Skip to main content
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1985 Jan 25;13(2):435–447. doi: 10.1093/nar/13.2.435

Point mutations in the 5' ICR and anticodon region of a Drosophila tRNAArg gene decrease in vitro transcription.

T S Stewart, D Söll, S Sharp
PMCID: PMC341006  PMID: 3923433

Abstract

We have examined the effects of various nucleotide substitutions in a Drosophila tRNAArg gene on in vitro transcription and stable transcription complex formation in Drosophila KcO and HeLa cell extracts. Substitutions in positions encoding the invariant G18 and G19 residues resulted in decreased transcription, however, the moderate decreases indicate that these nucleotides are not obligatory promoter recognition sites. An A21 to C21 mutation had no effect on transcription levels using homologous extract however, this mutant displayed decreased transcriptional abilities in HeLa cell extract. Nucleotide substitutions within the sequence encoding the anticodon led to a decrease in the transcription activity but not in the ability to form a stable transcription complex.

Full text

PDF
440

Images in this article

Selected References

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

  1. Allison D. S., Goh S. H., Hall B. D. The promoter sequence of a yeast tRNAtyr gene. Cell. 1983 Sep;34(2):655–664. doi: 10.1016/0092-8674(83)90398-7. [DOI] [PubMed] [Google Scholar]
  2. Cooley L., Appel B., Söll D. Post-transcriptional nucleotide addition is responsible for the formation of the 5' terminus of histidine tRNA. Proc Natl Acad Sci U S A. 1982 Nov;79(21):6475–6479. doi: 10.1073/pnas.79.21.6475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cortese R., Harland R., Melton D. Transcription of tRNA genes in vivo: single-stranded compared to double-stranded templates. Proc Natl Acad Sci U S A. 1980 Jul;77(7):4147–4151. doi: 10.1073/pnas.77.7.4147. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Dingermann T., Sharp S., Appel B., DeFranco D., Mount S., Heiermann R., Pongs O., Söll D. Transcription of cloned tRNA and 5S RNA genes in a Drosophila cell free extract. Nucleic Acids Res. 1981 Aug 25;9(16):3907–3918. doi: 10.1093/nar/9.16.3907. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Dingermann T., Sharp S., Schaack J., Söll D. Stable transcription complex formation of eukaryotic tRNA genes is dependent on a limited separation of the two intragenic control regions. J Biol Chem. 1983 Sep 10;258(17):10395–10402. [PubMed] [Google Scholar]
  6. Folk W. R., Hofstetter H. A detailed mutational analysis of the eucaryotic tRNAmet1 gene promoter. Cell. 1983 Jun;33(2):585–593. doi: 10.1016/0092-8674(83)90439-7. [DOI] [PubMed] [Google Scholar]
  7. Fuhrman S. A., Engelke D. R., Geiduschek E. P. HeLa cell RNA polymerase III transcription factors. Functional characterization of a fraction identified by its activity in a second template rescue assay. J Biol Chem. 1984 Feb 10;259(3):1934–1943. [PubMed] [Google Scholar]
  8. Galli G., Hofstetter H., Birnstiel M. L. Two conserved sequence blocks within eukaryotic tRNA genes are major promoter elements. Nature. 1981 Dec 17;294(5842):626–631. doi: 10.1038/294626a0. [DOI] [PubMed] [Google Scholar]
  9. Hanahan D. Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983 Jun 5;166(4):557–580. doi: 10.1016/s0022-2836(83)80284-8. [DOI] [PubMed] [Google Scholar]
  10. Hawley D. K., McClure W. R. Compilation and analysis of Escherichia coli promoter DNA sequences. Nucleic Acids Res. 1983 Apr 25;11(8):2237–2255. doi: 10.1093/nar/11.8.2237. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hofstetter H., Kressman A., Birnstiel M. L. A split promoter for a eucaryotic tRNA gene. Cell. 1981 May;24(2):573–585. doi: 10.1016/0092-8674(81)90348-2. [DOI] [PubMed] [Google Scholar]
  12. Laski F. A., Belagaje R., RajBhandary U. L., Sharp P. A. An amber suppressor tRNA gene derived by site-specific mutagenesis: cloning and function in mammalian cells. Proc Natl Acad Sci U S A. 1982 Oct;79(19):5813–5817. doi: 10.1073/pnas.79.19.5813. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lassar A. B., Martin P. L., Roeder R. G. Transcription of class III genes: formation of preinitiation complexes. Science. 1983 Nov 18;222(4625):740–748. doi: 10.1126/science.6356356. [DOI] [PubMed] [Google Scholar]
  14. Messing J. New M13 vectors for cloning. Methods Enzymol. 1983;101:20–78. doi: 10.1016/0076-6879(83)01005-8. [DOI] [PubMed] [Google Scholar]
  15. Newman A. J., Ogden R. C., Abelson J. tRNA gene transcription in yeast: effects of specified base substitutions in the intragenic promoter. Cell. 1983 Nov;35(1):117–125. doi: 10.1016/0092-8674(83)90214-3. [DOI] [PubMed] [Google Scholar]
  16. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Schaack J., Sharp S., Dingermann T., Burke D. J., Cooley L., Söll D. The extent of a eukaryotic tRNA gene. 5'- and 3'-flanking sequence dependence for transcription and stable complex formation. J Biol Chem. 1984 Feb 10;259(3):1461–1467. [PubMed] [Google Scholar]
  18. Sharp S., DeFranco D., Dingermann T., Farrell P., Söll D. Internal control regions for transcription of eukaryotic tRNA genes. Proc Natl Acad Sci U S A. 1981 Nov;78(11):6657–6661. doi: 10.1073/pnas.78.11.6657. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Sharp S., Dingermann T., Schaack J., DeFranco D., Söll D. Transcription of eukaryotic tRNA genes in vitro. I. Analysis of control regions using a competition assay. J Biol Chem. 1983 Feb 25;258(4):2440–2446. [PubMed] [Google Scholar]
  20. Stillman D. J., Geiduschek E. P. Differential binding of a S. cerevisiae RNA polymerase III transcription factor to two promoter segments of a tRNA gene. EMBO J. 1984 Apr;3(4):847–853. doi: 10.1002/j.1460-2075.1984.tb01895.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Stillman D. J., Sivertsen A. L., Zentner P. G., Geiduschek E. P. Correlations between transcription of a yeast tRNA gene and transcription factor-DNA interactions. J Biol Chem. 1984 Jun 25;259(12):7955–7962. [PubMed] [Google Scholar]
  22. Temple G. F., Dozy A. M., Roy K. L., Kan Y. W. Construction of a functional human suppressor tRNA gene: an approach to gene therapy for beta-thalassaemia. Nature. 1982 Apr 8;296(5857):537–540. doi: 10.1038/296537a0. [DOI] [PubMed] [Google Scholar]
  23. Traboni C., Ciliberto G., Cortese R. A novel method for site-directed mutagenesis: its application to an eukaryotic tRNAPro gene promoter. EMBO J. 1982;1(4):415–420. doi: 10.1002/j.1460-2075.1982.tb01184.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Traboni C., Ciliberto G., Cortese R. Mutations in Box B of the promoter of a eucaryotic tRNAPro gene affect rate of transcription, processing, and stability of the transcripts. Cell. 1984 Jan;36(1):179–187. doi: 10.1016/0092-8674(84)90087-4. [DOI] [PubMed] [Google Scholar]
  25. Zoller M. J., Smith M. Oligonucleotide-directed mutagenesis using M13-derived vectors: an efficient and general procedure for the production of point mutations in any fragment of DNA. Nucleic Acids Res. 1982 Oct 25;10(20):6487–6500. doi: 10.1093/nar/10.20.6487. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES