Stop making sense or Regulation at the level of termination in eukaryotic protein synthesis

Valle, Rosaura P.C.; Morch, Marie-Dominique

doi:10.1016/0014-5793(88)81225-0

Cited by 79 publications

(50 citation statements)

References 109 publications

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“…of nonsense suppression (45). As above, the ability of the two nonsense codons to promote reinitiation at GCN4 could be explained by this hypothesis, since the presence of these two triplets would eliminate the proposed tRNA-tRNA interactions that hinder reinitiation.…”

Section: Resultsmentioning

confidence: 82%

Effect of sequence context at stop codons on efficiency of reinitiation in GCN4 translational control.

Grant¹,

Hinnebusch²

1994

Mol. Cell. Biol.

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Translational control of the GCN4 gene involves two short open reading frames in the mRNA leader (uORF1 and uORF4) that differ greatly in the ability to allow reinitiation at GCN4 following their own translation. The low efficiency of reinitiation characteristic of uORF4 can be reconstituted in a hybrid element in which the last codon of uORF1 and 10 nucleotides 3' to its stop codon (the termination region) are substituted with the corresponding nucleotides from uORF4. To define the features of these 13 nucleotides that determine their effects on reinitiation, we separately randomized the sequence of the third codon and termination region of the uORF1-u0RF4 hybrid and selected mutant alleles with the high-level reinitiation that is characteristic of uORF1. The results indicate that many different A+U-rich triplets present at the third codon of uORF1 can overcome the inhibitory effect of the termination region derived from uORF4 on the efficiency of reinitiation at GCN4. Efficient reinitiation is not associated with codons specifying a particular amino acid or isoacceptor tRNA. Similarly, we found that a diverse collection of A+U-rich sequences present in the termination region of uORF1 could restore efficient reinitiation at GCN4 in the presence of the third codon derived from uORF4. To explain these results, we propose that reinitiation can be impaired by stable base pairing between nucleotides flanking the uORF1 stop codon and either the tRNA which pairs with the third codon, the rRNA, or sequences located elsewhere in GCN4 mRNA. We suggest that these interactions delay the resumption of scanning following peptide chain termination at the uORF and thereby lead to ribosome dissociation from the mRNA.Regulation of the GCN4 gene of Saccharomyces cerevisiae is one of the best-characterized examples of genespecific translational control in a eukaryotic organism (reviewed in references 21 and 22). In wild-type yeast cells, GCN4 mRNA is translated efficiently only when cells are starved for an amino acid. Because GCN4 is a transcriptional activator of amino acid biosynthetic genes, increased translation of GCN4 mRNA can overcome the amino acid limitation which signalled its derepression. Translational control of GCN4 depends on four short open reading frames of two or three codons in length present in the GCN4 mRNA leader, of which the first (uORF1) and fourth (uORF4) are sufficient for nearly wild-type regulation. The presence of uORF4 alone results in constitutively repressed GCN4 translation. In contrast, uORF1 is a relatively weak translational barrier that must be present upstream of uORF4 for the derepression of GCN4 translation that occurs in response to amino acid starvation (32).In addition to the uORFs, derepression of GCN4 translation in starved cells is dependent on a reduction in the level of active eukaryotic translation initiation factor 2 (eIF-2) (13). The function of eIF-2 in the initiation process is to bind charged initiator tRNA"' in a ternary complex with GTP to the small ribosomal subunit, formi...

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Section: Resultsmentioning

confidence: 82%

Effect of sequence context at stop codons on efficiency of reinitiation in GCN4 translational control.

Grant¹,

Hinnebusch²

1994

Mol. Cell. Biol.

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“…If these viral blocks are involved in ribosome binding at the 25 g and CP OgFs then it has implications on the modeof translation of the other open reading frames, as these blocks were not present upstream from their AUGs, The e~pression of the other ORFs could therefore be by a 'relay race' mechanism of translation or translational frameshifting [23,24].…”

Section: Resultsmentioning

confidence: 99%

Occurrence of chloroplast ribosome recognition sites within conserved elements of the RNA genomes of carlaviruses

