Coupling of Open Reading Frames by Translational Bypassing

Herr, Alan J.; Atkins, John F.; Gesteland, Raymond F.

doi:10.1146/annurev.biochem.69.1.343

Cited by 61 publications

(72 citation statements)

References 96 publications

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“…14), which subsequently was shown to have a role in normally restraining forward mRNA slippage 11 . These diverse stimulatory elements are likely to provide a complex regulatory network of physical interactions and, presumably, kinetic effects that influence bypassing 15 .…”

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High-efficiency translational bypassing of non-coding nucleotides specified by mRNA structure and nascent peptide

et al. 2014

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The gene product 60 (gp60) of bacteriophage T4 is synthesized as a single polypeptide chain from a discontinuous reading frame as a result of bypassing of a non-coding mRNA region of 50 nucleotides by the ribosome. To identify the minimum set of signals required for bypassing, we recapitulated efficient translational bypassing in an in vitro reconstituted translation system from Escherichia coli. We find that the signals, which promote efficient and accurate bypassing, are specified by the gene 60 mRNA sequence. Systematic analysis of the mRNA suggests unexpected contributions of sequences upstream and downstream of the non-coding gap region as well as of the nascent peptide. During bypassing, ribosomes glide forward on the mRNA track in a processive way. Gliding may have a role not only for gp60 synthesis, but also during regular mRNA translation for reading frame selection during initiation or tRNA translocation during elongation.

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High-efficiency translational bypassing of non-coding nucleotides specified by mRNA structure and nascent peptide

et al. 2014

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“…Long distance bypassing is required for expression of phage T4 gene 60 (8,9) where 50 nucleotides are bypassed by 50% of the ribosomes that initiate translation (10). Several different signals in the mRNA are required.…”

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Factors That Influence Selection of Coding Resumption Sites in Translational Bypassing

Herr

Wills

Nelson

et al. 2004

Journal of Biological Chemistry

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This study investigates bypassing initiated from codons immediately 5 of a stop codon. The mRNA slips and is scanned by the peptidyl-tRNA for a suitable landing site, and standard decoding resumes at the next 3 codon. This work shows that landing sites with potentially strong base pairing between the peptidyl-tRNA anticodon and mRNA are preferred, but sites with little or no potential for Watson-Crick or wobble base pairing can also be utilized. These results have implications for re-pairing in ribosomal frameshifting. Shine-Dalgarno sequences in the mRNA can alter the distribution of landing sites observed. The bacteriophage T4 gene 60 nascent peptide, known to influence take-off in its native context, imposes stringent P-site pairing requirements, thereby limiting the number of suitable landing sites.Standard genetic decoding is dependent on the formation and maintenance of cognate codon:anticodon pairing. Once established in the ribosomal A-site, this interaction continues through the ribosomal P-site. However, at specific sites in mRNAs ranging from those in retroviruses to Escherichia coli, ribosomes can shift reading frame for gene expression purposes. Most of these specific changes in reading frame involve dissociation of codon:anticodon pairing in the ribosomal P-site and re-pairing at an overlapping codon. Although mechanisms for ensuring the fidelity of pairing in the A-site have been well studied (1), much less is known about the stringency of pairing in the ribosomal P-site. In single nucleotide frameshifting events, the stringency of re-pairing is often difficult to separate from the efficiency of dissociation because the codons share two nucleotides in common. Dissociation and re-pairing, however, are uncoupled in the related phenomenon of translational bypassing. Following initiation of mRNA slippage (or take-off), ribosomes move further downstream, bypassing a block of nucleotides, before coding resumes at the codon 3Ј of the landing site. A careful analysis of landing site selection offers a means to investigate P-site pairing stringency.Long range translational bypassing in response to A-site codons whose aminoacyl-tRNAs are limiting has been demonstrated (2-4). In these cases, the peptidyl-tRNA dissociates from the ribosomal P-site codon, scans the coding gap, and re-pairs to mRNA at complementary triplets. There are also examples of short distance hopping over stop codons

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“…1). The prototypical case has been the translation of the phage T4 gene 60 mRNA coding for a topoisomerase subunit, which involves the bypassing of a 50-nt untranslated segment (or ''coding gap'') between a GGA ''takeoff'' triplet and a GGA ''landing site'' triplet.…”

