mrna by eukaryotic translation initiation factors. Mol Cell 1998, 2: Direct demonstration by atomic force microscopy that the eif4e/4g/pab1p interaction forms the core of a bridge between the mrna cap structure and poly(a) tail, and hence can mediate mrna circularisation. 11. Le H, Tanguay RL, Balasta ML, Wei CC, Browning KS, Metz AM, Goss DJ, Gallie DR: Translation initiation factors eif-iso4g and eif-4b interact with the poly(a)-binding protein and increase its RNA binding activity. J Biol Chem 1997, 272: Wei CC, Balasta ML, Ren J, Goss DJ: Wheat germ poly(a) binding protein enhances the binding affinity of eukaryotic initiation factor 4F and (iso)4f for cap analogues. Biochemistry 1998, 37: Piron M, Vende P, Cohen J, Poncet D: Rotavirus RNA-binding protein NSP3 interacts with eif4gi and evicts the poly(a) binding protein from eif4f. EMBO J 1998, 17: Discovery of a new viral strategy to reprogram cellular translation: NSP3 protein binds specifically to rotaviral mrna 3 ends and displaces PABP from eif4f. Evidence is provided for a PABP/eIF4G interaction in mammalian cells. 14. Gradi A, Imataka H, Svitkin YV, Rom E, Raught B, Morino S, Sonenberg N: A novel functional human eukaryotic translation initiation factor 4G. Mol Cell Biol 1998, 18: Imataka H, Gradi A, Sonenberg N: A newly identified N-terminal amino acid sequence of human eif4g binds poly(a)-binding protein and functions in poly(a)-dependent translation. EMBO J 1998, 17: Co-immunoprecipitation analysis identified a 29 amino acid stretch at the amino terminus of human eif4g I and II as the PABP-binding site. Thus, together with [13 ], this paper extends the concept of the eif4e/4g/pab1p bridging interactions to mammalian translation. 16. Novoa I, Carrasco L: Cleavage of eukaryotic translation initiation factor 4G by exogenously added hybrid proteins containing poliovirus 2Apro in HeLa cells: effects on gene expression. Mol Cell Biol 1999, 19: Possible differences between the first and subsequent rounds of translation initiation on the same mrna molecule are so far largely unexplored. This study takes a first step in this direction by showing a differential requirement for physically intact eif4g. 17. Gradi A, Svitkin YV, Imataka H, Sonenberg N: Proteolysis of human eukaryotic translation initiation factor eif4gii, but not eif4gi, coincides with the shutoff of host protein synthesis after poliovirus infection. Proc Natl Acad Sci USA 1998, 95: Craig AW, Haghighat A, Yu AT, Sonenberg N: Interaction of polyadenylate-binding protein with the eif4g homologue PAIP enhances translation. Nature 1998, 392: This paper describes PAIP, a new human protein with homology to the central portion of eif4g. This adds complexity to mammalian translation initiation models: PAIP interacts with PABP and eif4a, but not with eif4e. 19. Jackson RJ: A comparative view of initiation site selection mechanisms. In Translational Control. Edited by Hershey JWB, Mathews MB, Sonenberg N. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press; 1996: Pestova TV, Borukhov SI, Hellen CU: Eukaryotic ribosomes require initiation factors 1 and 1A to locate initiation codons. Nature 1998, 394: Initiation reactions assembled from highly purified or recombinant components are combined elegantly with toe-printing analysis. This demonstrates that the 'Cinderella' factors eif1/1a are essential initiation factors that are absolutely required for positioning of 43S complexes at the start codon in vitro 21. Pestova TV, Hellen CU: Ribosome recruitment and scanning: what s new? Trends Biochem Sci 1999, 24: Fletcher CM, Pestova TV, Hellen CU, Wagner G: Structure and interactions of the translation initiation factor eif1. EMBO J 1999, 18: The solution structure of eif1 was determined by NMR spectroscopy. Availability of structural information for this interesting factor (see [20 ]) will help us understand its mode of action 23. Huang HK, Yoon H, Hannig EM, Donahue TF: GTP hydrolysis controls stringent selection of the AUG start codon during translation initiation in Saccharomyces cerevisiae. Genes Dev 1997, 11:

