PubMed Health⌕ Search

SEARCH · PubMed Health

Results for “Translating”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 559 records · Page 31Linked to original sources

Nuclear translation: what is the evidence?

Recently, several reports have been published in support of the idea that protein synthesis occurs in both the nucleus and the cytoplasm. This proposal has generated a great deal of excitement because, if true, it would mean that our thinking about the compartmentalization of cell functions would have to be re-evaluated. The significance and broad implications of this phenomenon require that the experimental evidence used to support it be carefully evaluated. Here, we critique the published evidence in support of, or in opposition to, the question of whether translation occurs in the nucleus. Arguments in support of nuclear translation focus on three issues: (1) the presence of translation factors and ribosomal components in the nucleus, and their recruitment to sites of transcription; (2) amino acid incorporation in isolated nuclei and in nuclei under conditions that should not permit protein import; and (3) the fact that nuclear translation would account for observations that are otherwise difficult to explain. Arguments against nuclear translation emphasize the absence (or low abundance) from nuclei of many translation factors; the likely inactivity of nascent ribosomes; and the loss of translation activity as nuclei are purified from contaminating cytoplasm. In our opinion, all of the experiments on nuclear translation published to date lack critical controls and, therefore, are not compelling; also, traditional mechanisms can explain the observations for which nuclear translation has been invoked. Thus, while we cannot rule out nuclear translation, in the absence of better supporting data we are reluctant to believe it occurs.

Cell Nucleus↗

A 250-nucleotide UA-rich element in the 3' untranslated region of Xenopus laevis Vg1 mRNA represses translation both in vivo and in vitro.

Xenopus laevis Vgl mRNA undergoes both localization and translational control during oogenesis. Vg1 protein does not appear until late stage IV, after localization is complete. To determine whether Vg1 translation is regulated by cytoplasmic polyadenylation, the RACE-PAT method was used. Vg1 mRNA has a constant poly(A) tail throughout oogenesis, precluding a role for cytoplasmic polyadenylation. To identify cis-acting elements involved in Vg1 translational control, the Vg1 3' UTR was inserted downstream of the luciferase ORF and in vitro transcribed, adenylated mRNA injected into stage III or stage VI oocytes. The Vg1 3' UTR repressed luciferase translation in both stages. Deletion analysis of the Vg1 3' UTR revealed that a 250-nt UA-rich fragment, the Vg1 translational element or VTE, which lies 118 nt downstream of the Vg1 localization element, could repress translation as well as the full-length Vg1 3' UTR. Poly(A)-dependent translation is not necessary for repression as nonadenylated mRNAs are also repressed, but cap-dependent translation is required as introduction of the classical swine fever virus IRES upstream of the luciferase coding region prevents repression by the VTE. Repression by the Vg1 3' UTR has been reproduced in Xenopus oocyte in vitro translation extracts, which show a 10-25-fold synergy between the cap and poly(A) tail. A number of proteins UV crosslink to the VTE including FRGY2 and proteins of 36, 42, 45, and 60 kDa. The abundance of p42, p45, and p60 is strikingly higher in stages I-III than in later stages, consistent with a possible role for these proteins in Vg1 translational control.

3' Untranslated Regions↗

Cap-dependent translation and control of the cell cycle.

The control of gene expression at the translational level has emerged in the past decade as an important aspect of cell growth, proliferation and malignant transformation. Translation is primarily regulated at the initiation step, and mitogen-dependent signaling pathways converge to modulate the activity of translation initiation factors. In most tumors tested, at least one translation initiation factor is overexpressed and overexpression of translation initiation factors often provokes transformation. Malignant transformation could be caused by the increased translation of a subset of mRNAs encoding important proteins which are required for cell growth and proliferation. These mRNAs usually possess regulatory sequences that render their translation more sensitive to changes in the activity of translation initiation factors. In this chapter, we describe recent advances illustrating the importance of translation in cell cycle progression and cell transformation. Control of translation initiation may represent an excellent target for antitumor drugs.