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“…This case of bypassing depends on very specific features of the sequence (1-3): (i) identity of the takeoff and landing site triplets, implying that peptidyl-tRNA is what makes the trip; (ii) optimum spacing of the takeoff and landing sites; (iii) a terminator triplet immediately 3Ј of the takeoff triplet; (iv) a stem-loop of optimal stability and placement in the region after the takeoff site; and (v) a particular amino acid sequence of the peptide encoded in a region 5Ј of the takeoff site. The role of these sequence elements and of participating factors has been investigated in detail (1,(4)(5)(6).…”

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“…It seems intuitively likely that the complex slides through the intervening or coding gap region while remaining associated with the mRNA and influenced by elements in its sequence, a model Herr et al (1) refer to as ''scanning.'' However, it remains conceivable that the ribosome::peptidyl-tRNA complex detaches completely from the message and somehow hops over the coding gap to reengage at the landing site, a theoretical possibility that the authors of the aforementioned review term ''bridging.…”

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Evidence that the bypassing ribosome travels through the coding gap

Gallant¹,

Bonthuis²,

Lindsley³

2003

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

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In translational bypassing, a peptidyl-tRNA::ribosome complex skips over a number of nucleotides in a messenger sequence and resumes protein chain elongation after a ''landing site'' downstream of the bypassed region. The present experiments demonstrate that the complex ''scans'' processively through the bypassed region. This conclusion rests on three observations. (i) When two potential ''landing sites'' are present, the protein sequence of the product shows that virtually all ribosomes land at the first and virtually none at the second. (ii) In such a sequence with two landing sites, the presence of a terminator triplet in phase in the coding region immediately after the first landing site drastically reduces the efficiency of bypassing. (iii) Internally complementary sequences that can form a stable stem͞loop in the bypassed region significantly reduce the efficiency of bypassing. We analyze bypassing from a given ''takeoff'' site to ''landing sites'' at different distances downstream so as to derive estimates of the frequency of ribosome takeoff and of the stability of the bypassing complex.I n ''translational bypassing,'' a ribosome skips over a number of nucleotides in a messenger sequence so as to join the information from two noncontiguous ORFs into the sequence of a single, continuous polypeptide chain (reviewed in ref. 1). The prototypical case has been the translation of the phage T4 gene 60 mRNA coding for a topoisomerase subunit, which involves the bypassing of a 50-nt untranslated segment (or ''coding gap'') between a GGA ''takeoff'' triplet and a GGA ''landing site'' triplet. (In the text, we use boldface to show RNA triplets and normal type for triplets or other sequences in DNA.) This case of bypassing depends on very specific features of the sequence (1-3): (i) identity of the takeoff and landing site triplets, implying that peptidyl-tRNA is what makes the trip; (ii) optimum spacing of the takeoff and landing sites; (iii) a terminator triplet immediately 3Ј of the takeoff triplet; (iv) a stem-loop of optimal stability and placement in the region after the takeoff site; and (v) a particular amino acid sequence of the peptide encoded in a region 5Ј of the takeoff site. The role of these sequence elements and of participating factors has been investigated in detail (1, 4-6).The sequence rules governing this kind of bypass event seem very constraining. We have reported on a type of bypass event that seems to be subject to more relaxed rules and may therefore occur on a wide variety of sequences. This is a bypass induced by ribosome pausing at a ''hungry'' codon calling for an aminoacyltRNA in short supply (7,8). We showed that this kind of bypass shared with the gene 60 case only the requirement for synonymous takeoff and landing sites. Otherwise, it could be demonstrated over various coding gap distances (from 7 to at least 40 nt), and seemed largely independent of the sequence in the coding gap and upstream of it (7). In those experiments and the present ones, we inhibited the activation of isole...

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Coupling of Open Reading Frames by Translational Bypassing

Cited by 61 publications

References 96 publications

High-efficiency translational bypassing of non-coding nucleotides specified by mRNA structure and nascent peptide

High-efficiency translational bypassing of non-coding nucleotides specified by mRNA structure and nascent peptide

Factors That Influence Selection of Coding Resumption Sites in Translational Bypassing

Evidence that the bypassing ribosome travels through the coding gap

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