7 Translation and translational control Preiss and Hentze Asano K, Phan L, Anderson J, Hinnebusch AG: Complex formation by all five homologues of mammalian translation initiation factor 3 subunits from yeast Saccharomyces cerevisiae. J Biol Chem 1998, 273: See annotation [25 ]. 25. Phan L, Zhang X, Asano K, Anderson J, Vornlocher HP, Greenberg JR, Qin J, Hinnebusch AG: Identification of a translation initiation factor 3 (eif3) core complex, conserved in yeast and mammals, that interacts with eif5. Mol Cell Biol 1998, 18: Together with [24 ], this study identifies an eif3 core complex conserved between yeast and mammals and charts a map of intersubunit interactions and contacts with other initiation factors. 26. Das S, Maiti T, Das K, Maitra U: Specific interaction of eukaryotic translation initiation factor 5 (eif5) with the beta-subunit of eif2. J Biol Chem 1997, 272: Block KL, Vornlocher HP, Hershey JW: Characterization of cdnas encoding the p44 and p35 subunits of human translation initiation factor eif3. J Biol Chem 1998, 273: Naranda T, MacMillan SE, Hershey JW: Purified yeast translational initiation factor eif-3 is an RNA-binding protein complex that contains the PRT1 protein. J Biol Chem 1994, 269: Danaie P, Wittmer B, Altmann M, Trachsel H: Isolation of a protein complex containing translation initiation factor Prt1 from Saccharomyces cerevisiae. J Biol Chem 1995, 270: Wang X, Flynn A, Waskiewicz AJ, Webb BL, Vries RG, Baines IA, Cooper JA, Proud CG: The phosphorylation of eukaryotic initiation factor eif4e in response to phorbol esters, cell stresses, and cytokines is mediated by distinct MAP kinase pathways. J Biol Chem 1998, 273: Sonenberg N, Gingras AC: The mrna 5 cap-binding protein eif4e and control of cell growth. Curr Opin Cell Biol 1998, 10: Marcotrigiano J, Gingras AC, Sonenberg N, Burley SK: Cocrystal structure of the messenger RNA 5 cap-binding protein (eif4e) bound to 7-methyl-GDP. Cell 1997, 89: Waskiewicz AJ, Johnson JC, Penn B, Mahalingam M, Kimball SR, Cooper JA: Phosphorylation of the cap-binding protein eukaryotic translation initiation factor 4E by protein kinase Mnk1 in vivo. Mol Cell Biol 1999, 19: See annotation [34 ]. 34. Pyronnet S, Imataka H, Gingras AC, Fukunaga R, Hunter T, Sonenberg N: Human eukaryotic translation initiation factor 4G (eif4g) recruits Mnk1 to phosphorylate eif4e. EMBO J 1999, 18: This paper and [33 ] identify a physical link between mitogen- and stressactivated signalling and translation initiation. Mnk1 docks onto eif4g to phosphorylate bound eif4e. 35. Choi SK, Lee JH, Zoll WL, Merrick WC, Dever TE: Promotion of met Met-tRNA Met i binding to ribosomes by yif2, a bacterial IF2 homolog in yeast. Science 1998, 280: In addition to highlighting an unexpected universal conservation of a bacterial and a yeast initiation factor, this report and [36 ]. indicate the existence of an additional GTP-hydrolysis step in eukaryotic initiation. 36. Lee JH, Choi SK, Roll-Mecak A, Burley SK, Dever TE: Universal conservation in translation initiation revealed by human and archaeal homologs of bacterial translation initiation factor IF2. Proc Natl Acad Sci USA 1999, 96: See annotation [35 ]. 37. Muckenthaler M, Gray NK, Hentze MW: IRP-1 binding to ferritin mrna prevents the recruitment of the small ribosomal subunit by the cap-binding complex eif4f. Mol Cell 1998, 2: This paper describes a new method to specifically capture and purify translation initiation complexes formed on a given mrna. Analysis of the composition of such complexes reveals the mechanism of translational control by a 5 UTR-binding protein at 'initiation factor resolution'. 38. Paraskeva E, Gray NK, Schlager B, Wehr K, Hentze MW: Ribosomal pausing and scanning arrest as mechanisms of translational regulation from cap-distal iron-responsive elements. Mol Cell Biol 1999, 19: Wickens M, Kimble J, Strickland S: Translational control of developmental decisions. In Translational Control. Edited by Hershey JWB, Mathews MB, Sonenberg N. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press; 1996: Stutz A, Conne B, Huarte J, Gubler P, Volkel V, Flandin P, Vassalli JD: Masking, unmasking, and regulated polyadenylation cooperate in the translational control of a dormant mrna in mouse oocytes. Genes Dev 1998, 12: The complex interdependence between translational masking and control of cytoplasmic polyadenylation is elegantly demonstrated for tissue-type plasminogen activator mrna. 41. Culp PA, Musci TJ: Translational activation and cytoplasmic polyadenylation of FGF receptor-1 are independently regulated during Xenopus oocyte maturation. Dev Biol 1998, 193: The role of a 3 UTR motif in the appropriate timing of Xenopus FGF-receptor 1 translation during meiotic maturation is highlighted. The study provides an example of an apparent lack of a relationship between translational activation and polyadenylation. 42. de Moor CH, Richter JD: Cytoplasmic polyadenylation elements mediate masking and unmasking of cyclin B1 mrna. EMBO J 1999, 18: Here, an involvement of 3 UTR CPEs in both translational masking and unmasking of cyclin B1 mrna during Xenopus oocyte maturation is demonstrated. 43. Kuge H, Richter JD: Cytoplasmic 3 poly(a) addition induces 5 cap ribose methylation: implications for translational control of maternal mrna. EMBO J 1995, 14: Walker J, Minshall N, Hake L, Richter J, Standart N: The clam 3 UTR masking element-binding protein p82 is a member of the CPEB family. RNA 1999, 5: See annotation [45 ]. 45. Minshall N, Walker J, Dale M, Standart N: Dual roles of p82, the clam CPEB homolog, in cytoplasmic polyadenylation and translational masking. RNA1999, 5: This paper and [44 ] illustrate a functional linkage between masking/unmasking and cytoplasmic polyadenylation. 46. Jan E, Yoon JW, Walterhouse D, Iannaccone P, Goodwin EB: Conservation of the C. elegans tra-2 3 UTR translational control. EMBO J 1997, 16: Jan E, Motzny CK, Graves LE, Goodwin EB: The STAR protein, GLD 1, is a translational regulator of sexual identity in Caenorhabditis elegans. EMBO J 1999, 18: Translational repression through 3 UTR elements is mediated by the KHdomain protein GLD-1, a member of the STAR- family of proteins. A comprehensive study providing both in vivo and biochemical evidences. 48. Gallie DR, Lewis NJ, Marzluff WF: The histone 3 -terminal stemloop is necessary for translation in Chinese hamster ovary cells. Nucleic Acids Res 1996, 24: Martin F, Schaller A, Eglite S, Schumperli D, Muller B: The gene for histone RNA hairpin binding protein is located on human chromosome 4 and encodes a novel type of RNA binding protein. EMBO J 1997, 16: Wang ZF, Ingledue TC, Dominski Z, Sanchez R, Marzluff WF: Two Xenopus proteins that bind the 3 end of histone mrna: implications for translational control of histone synthesis during oogenesis. Mol Cell Biol 1999, 19: This study demonstrates that Xenopus oocytes have two SLBPs whose levels are inversely regulated during oogenesis. The authors suggest that an exchange of SLBP isoforms on the histone mrna 3 ends regulates their translational activation. 51. Macdonald PM, Smibert CA: Translational regulation of maternal mrnas. Curr Opin Genet Dev 1996, 6: Kim-Ha J, Kerr K, Macdonald PM: Translational regulation of oskar mrna by bruno, an ovarian RNA-binding protein, is essential. Cell 1995, 81: Gunkel N, Yano T, Markussen FH, Olsen LC, Ephrussi A: Localization-dependent translation requires a functional interaction between the 5 and 3 ends of oskar mrna. Genes Dev 1998, 12: derepressor and 3 UTR repressor elements cooperate to achieve spatially restricted oskar mrna translation during Drosophila oogenesis. This is the first detailed report that identifies an active role of a 5 element in mediating derepression from 3 UTR silencing.