Animals↗

[Translational regulation of potato virus X RNA-coat protein complexes: the key role of a coat protein N-terminal peptide].

The efficiency of in vitro translation of potato virus X (PVX) RNA within vRNP complexes assembled from genomic RNA and viral CP was examined. The vRNP particles contain the 5'-proximal RNA segments encapsidated by helically arranged CP head-like portions heterogeneous in length and the CP-free RNA tail. Translation of RNA is completely repressed upon incubation with PVX CP and is accompanied by vRNP particles production. By contrast, translation is activated in vRNPs in vitro assembled using two CP forms, differing in the principals of their N-terminal peptides modification. The N-terminal peptide of PVX CP represents the major phosphorylation site(s) for Thr/Ser-specific protein kinases. It was shown that: (i) CP phosphorylation results in a translational activation of vRNP; (ii) removal of N-terminal peptide from CP abolished activation and CP retains the translation repressing ability. It was suggested that substitution of Ser/Thr residues by non-phosphorylated Ala/Gly in N-terminal peptide of the mutant CP will led to a complete inhibition of vRNP translation. However, opposite results were obtained in our experiments: (i) RNA of such mutant virus (PVX-ST) was efficiently translated within the virions; (ii) RNA of a wild-type (wt) PVX also efficiently translated in mixedly assembled vRNP "wt PVX RNA + PVX-ST CP"; (iii) opposite result (repression of translation) was obtained with "mixed" vRNP (PVX-ST RNA + wtPVX CP). Therefore, the N-terminal peptide located at the surface of the particle and of the particles plays a key role in translation activation of the RNA encapsidated in vRNP and native virions.

Capsid Proteins↗

Translational control of globin chain ontogeny in hamster yolk sac erythroid cells.

Prior research has demonstrated that globin ontogeny of hamster proceeds nearly to completion during the several days that yolk sac erythroid cells (YSEC) circulate in the embryo; synthesis of embryonic globin chains gives way to synthesis of adult globin chains in these primitive cells. In the present study, we translated total cell RNA extracted from YSEC on days 9-13 of gestation in wheat germ cell-free extract, expecting to observe the same progressive rise that occurs in vivo in rates of translation of alpha- and beta-globin mRNA during ontogeny. The opposite occurred; translation rates of both globins decreased sharply. This disparity between synthesis of alpha- and beta-globins in vivo and in vitro suggested an element of control of translation attributable to the YSEC cytoplasm. We therefore assayed the effect of RNA-free clarified YSEC cytoplasm on cell-free translation of YSEC RNA. A repression of translation was detected of alpha- and beta-globin mRNA (not of embryonic globin mRNA), exercised strongly by cytoplasm from YSEC early in ontogeny (gestational day 9), and weakening as ontogeny progressed. The same effect was noted on alpha- and beta-globin mRNA of adult hamster and of rabbit. Heat treatment of cytoplasm abolished the greater part of the translation regulation, suggesting that the active agent is protein. Further characterization of this translational regulator included: (a) it binds to globin poly(A) mRNA but not to poly(A), (b) it was not detected in cell lysate of adult hamster brain, lung, or erythrocytes, and (c) it did not inhibit translation of adult hamster brain and liver RNA. We conclude that hamster globin ontogeny is substantially modulated by this translational regulation of alpha- and beta-globin expression.

Animals↗

Eukaryotic initiation factor (eIF)-4F. Implications for a role in internal initiation of translation.

In order to study the eukaryotic translation initiation mechanisms of "internal initiation," "re-initiation," and/or "coupled internal initiation," a series of model mRNAs have been constructed which contain two non-overlapping open reading frames (ORFs) that encode different lengths of rabbit alpha globin. These mRNAs, along with the bicistronic constructs TK/CAT and TK/P2CAT developed by Pelletier and Sonenberg (Pelletier, J., and Sonenberg, N. (1988) Nature 334, 320-325, 1988), were used to program an in vitro rabbit reticulocyte lysate translation system. Cap-dependent and cap-independent translation were distinguished by monitoring translation in the presence or absence of exogenously added cap analog (m7GTP). Messenger RNAs which translate both ORF1 and ORF2 by a cap-dependent mechanism, as well as mRNAs that translate ORF2 by a cap-independent mechanism while still translating ORF1 in a cap-dependent fashion have been obtained. These same alpha globin mRNAs differ by no more than 45 nucleotides in intercistronic length. Initiation factor addition studies were performed in this same in vitro translation system. Both eukaryotic initiation factor (eIF)-4F and, to a lesser extent, eIF-4B can stimulate translation of an internally located ORF independent of upstream ORF translation and in a manner not dependent on mRNA cap recognition. This indicates that the cap-recognition initiation factor, eIF-4F, and eIF-4B facilitate cap-independent and internal initiation of an open reading frame.

Animals↗

The effect of Escherichia coli ribosomal protein S1 on the translational specificity of bacterial ribosomes.

Ribosomes from Gram-negative bacteria such as Escherichia coli exhibit non-specific translation of bacterial mRNAs. That is, they are able to translate mRNAs from a variety of sources in a manner independent of the "strength" of the Shine-Dalgarno region, in contrast to ribosomes from many Gram-positive bacteria, such as Bacillus subtilis, which show specific translation in only being able to translate other Gram-positive mRNA, or mRNAs that have "strong" Shine-Dalgarno regions. There is an evolutionary correlation between the translational specificity and the absence of a protein analogous to E. coli ribosomal protein S1. The specificity observed with B. subtilis ribosomes is a function of their 30 S subunit which lacks S1; translation of Gram-negative mRNA can occur with heterologous ribosomes containing the 30 S subunit of E. coli ribosomes and the 50 S subunit of B. subtilis ribosomes. However, the addition of E. coli S1 alone to B. subtilis ribosome does not overcome their characteristic inability to translate mRNA from Gram-negative organisms. By contrast, the removal of S1 from E. coli ribosomes results in translational behavior similar to that shown by B. subtilis ribosomes in that the S1-depleted E. coli ribosomes can translate mRNA from Gram-positive sources in the absence of added S1, although addition of S1 stimulates further translation of such mRNAs by the E. coli ribosomes.

Bacteria↗

Cell-free translation of adenovirus 2 E1a- and E1b-specific mRNAs and evidence that E1a-related polypeptides are produced from E1a-E1b overlapping mRNA.

We have characterized the polypeptides translated in vitro by mRNAs of early region 1 (E1) of human adenovirus (Ad) type 2. Poly (A+) polyribosomal RNA was isolated from early Ad2-infected cells, the viral specific mRNAs were selected by hybridization to Ad2 E1a and E1b DNA, and the mRNAs were translated in vitro using [35S]methionine as a labeled precursor with a rabbit reticulocyte lysate. E1a-selected mRNA was translated to the 45-58-kDa cluster of polypeptides. We show here that E1b-selected mRNA can also be translated to the 45-58-kDa cluster of polypeptides in addition to the major 19-kDa polypeptide. The E1b 58-kDa polypeptide was produced only at a low level unless E1b mRNA is fractionated before translation to enrich for the 58-kDa mRNA. Translation of E1b region-selected mRNAs that have been fractionated by size shows that the 22 S mRNA fraction is translated to at least the 53-58-kDa E1a-related polypeptides as well as to E1b 58- and 19-kDa polypeptides. Our experiments suggest that the 22 S mRNA fraction includes E1a-E1b overlapping mRNA which was translated to E1a-related polypeptides as well as E1b 22 S mRNA. When compared by two-dimensional gel electrophoresis and by tryptic peptide mapping, the cluster of polypeptides translated from E1a-selected mRNA and the cluster translated from E1b-selected mRNA were distinguishable. A possible explanation for this is discussed, based upon splicing sites of the E1a-E1b overlapping mRNA which would result in an amino acid sequence with a COOH-terminal end slightly different from that of E1a polypeptides.

