Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1984 Feb 10;12(3):1377–1390. doi: 10.1093/nar/12.3.1377

The in vivo and in vitro initiation site for transcription of the rRNA operon of Saccharomyces carlsbergensis.

J Klootwijk, M P Verbeet, G M Veldman, V C de Regt, H van Heerikhuizen, J Bogerd, R J Planta
PMCID: PMC318583  PMID: 6322105

Abstract

We have performed a detailed analysis of the transcription initiation of the rRNA operon in the yeast Saccharomyces carlsbergensis. Electron microscopic analysis of R-looped pre-rRNA molecules together with a very sensitive S1-nuclease mapping showed the use of only a single transcription start at about 700 bp upstream of the 17S rRNA gene and not of the minor start sites proposed for the very closely related species S. cerevisiae by others [Bayev et al. (5), Swanson and Holland (6)]. The sequence of 730 bp of the initiating region is presented. In vitro transcription in concentrated lysates of yeast spheroplasts in the presence of (gamma-SH)ATP or (gamma-SH)GTP, followed by purification of the in vitro initiated RNA via Hg-agarose, revealed that on the endogenous template exactly the same site is used for transcription initiation as in vivo.

Full text

PDF
1388

Images in this article

1381Image
on p.1381
1383Image
on p.1383
1385Image
on p.1385
1386Image
on p.1386

Selected References

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

  1. Bayev A. A., Georgiev O. I., Hadjiolov A. A., Kermekchiev M. B., Nikolaev N., Skryabin K. G., Zakharyev V. M. The structure of the yeast ribosomal RNA genes. 2. The nucleotide sequence of the initiation site for ribosomal RNA transcription. Nucleic Acids Res. 1980 Nov 11;8(21):4919–4926. doi: 10.1093/nar/8.21.4919. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bell G. I., DeGennaro L. J., Gelfand D. H., Bishop R. J., Valenzuela P., Rutter W. J. Ribosomal RNA genes of Saccharomyces cerevisiae. I. Physical map of the repeating unit and location of the regions coding for 5 S, 5.8 S, 18 S, and 25 S ribosomal RNAs. J Biol Chem. 1977 Nov 25;252(22):8118–8125. [PubMed] [Google Scholar]
  3. Clewell D. B., Helinski D. R. Properties of a supercoiled deoxyribonucleic acid-protein relaxation complex and strand specificity of the relaxation event. Biochemistry. 1970 Oct 27;9(22):4428–4440. doi: 10.1021/bi00824a026. [DOI] [PubMed] [Google Scholar]
  4. Hindley J., Page S. M. Nucleotide sequence of yeast 5 S ribosomal RNA. FEBS Lett. 1972 Oct 1;26(1):157–160. doi: 10.1016/0014-5793(72)80563-5. [DOI] [PubMed] [Google Scholar]
  5. Hipskind R. A., Reeder R. H. Initiation of ribosomal RNA chains in homogenates of oocyte nuclei. J Biol Chem. 1980 Aug 25;255(16):7896–7906. [PubMed] [Google Scholar]
  6. Ide G. J. Nucleoside 5'-[gamma-S]triphosphates will initiate transcription in isolated yeast nuclei. Biochemistry. 1981 Apr 28;20(9):2633–2638. doi: 10.1021/bi00512a042. [DOI] [PubMed] [Google Scholar]
  7. Inman J. K., Dintzis H. M. The derivatization of cross-linked polyacrylamide beads. Controlled introduction of functional groups for the preparation of special-purpose, biochemical adsorbents. Biochemistry. 1969 Oct;8(10):4074–4082. doi: 10.1021/bi00838a026. [DOI] [PubMed] [Google Scholar]
  8. Klekamp M. S., Weil P. A. Specific transcription of homologous class III genes in yeast-soluble cell-free extracts. J Biol Chem. 1982 Jul 25;257(14):8432–8441. [PubMed] [Google Scholar]
  9. Klemenz R., Geiduschek E. P. The 5' terminus of the precursor ribosomal RNA of Saccharomyces cerevisiae. Nucleic Acids Res. 1980 Jun 25;8(12):2679–2689. doi: 10.1093/nar/8.12.2679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Klootwijk J., de Jonge P., Planta R. J. The primary transcript of the ribosomal repeating unit in yeast. Nucleic Acids Res. 1979 Jan;6(1):27–39. doi: 10.1093/nar/6.1.27. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Maxam A. M., Gilbert W. A new method for sequencing DNA. Proc Natl Acad Sci U S A. 1977 Feb;74(2):560–564. doi: 10.1073/pnas.74.2.560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Moss T., Boseley P. G., Birnstiel M. L. More ribosomal spacer sequences from Xenopus laevis. Nucleic Acids Res. 1980 Feb 11;8(3):467–485. doi: 10.1093/nar/8.3.467. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Reeve A. E., Smith M. M., Pigiet V., Huang R. C. Incorporation of purine nucleoside 5'-[gamma-S]triphosphates as affinity probes for initiation of RNA synthesis in vitro. Biochemistry. 1977 Oct 4;16(20):4464–4470. doi: 10.1021/bi00639a021. [DOI] [PubMed] [Google Scholar]
  14. Skriabin K. G., Zakhar'ev V. M., Rubtsov P. M., Baev A. A. Posledovatel'nost' nukleotidov predpolagaemoi oblasti initsiatsii transkriptsii ribosomnogo operona drozhzhei. Dokl Akad Nauk SSSR. 1979;247(5):1275–1277. [PubMed] [Google Scholar]
  15. Sluyterman L. A., Wijdenes J. An agarose mercurial column for the separation of mercaptopapain and nonmercaptopapain. Biochim Biophys Acta. 1970 Mar 31;200(3):593–595. doi: 10.1016/0005-2795(70)90122-4. [DOI] [PubMed] [Google Scholar]
  16. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  17. Swanson M. E., Holland M. J. RNA polymerase I-dependent selective transcription of yeast ribosomal DNA. Identification of a new cellular ribosomal RNA precursor. J Biol Chem. 1983 Mar 10;258(5):3242–3250. [PubMed] [Google Scholar]
  18. Tekamp P. A., Valenzuela P., Maynard T., Bell G. I., Rutter W. J. Specific gene transcription in yeast nuclei and chromatin by added homologous RNA polymerases I and II. J Biol Chem. 1979 Feb 10;254(3):955–963. [PubMed] [Google Scholar]
  19. Valenzuela P., Bell G. I., Venegas A., Sewell E. T., Masiarz F. R., DeGennaro L. J., Weinberg F., Rutter W. J. Ribosomal RNA genes of Saccharomyces cerevisiae. II. Physical map and nucleotide sequence of the 5 S ribosomal RNA gene and adjacent intergenic regions. J Biol Chem. 1977 Nov 25;252(22):8126–8135. [PubMed] [Google Scholar]
  20. Verbeet M. P., Klootwijk J., van Heerikhuizen H., Fontijn R., Vreugdenhil E., Planta R. J. Molecular cloning of the rDNA of Saccharomyces rosei and comparison of its transcription initiation region with that of Saccharomyces carlsbergensis. Gene. 1983 Jul;23(1):53–63. doi: 10.1016/0378-1119(83)90216-0. [DOI] [PubMed] [Google Scholar]
  21. van Keulen H., Thomas D. Y. A yeast transcription system for the 5S rRNA gene. Nucleic Acids Res. 1982 Sep 11;10(17):5223–5238. doi: 10.1093/nar/10.17.5223. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES