Pseudouridine in the Anticodon G Psi A of Plant Cytoplasmic TRNA(Tyr) is Required for UAG and UAA Suppression in the TMV-specific Context

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Specialty Biochemistry

Date 1992 Nov 25

PMID 1461724

Citations 40

Authors

K Zerfass

H Beier

Affiliations

Soon will be listed here.

Abstract

We have previously isolated and sequenced Nicotiana cytoplasmic tRNA(Tyr) with G psi A anticodon which promotes readthrough over the leaky UAG termination codon at the end of the 126 K cistron of tobacco mosaic virus RNA and we have demonstrated that tRNA(Tyr) with Q psi A anticodon is no UAG suppressor. Here we show that the nucleotide in the middle of the anticodon (i.e., psi 35) also contributes to the suppressor efficiency displayed by cytoplasmic tRNA(Tyr). A tRNA(Tyr) with GUA anticodon was synthesized in vitro using T7 RNA polymerase transcription. This tRNA(Tyr) was unable to suppress the UAG codon, indicating that nucleotide modifications in the anticodon of tRNA(Tyr) have either stimulating (i.e., psi 35) or inhibitory (i.e., Q34) effects on suppressor activity. Furthermore, we have shown that the UAA but not the UGA stop codon is also efficiently recognized by tobacco tRNA(G psi ATyr), if placed in the TMV context. Hence this is the first naturally occurring tRNA for which UAA suppressor activity has been demonstrated. In order to study the influence of neighbouring nucleotides on the readthrough capacity of tRNA(Tyr), we have established a system, in which part of the sequence around the leaky UAG codon of TMV RNA was inserted into a zein pseudogene which naturally harbours an UAG codon in the middle of the gene. The construct was cloned into the vector pSP65 and in vitro transcripts, generated by SP6 RNA polymerase, were translated in a wheat germ extract depleted of endogenous mRNAs and tRNAs. A number of mutations in the codons flanking the UAG were introduced by site-directed mutagenesis. It was found that changes at specific positions of the two downstream codons completely abolished the readthrough over the UAG by Nicotiana tRNA(Tyr), indicating that this tRNA needs a very specific codon context for its suppressor activity.

Citing Articles

Variation of tRNA modifications with and without intron dependency.

Hayashi S Front Genet. 2024; 15:1460902.

PMID: 39296543 PMC: 11408192. DOI: 10.3389/fgene.2024.1460902.

Identification of permissive amber suppression sites for efficient non-canonical amino acid incorporation in mammalian cells.

Bartoschek M, Ugur E, Nguyen T, Rodschinka G, Wierer M, Lang K Nucleic Acids Res. 2021; 49(11):e62.

PMID: 33684219 PMC: 8216290. DOI: 10.1093/nar/gkab132.

Epitranscriptomic marks: Emerging modulators of RNA virus gene expression.

Netzband R, Pager C Wiley Interdiscip Rev RNA. 2019; 11(3):e1576.

PMID: 31694072 PMC: 7169815. DOI: 10.1002/wrna.1576.

Deciphering the reading of the genetic code by near-cognate tRNA.

Blanchet S, Cornu D, Hatin I, Grosjean H, Bertin P, Namy O Proc Natl Acad Sci U S A. 2018; 115(12):3018-3023.

PMID: 29507244 PMC: 5866558. DOI: 10.1073/pnas.1715578115.

Pseudouridine in the Anticodon of Escherichia coli tRNATyr(QΨA) Is Catalyzed by the Dual Specificity Enzyme RluF.

Addepalli B, Limbach P J Biol Chem. 2016; 291(42):22327-22337.

PMID: 27551044 PMC: 5064010. DOI: 10.1074/jbc.M116.747865.

References

Grosjean H, de Henau S, Crothers D . On the physical basis for ambiguity in genetic coding interactions. Proc Natl Acad Sci U S A. 1978; 75(2):610-4. PMC: 411305. DOI: 10.1073/pnas.75.2.610. View

Ward D, Reich E . Conformational properties of polyformycin: a polyribonucleotide with individual residues in the syn conformation. Proc Natl Acad Sci U S A. 1968; 61(4):1494-501. PMC: 225282. DOI: 10.1073/pnas.61.4.1494. View

Stark B, Kole R, Bowman E, Altman S . Ribonuclease P: an enzyme with an essential RNA component. Proc Natl Acad Sci U S A. 1978; 75(8):3717-21. PMC: 392857. DOI: 10.1073/pnas.75.8.3717. View

Pelham H . Leaky UAG termination codon in tobacco mosaic virus RNA. Nature. 1978; 272(5652):469-71. DOI: 10.1038/272469a0. View

Siegel A, Hari V, Kolacz K . The effect of tobacco mosaic virus infection on host and virus-specific protein synthesis in protoplasts. Virology. 1978; 85(2):494-503. DOI: 10.1016/0042-6822(78)90456-7. View

Fritsch C, Mayo M, Hirth L . Further studies on the translation products of tobacco rattle virus RNA in vitro. Virology. 1977; 77(2):722-32. DOI: 10.1016/0042-6822(77)90494-9. View

Jank P, Nishimura S, Gross H . Rabbit liver tRNA1Val:I. Primary structure and unusual codon recognition. Nucleic Acids Res. 1977; 4(6):1999-2008. PMC: 342537. DOI: 10.1093/nar/4.6.1999. View

Bruening G, Beachy R, Scalla R, Zaitlin M . In vitro and in vivo translation of the ribonucleic acids of a cowpea strain of tobacco mosaic virus. Virology. 1976; 71(2):498-517. DOI: 10.1016/0042-6822(76)90377-9. View

Topal M, Fresco J . Complementary base pairing and the origin of substitution mutations. Nature. 1976; 263(5575):285-9. DOI: 10.1038/263285a0. View

10.

Topal M, Fresco J . Base pairing and fidelity in codon-anticodon interaction. Nature. 1976; 263(5575):289-93. DOI: 10.1038/263289a0. View

11.

Kasai H, Oashi Z, Harada F, Nishimura S, Oppenheimer N, Crain P . Structure of the modified nucleoside Q isolated from Escherichia coli transfer ribonucleic acid. 7-(4,5-cis-Dihydroxy-1-cyclopenten-3-ylaminomethyl)-7-deazaguanosine. Biochemistry. 1975; 14(19):4198-208. DOI: 10.1021/bi00690a008. View

12.

Paterson R, Knight C . Protein synthesis in tobacco protoplasts infected with tobacco mosaic virus. Virology. 1975; 64(1):10-22. DOI: 10.1016/0042-6822(75)90074-4. View

13.

Laskey R, Mills A . Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem. 1975; 56(2):335-41. DOI: 10.1111/j.1432-1033.1975.tb02238.x. View

14.

Zerfass K, Beier H . The leaky UGA termination codon of tobacco rattle virus RNA is suppressed by tobacco chloroplast and cytoplasmic tRNAs(Trp) with CmCA anticodon. EMBO J. 1992; 11(11):4167-73. PMC: 556927. DOI: 10.1002/j.1460-2075.1992.tb05510.x. View

15.

Alonso E, Garcia-Luque I, De La Cruz A, Wicke B, Avila-Rincon M, Serra M . Nucleotide sequence of the genomic RNA of pepper mild mottle virus, a resistance-breaking tobamovirus in pepper. J Gen Virol. 1991; 72 ( Pt 12):2875-84. DOI: 10.1099/0022-1317-72-12-2875. View

16.

Ugaki M, Tomiyama M, Kakutani T, Hidaka S, Kiguchi T, Nagata R . The complete nucleotide sequence of cucumber green mottle mosaic virus (SH strain) genomic RNA. J Gen Virol. 1991; 72 ( Pt 7):1487-95. DOI: 10.1099/0022-1317-72-7-1487. View

17.

Entwistle J, Knudsen S, Muller M, Cameron-Mills V . Amber codon suppression: the in vivo and in vitro analysis of two C-hordein genes from barley. Plant Mol Biol. 1991; 17(6):1217-31. DOI: 10.1007/BF00028737. View

18.

Skuzeski J, Nichols L, GESTELAND R, Atkins J . The signal for a leaky UAG stop codon in several plant viruses includes the two downstream codons. J Mol Biol. 1991; 218(2):365-73. DOI: 10.1016/0022-2836(91)90718-l. View

19.

Zawadzki V, Gross H . Rapid and simple purification of T7 RNA polymerase. Nucleic Acids Res. 1991; 19(8):1948. PMC: 328134. DOI: 10.1093/nar/19.8.1948. View

20.

Sprinzl M, Dank N, Nock S, Schon A . Compilation of tRNA sequences and sequences of tRNA genes. Nucleic Acids Res. 1991; 19 Suppl:2127-71. PMC: 331350. DOI: 10.1093/nar/19.suppl.2127. View