Adenoviruses, Human↗

Translational regulation of lipoprotein lipase by thyroid hormone is via a cytoplasmic repressor that interacts with the 3' untranslated region.

To better characterize the increase in lipoprotein lipase (LPL) translation by hypothyroidism, adipocytes were prepared from control and hypothyroid rats. Whereas LPL synthesis was higher in hypothyroid adipocytes, with no change in mRNA levels, there was no increase in hormone-sensitive lipase (HSL) synthesis. To determine whether a transacting translation regulatory factor was present, a cytoplasmic fraction was prepared from control and hypothyroid adipocytes, and added to an in vitro translation system containing the hLPL mRNA. The hypothyroid cell fraction from adipose and heart yielded an increase in LPL translation, when compared to control extracts. Further experiments determined that the control adipocyte extract contained a translation-inhibitory factor that was 8-fold lower in activity in the hypothyroid extract. Using different LPL mRNA constructs in the in vitro translation reaction, the region that controlled translation was localized to nucleotides 1599 to 1638 (proximal 3' untranslated region (UTR)). To confirm the presence of a transacting factor, a sense RNA strand corresponding to this region was added to the in vitro translation reaction. This sense strand competed for the transacting factor in the control cell extract, yet had no effect on the hypothyroid cell extract. Thus, there is a translation repressor factor in the cytoplasm of rat adipocytes, and this factor is greatly reduced in activity in hypothyroid rat adipocytes. Because a similar mechanism of LPL regulation occurs in response to epinephrine, the absence of the translation repressor may be a mechanism for the loss of sensitivity of hypothyroid cells for catecholamines.

Adipose Tissue↗

Translational control genes in the sea urchin genome.

Sea urchin eggs and early cleavage stage embryos provide an example of regulated gene expression at the level of translation. The availability of the sea urchin genome offers the opportunity to investigate the "translational control" toolkit of this model system. The annotation of the genome reveals that most of the factors implicated in translational control are encoded by nonredundant genes in echinoderm, an advantage for future functional studies. In this paper, we focus on translation factors that have been shown or suggested to play crucial role in cell cycle and development of sea urchin embryos. Addressing the cap-binding translational control, three closely related eIF4E genes (class I, II, III) are present, whereas its repressor 4E-BP and its activator eIF4G are both encoded by one gene. Analysis of the class III eIF4E proteins in various phyla shows an echinoderm-specific amino acid substitution. Furthermore, an interaction site between eIF4G and poly(A)-binding protein is uncovered in the sea urchin eIF4G proteins and is conserved in metazoan evolution. In silico screening of the sea urchin genome has uncovered potential new regulators of eIF4E sharing the common eIF4E recognition motif. Taking together, these data provide new insights regarding the strong requirement of cap-dependent translation following fertilization. The genome analysis gives insights on the complexity of eEF1B structure and motifs of functional relevance, involved in the translational control of gene expression at the level of elongation. Finally, because deregulation of translation process can lead to diseases and tumor formation in humans, the sea urchin orthologs of human genes implicated in human diseases and signaling pathways regulating translation were also discussed.

Amino Acid Sequence↗

Cross-cultural comparisons of the content of SF-36 translations across 10 countries: results from the IQOLA Project. International Quality of Life Assessment.

Increasingly, translated and culturally adapted health-related quality of life measures are being used in cross-cultural research. To assess comparability of results, researchers need to know the comparability of the content of the questionnaires used in different countries. Based on an item-by-item discussion among International Quality of Life Assessment (IQOLA) investigators of the content of the translated versions of the SF-36 in 10 countries, we discuss the difficulties that arose in translating the SF-36. We also review the solutions identified by IQOLA investigators to translate items and response choices so that they are appropriate within each country as well as comparable across countries. We relate problems and solutions to ratings of difficulty and conceptual equivalence for each item. The most difficult items to translate were physical functioning items that refer to activities not common outside the United States and items that use colloquial expressions in the source version. Identifying the origin of the source items, their meaning to American English-speaking respondents and American English synonyms, in response to country-specific translation issues, greatly helped the translation process. This comparison of the content of translated SF-36 items suggests that the translations are culturally appropriate and comparable in their content.

