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. 1984 May;3(5):1091–1096. doi: 10.1002/j.1460-2075.1984.tb01934.x

The molecular basis for the differential translation of TMV RNA in tobacco protoplasts and wheat germ extracts

Hildburg Beier 1,*, Miroslawa Barciszewska 1,1, Hans-Dieter Sickinger 1
PMCID: PMC557478  PMID: 16453524

Abstract

Translation of tobacco mosaic virus (TMV) RNA in tobacco protoplasts yields the 17.5-K coat protein, a 126-K protein and a 183-K protein which is generated by an efficient readthrough over the UAG termination codon at the end of the 126-K cistron. In wheat germ extracts, however, only the 5'-proximal 126-K cistron is translated whereas the 183-K readthrough protein is not synthesized. Purification and sequence analysis of the endogenous tyrosine tRNAs revealed that the uninfected tobacco plant contains two tRNAsTyr, both with GΨA anticodons which stimulate the UAG readthrough in vitro and presumably in vivo. In contrast, ˜85% of the tRNATyr from wheat germ contains a QΨA anticodon and ˜15% has a GΨA anticodon. Otherwise the sequences of tRNAsTyr from wheat germ and tobacco are identical. UAG readthrough and hence synthesis of the 183-K protein is only stimulated by tRNATyrGΨA and not at all by tRNATyrQΨA. The tRNAsTyr from wheat leaves were also sequenced. This revealed that adult wheat contains tRNATyrGΨA only. This is very much in contrast to the situation in animals, where Q-containing tRNAs are characteristic for adult tissues whereas Q deficiency is typical for the neoplastic and embryonic state.

Keywords: readthrough, tobacco mosaic virus (TMV), wheat germ tRNAsTyr, translation, UAG suppression

