Reported translational bypass in a trpR ′- lacZ ′ fusion is accounted for by unusual initiation and +1 frameshifting 1 1Edited by J. H. Miller

Wills, Norma M.; Ingram, Jennifer A; Gesteland, Raymond F.; Atkins, John F.

doi:10.1006/jmbi.1997.1187

Cited by 16 publications

(11 citation statements)

References 19 publications

(19 reference statements)

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“…8,9 In fact, the data exemplify this pitfall in two cases: the triplets GUG and UUG are known to support initiation at substantial frequencies, 10 and we observed a very high rate of enzyme synthesis with either of these triplets as the takeoff/landing site couple: about Reporters with the indicated takeoff sites (and a matching landing site 16 nt downstream) were grown into log phase in M63-glucose medium and induced for one mass doubling. Results are induced b-galactosidase differential rate, meanGstandard error, with number of replicates in parentheses.…”

Section: Resultsmentioning

confidence: 88%

Spontaneous Ribosome Bypassing in Growing Cells

Lindsley

Gallant

Doneanu

et al. 2005

Journal of Molecular Biology

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Section: Resultsmentioning

confidence: 88%

Spontaneous Ribosome Bypassing in Growing Cells

Lindsley

Gallant

Doneanu

et al. 2005

Journal of Molecular Biology

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“…As the frameshift on the G string is relatively inefficient, it may be that many other examples simply go unnoticed. There has been a report of an inefficient "out-offrame" net ϩ1 recoding event in lacZ that shifted ribosomes back into the wild-type reading frame (32); could this be an example of a bacterial translational compensation mechanism? Investigations of other mutations that confer clinically relevant ACV r on HSV may reveal other interesting mechanisms that permit these viruses to evade chemotherapy and retain pathogenicity.…”

Section: Discussionmentioning

confidence: 99%

Translational Compensation of a Frameshift Mutation Affecting Herpes Simplex Virus Thymidine Kinase Is Sufficient To Permit Reactivation from Latency

Griffiths

Chen

Horsburgh

et al. 2003

J Virol

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Herpes simplex virus thymidine kinase is important for reactivation of virus from its latent state and is a target for the antiviral drug acyclovir. Most acyclovir-resistant isolates have mutations in the thymidine kinase gene; however, how these mutations confer clinically relevant resistance is unclear. Reactivation from explanted mouse ganglia was previously observed with a patient-derived drug-resistant isolate carrying a single guanine insertion within a run of guanines in the thymidine kinase gene. Despite this mutation, low levels of active enzyme were synthesized following an unusual ribosomal frameshift. Here we report that a virus, generated from a pretherapy isolate from the same patient, engineered to lack thymidine kinase activity, was competent for reactivation. This suggested that the clinical isolate contains alleles of other genes that permit reactivation in the absence of thymidine kinase. Therefore, to establish whether thymidine kinase synthesized via a ribosomal frameshift was sufficient for reactivation under conditions where reactivation requires this enzyme, we introduced the mutation into the well-characterized strain KOS. This mutant virus reactivated from latency, albeit less efficiently than KOS. Plaque autoradiography revealed three phenotypes of reactivating viruses: uniformly low thymidine kinase activity, mixed high and low activity, and uniformly high activity. We generated a recombinant thymidine kinase-null virus from a reactivating virus expressing uniformly low activity. This virus did not reactivate, confirming that mutations in other genes that would influence reactivation had not arisen. Therefore, in strains that require thymidine kinase for reactivation from latency, low levels of enzyme synthesized via a ribosomal frameshift can suffice.