Foster

Mills

1991

FEBS Letters

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The nucl¢otide sequences upstream from the ,=~rlavirus olin reading frames wcrc exan~ined for direct sequence homology, Blocks of homology were evident upstream from the 25 K ORFs eF potato virus S (PVS), potato virus M (PVM} and lily symptomless virus (LSV), and upstream from th,= coast protein initiation codons of PVS, PVM, LSV. carnation latent virus and Hdenium virus S, Then,= blocks, which correspond to the Y.terminal resions of the subjenomic RNAs. were sl~own to contain potential ribosome rcco~,nitlon sequences, The distances between tit,= bindtn8 sites and initiation codons ransed from 20 to 40 nucleotides on the viral RNAs, Whilsl tit,= m.'tjority of chloroplasts mRNAs have a distance of 8 nucleotides between bindin8 site and initiation codon, the remaininB have a distance o1" 23 nucleotides which is similar to that reported here for the carlaviru~s.Subgenomic promoter: Seq nonce homology; Carlavirus; Ribosome bindin6 sit'=

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“…To date a mammalian UAA suppressor has not been isolated and indeed, UAA was once considered to be an absolute termination codon in mammalian cells (20,21). Recently, it has been possible to demonstrate readthrough suppression of UAA in viral systems where the natural UAG codon [MuLV (2, 9) and TMV (7) (Fig.…”

Section: Discussionmentioning

confidence: 99%

Identification of amino acids inserted during suppression of UAA and UGA termination codons at the gag-pol junction of Moloney murine leukemia virus.

Feng

Copeland

Oroszlan

et al. 1990

Proc. Natl. Acad. Sci. U.S.A.

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Expression of the murine leukemia virus poi gene occurs by translational readthrough of an in-frame UAG codon between the gag and poi coding regions. In a previous study, we mutated the UAG codon to UAA or UGA and demonstrated that both of these termination codons could be suppressed in reticulocyte lysates and in infected cells with the same efficiency as UAG. We now report the identity of the amino acids inserted in vitro in response to UAA and UGA in fusion products containing the gag-pol junction region. The results show that UAA, like UAG, directs the incorporation of glutamine, whereas UGA directs the incorporation of three amino acids, arginine, cysteine, and tryptophan. To our knowledge, this is the first report indicating misreading of UAA as glutamine and UGA as arginine and cysteine in higher eukaryotes. Interestingly, although our protein synthesis system presumably contains other known UAG and UGA suppressors, these tRNAs did not suppress the termination codons in our experiments. Thus, it seems possible that the sequence surrounding the gag-pol junction not only promotes suppression but also helps determine which tRNAs function in suppression.Readthrough suppression of termination codons is a mechanism of translational regulation that is used by a number of viruses infecting higher eukaryotes. The analysis of this phenomenon thus provides new information on the functional capabilities of the translational apparatus of these organisms. We have been investigating suppression in the murine leukemia virus (MuLV) system (1, 2). MuLV, like other mammalian type C retroviruses, regulates pol gene expression by translational readthrough ofan in-frame UAG codon between the gag and pol coding regions (3-5). Suppression of this termination codon occurs with a frequency of about 5% (3)(4)(5) and is mediated by insertion of a glutamine residue at the position corresponding to the UAG (6). The readthrough product is a 200-kDa gag-pol fusion protein, which is cleaved at a late stage in virus assembly to give rise to protease, reverse transcriptase, and integrase (3).To determine whether the signals that govern readthrough are unique for UAG, we used oligonucleotide-directed mutagenesis to change the UAG at the Moloney (Mo)-MuLV gag-pol junction to UAA or UGA (2). Interestingly, we found that these termination codons could be efficiently suppressed both in reticulocyte lysates and in infected cells (2). These results demonstrated that the suppression signals are not specific for UAG and that mammalian cells and cell extracts contain tRNAs capable of suppressing UGA and UAA codons which appear in a retroviral context. Similar findings have been reported by other investigators working with tobacco mosaic virus (TMV) (7), Sindbis virus (8), and MuLV (9).In the present study we have identified the amino acids inserted in response to UAA and UGA by using an in vitro MuLV suppression system (1). We find that UAA, like UAG, directs the incorporation of glutamine into protein, whereas UGA directs the incorporation ...

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Stop making sense or Regulation at the level of termination in eukaryotic protein synthesis

Cited by 79 publications

References 109 publications

Effect of sequence context at stop codons on efficiency of reinitiation in GCN4 translational control.

Effect of sequence context at stop codons on efficiency of reinitiation in GCN4 translational control.

Occurrence of chloroplast ribosome recognition sites within conserved elements of the RNA genomes of carlaviruses

Identification of amino acids inserted during suppression of UAA and UGA termination codons at the gag-pol junction of Moloney murine leukemia virus.

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