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Beachy R. N., Zaitlin M. Characterization and in vitro translation of the RNAs from less-than-full-length, virus-related, nucleoprotein rods present in tobacco mosaic virus preparations. Virology. 1977 Aug;81(1):160–169. doi: 10.1016/0042-6822(77)90068-x. [DOI] [PubMed] [Google Scholar]
  2. Beier H., Barciszewska M., Krupp G., Mitnacht R., Gross H. J. UAG readthrough during TMV RNA translation: isolation and sequence of two tRNAs with suppressor activity from tobacco plants. EMBO J. 1984 Feb;3(2):351–356. doi: 10.1002/j.1460-2075.1984.tb01810.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Beier H., Mundry K. W., Issinger O. G. In vivo and in vitro translation of the RNAs of four tobamoviruses. Intervirology. 1980;14(5-6):292–299. doi: 10.1159/000149199. [DOI] [PubMed] [Google Scholar]
  4. Bruening G., Beachy R. N., Scalla R., Zaitlin M. In vitro and in vivo translation of the ribonucleic acids of a cowpea strain of tobacco mosaic virus. Virology. 1976 Jun;71(2):498–517. doi: 10.1016/0042-6822(76)90377-9. [DOI] [PubMed] [Google Scholar]
  5. Davies J. W., Kaesberg P. Translation of virus mRNA: protein synthesis directed by several virus RNAs in a cell-free extract from wheat germ. J Gen Virol. 1974 Oct;25(1):11–20. doi: 10.1099/0022-1317-25-1-11. [DOI] [PubMed] [Google Scholar]
  6. Gillum A. M., Urquhart N., Smith M., RajBhandary U. L. Nucleotide sequence of salmon testes and salmon liver cytoplasmic initiator tRNA. Cell. 1975 Nov;6(3):395–405. doi: 10.1016/0092-8674(75)90189-0. [DOI] [PubMed] [Google Scholar]
  7. Goelet P., Lomonossoff G. P., Butler P. J., Akam M. E., Gait M. J., Karn J. Nucleotide sequence of tobacco mosaic virus RNA. Proc Natl Acad Sci U S A. 1982 Oct;79(19):5818–5822. doi: 10.1073/pnas.79.19.5818. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gross H. J., Krupp G., Domdey H., Raba M., Jank P., Lossow C., Alberty H., Ramm K., Sänger H. L. Nucleotide sequence and secondary structure of citrus exocortis and chrysanthemum stunt viroid. Eur J Biochem. 1982 Jan;121(2):249–257. doi: 10.1111/j.1432-1033.1982.tb05779.x. [DOI] [PubMed] [Google Scholar]
  9. Gupta R. C., Randerath K. Use of specific endonuclease cleavage in RNA sequencing-an enzymic method for distinguishing between cytidine and uridine residues. Nucleic Acids Res. 1977 Oct;4(10):3441–3454. doi: 10.1093/nar/4.10.3441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Harada F., Nishimura S. Possible anticodon sequences of tRNA His , tRNA Asm , and tRNA Asp from Escherichia coli B. Universal presence of nucleoside Q in the first postion of the anticondons of these transfer ribonucleic acids. Biochemistry. 1972 Jan 18;11(2):301–308. doi: 10.1021/bi00752a024. [DOI] [PubMed] [Google Scholar]
  11. Holmes W. M., Hurd R. E., Reid B. R., Rimerman R. A., Hatfield G. W. Separation of transfer ribonucleic acid by sepharose chromatography using reverse salt gradients. Proc Natl Acad Sci U S A. 1975 Mar;72(3):1068–1071. doi: 10.1073/pnas.72.3.1068. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Jank P., Shindo-Okada N., Nishimura S., Gross H. J. Rabbit liver tRNA1Val:I. Primary structure and unusual codon recognition. Nucleic Acids Res. 1977 Jun;4(6):1999–2008. doi: 10.1093/nar/4.6.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jay E., Bambara R., Padmanabhan R., Wu R. DNA sequence analysis: a general, simple and rapid method for sequencing large oligodeoxyribonucleotide fragments by mapping. Nucleic Acids Res. 1974 Mar;1(3):331–353. doi: 10.1093/nar/1.3.331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Johnson P. F., Abelson J. The yeast tRNATyr gene intron is essential for correct modification of its tRNA product. Nature. 1983 Apr 21;302(5910):681–687. doi: 10.1038/302681a0. [DOI] [PubMed] [Google Scholar]
  15. Kasai H., Nakanishi K., Macfarlane R. D., Torgerson D. F., Ohashi Z., McCloskey J. A., Gross H. J., Nishimura S. Letter: The structure of Q* nucleoside isolated from rabbit liver transfer ribonucleic acid. J Am Chem Soc. 1976 Aug 4;98(16):5044–5046. doi: 10.1021/ja00432a071. [DOI] [PubMed] [Google Scholar]
  16. Katze J. R. Alterations in SVT2 cell transfer RNAs in response to cell density and serum type. Biochim Biophys Acta. 1975 Mar 10;383(2):131–139. doi: 10.1016/0005-2787(75)90254-3. [DOI] [PubMed] [Google Scholar]
  17. Katze J. R., Beck W. T. Administration of queuine to mice relieves modified nucleoside queuosine deficiency in Ehrlich ascites tumor tRNA. Biochem Biophys Res Commun. 1980 Sep 16;96(1):313–319. doi: 10.1016/0006-291x(80)91216-4. [DOI] [PubMed] [Google Scholar]
  18. Katze J. R., Mosteller R. D. Inhibition of nucleoside Q formation in transfer ribonucleic acid during methionine starvation of relaxed-control Escherichia coli. J Bacteriol. 1976 Jan;125(1):205–210. doi: 10.1128/jb.125.1.205-210.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  20. Landin R. M., Boisnard M., Petrissant G. Correlation between the presence of tRNA His GUG and the erythropoietic function in foetal sheep liver. Nucleic Acids Res. 1979 Nov 24;7(6):1635–1648. doi: 10.1093/nar/7.6.1635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Laskey R. A., Mills A. D. Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem. 1975 Aug 15;56(2):335–341. doi: 10.1111/j.1432-1033.1975.tb02238.x. [DOI] [PubMed] [Google Scholar]