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“…The resulting fragment was cloned into the HindIII and ApaI restriction sites of the plasmid vector pSKAGS (46). The gst-ЈlacZ fusions of the (postulated) SECIS elements of hydV (Desulfomicrobium baculatum), selD (Treponema denticola), and the selW-like gene (Campylobacter jejuni) were generated in the same way, except that the respective gene regions were amplified from synthetic oligonucleotides.…”

Section: Methodsmentioning

confidence: 99%

Factors and Selenocysteine Insertion Sequence Requirements for the Synthesis of Selenoproteins from a Gram-Positive Anaerobe in Escherichia coli

Gursinsky

Gröbe

Schierhorn

et al. 2008

Appl Environ Microbiol

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Selenoprotein synthesis in Escherichia coli strictly depends on the presence of a specific selenocysteine insertion sequence (SECIS) following the selenocysteine-encoding UGA codon of the respective mRNA. It is recognized by the selenocysteine-specific elongation factor SelB, leading to cotranslational insertion of selenocysteine into the nascent polypeptide chain. The synthesis of three different selenoproteins from the grampositive anaerobe Eubacterium acidaminophilum in E. coli was studied. Incorporation of 75 Se into glycine reductase protein B (GrdB1), the peroxiredoxin PrxU, and selenophosphate synthetase (SelD1) was negligible in an E. coli wild-type strain and was fully absent in an E. coli SelB mutant. Selenoprotein synthesis, however, was strongly increased if selB and selC (tRNA Sec ) from E. acidaminophilum were coexpressed. Putative secondary structures downstream of the UGA codons did not show any sequence similarity to each other or to the E. coli SECIS element. However, mutations in these structures strongly reduced the amount of 75 Se-labeled protein, indicating that they indeed act as SECIS elements. UGA readthrough mediated by the three different SECIS elements was further analyzed using gst-lacZ translational fusions. In the presence of selB and selC from E. acidaminophilum, UGA readthrough was 36 to 64% compared to the respective cysteine-encoding UGC variant. UGA readthrough of SECIS elements present in Desulfomicrobium baculatum (hydV), Treponema denticola (selD), and Campylobacter jejuni (selW-like gene) was also considerably enhanced in the presence of E. acidaminophilum selB and selC. This indicates recognition of these SECIS elements and might open new perspectives for heterologous selenoprotein synthesis in E. coli.The 21st amino acid selenocysteine (Sec) is present in a variety of different eukaryotic, archaeal, and bacterial proteins (11,25,26,29,50). The insertion of selenocysteine into growing peptide chains is directed by an opal (UGA) codon (8, 51), which in the standard genetic code signals termination of translation. Selenoprotein synthesis in Escherichia coli has been elucidated and intensively studied by Böck and coworkers (6) and requires the products of the genes selA, selB, selC, and selD. selC encodes a selenocysteine-specific tRNA Sec that is first charged with serine by seryl-tRNA synthetase. It is converted to selenocysteyl-tRNA Sec by selenocysteine synthase, the gene product of selA. A low-molecular-weight selenium donor, selenophosphate, is required for this reaction. It is synthesized from ATP and selenide by selenophosphate synthetase, SelD (41).Recoding of UGA into selenocysteine requires a special mRNA motif, the selenocysteine insertion sequence (SECIS) (22). In mRNAs encoding bacterial selenoproteins, it is located at the 3Ј site of the UGA codon (52). This mRNA structure is recognized by the bacterial selenocysteine-specific elongation factor SelB, which functions homologously to the standard elongation factor EF-Tu (9, 24). SelB binds GTP and selenocysteyl-...

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Reported translational bypass in a trpR ′- lacZ ′ fusion is accounted for by unusual initiation and +1 frameshifting 1 1Edited by J. H. Miller

Cited by 16 publications

References 19 publications

Spontaneous Ribosome Bypassing in Growing Cells

Spontaneous Ribosome Bypassing in Growing Cells

Translational Compensation of a Frameshift Mutation Affecting Herpes Simplex Virus Thymidine Kinase Is Sufficient To Permit Reactivation from Latency

Factors and Selenocysteine Insertion Sequence Requirements for the Synthesis of Selenoproteins from a Gram-Positive Anaerobe in Escherichia coli

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