Cross-Cultural Comparison↗

The Danish SF-36 Health Survey: translation and preliminary validity studies.

This article reports on the Danish translation of SF-36 and discusses the procedures used for translation improvement, translation evaluation, and scale evaluation. We followed the standard procedures of the International Quality of Life Assessment (IQOLA) Project including forward and backward translation, independent assessment of translation quality, assessment of response-choice weighting through visual analogue scale (VAS) investigations, and psychometric testing of the translated questionnaire. We found that backward translation, independent quality assessment, and VAS studies provided useful information for translation improvement. The Danish SF-36 received a favorable translation evaluation by independent rating; however, interrater agreement was low. Preliminary validity studies generally supported the internal consistency and homogeneity of the Danish SF-36, and the questionnaire performed satisfactorily in distinguishing depressive patients from nonpatients. On the basis of this and other studies, we recommend use of the Danish SF-36 in research.

Denmark↗

Two rat surfactant protein A isoforms arise by a novel mechanism that includes alternative translation initiation.

A single gene for rat surfactant protein A (SP-A) encodes two isoforms that are distinguished by an isoleucine-lysine-cysteine (IKC) N-terminal extension (SP-A and IKC-SP-A). Available evidence suggests that the variants are generated by alternative signal peptidase cleavage of the nascent polypeptide at a primary site (Cys(-)(1)-Asn(1)) and a secondary site (Gly(-)(4)-Ile(-)(3)). In this study, we used site-directed mutagenesis and heterologous expression in vitro and in insect cells to the examine mechanisms that may lead to alternative signal peptidase cleavage including alternative translation initiation at two in-frame AUGs (Met(-)(30) and Met(-)(20)), a suboptimal context for hydrolysis at the primary cleavage site, or cotranslational protein modifications that expose an otherwise cryptic secondary cleavage site. In vitro translation of a rat cDNA for SP-A resulted in both 28 and 29 kDa primary translation products on SDS-PAGE analysis, while translation of cDNAs encoding Met-30Ala and Met-20Ala mutations resulted in only the single 28 and 29 kDa molecular mass species, respectively. These data are consistent with translation initiation at both Met(-)(30) and Met(-)(20) during in vitro synthesis of SP-A. The Met-30Ala mutation reduced expression of the longer isoform in insect cells, indicating that the Met(-)(30) site also contributes to eucaryotic protein expression. Forcing translation initiation at Met(-)(30) by optimizing the Kozak consensus sequence surrounding that codon or by mutating the Met(-)(20) codon resulted in preferential expression of the longer SP-A isoform but reduced overall expression of the protein almost 10-fold. Both isoforms were generated to some degree whether translation was initiated at the codon for Met(-)(30) or Met(-)(20), indicating that the site of translation initiation is not the sole determinant of isoform generation and suggesting that either the context of the primary cleavage site is suboptimal or that cotranslational modifications affect cleavage. Preventing N-terminal glycosylation at Asn(1) did not affect the site of signal peptidase cleavage. Disruption of interchain disulfide formation at Cys(-)(1) by substitution with serine markedly enhanced cleavage at the Gly(-)(4)-Ile(-)(3) bond, but substitution with alanine enhanced cleavage at the Cys(-)(1)-Asn(1) bond. We conclude that rat SP-A isoforms arise by a novel mechanism that includes both alternative translation initiation at two in-frame AUGs and a suboptimal context for signal peptidase hydrolysis at the primary cleavage site.

Amino Acid Sequence↗

Molecular mechanisms for the control of translation by insulin.

Insulin acutely stimulates protein synthesis in mammalian cells, and this involves activation of the process of mRNA translation. mRNA translation is a complex multi-step process mediated by proteins termed translation factors. Several translation factors are regulated in response to insulin, often as a consequence of changes in their states of phosphorylation. The initiation factor eIF4E binds to the cap structure at the 5'-end of the mRNA and mediates assembly of an initiation-factor complex termed eIF4F. Assembly of this complex can be regulated by eIF4E-binding proteins (4E-BPs), which inhibit eIF4F complex assembly. Insulin induces phosphorylation of the 4E-BPs, resulting in alleviation of the inhibition. This regulatory mechanism is likely to be especially important for the control of the translation of specific mRNAs whose 5'-untranslated regions (5'-UTRs) are rich in secondary structure. Translation of another class of mRNAs, those with 5'-UTRs containing polypyrimidine tracts is also activated by insulin and this, like phosphorylation of the 4E-BPs, appears to involve the rapamycin-sensitive signalling pathway which leads to activation of the 70 kDa ribosomal protein S6 kinase (p70 S6 kinase) and the phosphorylation of the ribosomal protein S6. Overall stimulation of translation may involve activation of initiation factor eIF2B, which is required for all initiation events. This effect is dependent upon phosphatidylinositol 3-kinase and may involve the inactivation of glycogen synthase kinase-3 and consequent dephosphorylation of eIF2B, leading to its activation. Peptide-chain elongation can also be activated by insulin, and this is associated with the dephosphorylation and activation of elongation factor eEF2, probably as a consequence of the insulin-induced reduction in eEF2 kinase activity. Thus multiple signalling pathways acting on different steps in translation are involved in the activation of this process by insulin and lead both to general activation of translation and to the selective regulation of specific mRNAs.

Gene Expression Regulation↗

Target-specific arrest of mRNA translation by antisense 2'-O-alkyloligoribonucleotides.

We describe a novel experimental approach to investigate mRNA translation. Antisense 2'-O-allyl oligoribonucleotides (oligos) efficiently arrest translation of targeted mRNAs in rabbit reticulocyte lysate and wheat germ extract while displaying minimal non-specific effects on translation. Oligo/mRNA-hybrids positioned anywhere within the 5' UTR or the first approximately 20 nucleotides of the open reading frame block cap-dependent translation initiation with high specificity. The thermodynamic stability of hybrids between 2'-O-alkyl oligos and RNA permits translational inhibition with oligos as short as 10 nucleotides. This inhibition is independent of RNase H cleavage or modifications which render the mRNA untranslatable. We show that 2'-O-alkyl oligos can also be employed to interfere with cap-independent internal initiation of translation and to arrest translation elongation. The latter is accomplished by UV-crosslinking of psoralen-tagged 2'-O-methyloligoribonucleotides to the mRNA within the open reading frame. The utility of 2'-O-alkyloligoribonucleotides to arrest translation from defined positions within an mRNA provides new approaches to investigate mRNA translation.

Base Sequence↗

Secretion in yeast: structural features influencing the post-translational translocation of prepro-alpha-factor in vitro.

In vitro, efficient translocation and glycosylation of the precursor of yeast alpha-factor can take place post-translationally. This property of prepro-alpha-factor appears to be unique as it could not be extended to other yeast protein precursors such as preinvertase or preprocarboxypeptidase Y. In order to determine if specific domains of prepro-alpha-factor were involved in post-translational translocation, we carried out a series of experiments in which major domains were either deleted or fused onto reporter proteins. Fusion of various domains of prepro-alpha-factor onto the reporter protein alpha-globin did not allow post-translational translocation to occur in the yeast in vitro system. Prepro-alpha-factor retained its ability to be post-translationally translocated when parts or all of the pro region were deleted. Removal of the C-terminal repeats containing mature alpha-factor had the most profound influence as post-translational translocation decreased in proportion to the number of repeats deleted. Taken together, these results suggest that efficient post-translational translocation requires a signal sequence and the four C-terminal repeats. There does not however, appear to be specific information contained within the C-terminus, as their presence in fusion did not enable the post-translational translocation of reporter proteins. Lastly, the ability to post-translationally translocate radiochemically pure prepro-alpha-factor that had been isolated by immuno-affinity chromatography required the addition of a yeast lysate fraction. Moreover, post-translational translocation is a function of the microsomal membrane of yeast microsomes and not of a factor peculiar to the yeast lysate, as reticulocyte lysate supported this as well.