  22. Lockard R. E., Alzner-Deweerd B., Heckman J. E., MacGee J., Tabor M. W., RajBhandary U. L. Sequence analysis of 5'[32P] labeled mRNA and tRNA using polyacrylamide gel electrophoresis. Nucleic Acids Res. 1978 Jan;5(1):37–56. doi: 10.1093/nar/5.1.37. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Okada N., Nishimura S. Enzymatic synthesis of Q nucleoside containing mannose in the anticodon of tRNA: isolation of a novel mannosyltransferase from a cell-free extract of rat liver. Nucleic Acids Res. 1977 Aug;4(8):2931–2938. doi: 10.1093/nar/4.8.2931. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Okada N., Noguchi S., Kasai H., Shindo-Okada N., Ohgi T., Goto T., Nishimura S. Novel mechanism of post-transcriptional modification of tRNA. Insertion of bases of Q precursors into tRNA by a specific tRNA transglycosylase reaction. J Biol Chem. 1979 Apr 25;254(8):3067–3073. [PubMed] [Google Scholar]
  25. Okada N., Shindo-Okada N., Sato S., Itoh Y. H., Oda K., Nishimura S. Detection of unique tRNA species in tumor tissues by Escherichia coli guanine insertion enzyme. Proc Natl Acad Sci U S A. 1978 Sep;75(9):4247–4251. doi: 10.1073/pnas.75.9.4247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Paterson R., Knight C. A. Protein synthesis in tobacco protoplasts infected with tobacco mosaic virus. Virology. 1975 Mar;64(1):10–22. doi: 10.1016/0042-6822(75)90074-4. [DOI] [PubMed] [Google Scholar]
  27. Pelham H. R., Jackson R. J. An efficient mRNA-dependent translation system from reticulocyte lysates. Eur J Biochem. 1976 Aug 1;67(1):247–256. doi: 10.1111/j.1432-1033.1976.tb10656.x. [DOI] [PubMed] [Google Scholar]
  28. Pelham H. R. Leaky UAG termination codon in tobacco mosaic virus RNA. Nature. 1978 Mar 30;272(5652):469–471. doi: 10.1038/272469a0. [DOI] [PubMed] [Google Scholar]
  29. Roberts B. E., Paterson B. M. Efficient translation of tobacco mosaic virus RNA and rabbit globin 9S RNA in a cell-free system from commercial wheat germ. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2330–2334. doi: 10.1073/pnas.70.8.2330. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Scalla R., Romaine P., Asselin A., Rigaud J., Zaitlin M. An in vivo study of a nonstructural polypeptide synthesize upon TMV infection and its identification with a polypeptide synthesized in vitro from TMV RNA. Virology. 1978 Nov;91(1):182–193. doi: 10.1016/0042-6822(78)90366-5. [DOI] [PubMed] [Google Scholar]
  31. Shindo-Okada N., Terada M., Nishimura S. Changes in amount of hypo-modified tRNA having guanine in place of queuine during erythroid differentiation of murine erythroleukemia cells. Eur J Biochem. 1981 Apr;115(2):423–428. doi: 10.1111/j.1432-1033.1981.tb05254.x. [DOI] [PubMed] [Google Scholar]
  32. Siegel A., Hari V., Kolacz K. The effect of tobacco mosaic virus infection on host and virus-specific protein synthesis in protoplasts. Virology. 1978 Apr;85(2):494–503. doi: 10.1016/0042-6822(78)90456-7. [DOI] [PubMed] [Google Scholar]
  33. Stanley J., Vassilenko S. A different approach to RNA sequencing. Nature. 1978 Jul 6;274(5666):87–89. doi: 10.1038/274087a0. [DOI] [PubMed] [Google Scholar]
  34. Söll D. Enzymatic modification of transfer RNA. Science. 1971 Jul 23;173(3994):293–299. doi: 10.1126/science.173.3994.293. [DOI] [PubMed] [Google Scholar]
  35. Topal M. D., Fresco J. R. Base pairing and fidelity in codon-anticodon interaction. Nature. 1976 Sep 23;263(5575):289–293. doi: 10.1038/263289a0. [DOI] [PubMed] [Google Scholar]
  36. Tyc K., Kikuchi Y., Konarska M., Filipowicz W., Gross H. J. Ligation of endogenous tRNA 3' half molecules to their corresponding 5' halves via 2'-phosphomonoester,3',5'-phosphodiester bonds in extracts of Chlamydomonas. EMBO J. 1983;2(4):605–610. doi: 10.1002/j.1460-2075.1983.tb01470.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Vold B. S. Post-transcriptional modifications of the anticodon loop region: alterations in isoaccepting species of tRNA's during development in Bacillus subtilis. J Bacteriol. 1978 Jul;135(1):124–132. doi: 10.1128/jb.135.1.124-132.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Walker R. T., RajBhandary U. L. The nucleotide sequence of formylmethionine tRNA from Mycoplasma mycoides sp. capri. Nucleic Acids Res. 1978 Jan;5(1):57–70. doi: 10.1093/nar/5.1.57. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Ward D. C., Reich E. Conformational properties of polyformycin: a polyribonucleotide with individual residues in the syn conformation. Proc Natl Acad Sci U S A. 1968 Dec;61(4):1494–1501. doi: 10.1073/pnas.61.4.1494. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. White B. N., Tener G. M. Activity of a transfer RNA modifying enzyme during the development of Drosophila and its relationship to the su(s) locus. J Mol Biol. 1973 Mar 15;74(4):635–651. doi: 10.1016/0022-2836(73)90054-5. [DOI] [PubMed] [Google Scholar]
  41. Zagórski W. Translational regulation of expression of the brome-mosaic-virus RNA genome in vitro. Eur J Biochem. 1978 May 16;86(2):465–472. doi: 10.1111/j.1432-1033.1978.tb12329.x. [DOI] [PubMed] [Google Scholar]

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