Glycosylation↗

How strong is the case for regulation of the initiation step of translation by elements at the 3' end of eukaryotic mRNAs?

The belief that initiation of translation requires communication between the 5' and 3' ends of the mRNA guides--or misguides--the interpretation of many experiments. The closed-loop model for initiation creates the expectation that sequences at the 3' end of eukaryotic mRNAs should regulate translation. This review looks closely at the evidence in three prominent cases where such regulation is claimed. The mRNAs in question encode 15-lipoxygenase, ceruloplasmin, and histones. Vertebrate histone mRNAs lack a poly(A) tail, instead of which a 3' stem-loop structure is said to promote translation by binding a protein which purportedly binds initiation factors. The proffered evidence for this hypothesis has many flaws. Temporal control of 15-lipoxygenase production in reticulocytes is often cited as another well-documented example of translational regulation via the 3' untranslated region, but inspection of the evidence reveals significant gaps and contradictions. Solid evidence is lacking also for the idea that a ribosomal protein binds to and shuts off translation of ceruloplasmin mRNA. Some viral RNAs that lack a poly(A) tail have alternative 3' structures which are said to promote translation via circularization of the mRNA, but in no case has this been shown convincingly. Interpretation of many experiments is compromised by possible effects of the 3' structures on mRNA stability rather than translation. The functional-half-life assay, which is often employed to rule out effects on mRNA stability, might not be adequate to settle the question. Other issues, such as the possibility of artifacts caused by overexpression of RNA-binding proteins, can complicate studies of translational regulation. There is no doubt that elements at the 3' end of eukaryotic mRNAs can regulate gene expression in a variety of ways. It has not been shown unequivocally that one of these ways involves direct participation of the 3' untranslated region in the initiation step of translation.

3' Untranslated Regions↗

Initiation of translation in prokaryotes and eukaryotes.

The mechanisms whereby ribosomes engage a messenger RNA and select the start site for translation differ between prokaryotes and eukaryotes. Initiation sites in polycistronic prokaryotic mRNAs are usually selected via base pairing with ribosomal RNA. That straightforward mechanism is made complicated and interesting by cis- and trans-acting elements employed to regulate translation. Initiation sites in eukaryotic mRNAs are reached via a scanning mechanism which predicts that translation should start at the AUG codon nearest the 5' end of the mRNA. Interest has focused on mechanisms that occasionally allow escape from this first-AUG rule. With natural mRNAs, three escape mechanisms - context-dependent leaky scanning, reinitiation, and possibly direct internal initiation - allow access to AUG codons which, although not first, are still close to the 5' end of the mRNA. This constraint on the initiation step of translation in eukaryotes dictates the location of transcriptional promoters and may have contributed to the evolution of splicing.The binding of Met-tRNA to ribosomes is mediated by a GTP-binding protein in both prokaryotes and eukaryotes, but the more complex structure of the eukaryotic factor (eIF-2) and its association with other proteins underlie some aspects of initiation unique to eukaryotes. Modulation of GTP hydrolysis by eIF-2 is important during the scanning phase of initiation, while modulating the release of GDP from eIF-2 is a key mechanism for regulating translation in eukaryotes. Our understanding of how some other protein factors participate in the initiation phase of translation is in flux. Genetic tests suggest that some proteins conventionally counted as eukaryotic initiation factors may not be required for translation, while other tests have uncovered interesting new candidates. Some popular ideas about the initiation pathway are predicated on static interactions between isolated factors and mRNA. The need for functional testing of these complexes is discussed. Interspersed with these theoretical topics are some practical points concerning the interpretation of cDNA sequences and the use of in vitro translation systems. Some human diseases resulting from defects in the initiation step of translation are also discussed.

Animals↗