Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1972 Feb;69(2):407–411. doi: 10.1073/pnas.69.2.407

In Vitro Synthesis of Ribosomal RNA by Bacillus subtilis RNA Polymerase

Caroline Hussey 1,*, Janice Pero 1, Rosalind G Shorenstein 1, Richard Losick 1
PMCID: PMC426468  PMID: 4621834

Abstract

Two kinds of hybridization competition experiments show that Bacillus subtilis RNA polymerase synthesizes ribosomal RNA (rRNA) in vitro with B. subtilis DNA as a template. First, RNA synthesized in vitro competes with the hybridization of [32P]rRNA synthesized in vivo to the heavy strand of B. subtilis DNA. Second, unlabeled rRNA synthesized in vivo competes with the hybridization of [3H]RNA synthesized in vitro to denatured DNA or heavy-strand DNA, but not to light-strand DNA. The ability of RNA polymerase holoenzyme to synthesize rRNA in vitro is not lost after extensive purification. RNA polymerase core enzyme, however, which is missing the σ factor, synthesizes little rRNA in vitro.

RNA polymerase purified from wild-type sporulating cells synthesizes little rRNA in vitro, while the in vitro synthesis of rRNA by RNA polymerase from stationary phase cells of the sporulation-defective mutant rfr 10 is apparently unimpaired.

Keywords: sporulation, hybridization, heavy strand of DNA

Full text

PDF

Selected References

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

  1. Babinet C. A new method for the purification of RNA-polymerase. Biochem Biophys Res Commun. 1967 Mar 21;26(6):639–644. doi: 10.1016/s0006-291x(67)80119-0. [DOI] [PubMed] [Google Scholar]
  2. Gillespie D., Spiegelman S. A quantitative assay for DNA-RNA hybrids with DNA immobilized on a membrane. J Mol Biol. 1965 Jul;12(3):829–842. doi: 10.1016/s0022-2836(65)80331-x. [DOI] [PubMed] [Google Scholar]
  3. Hussey C., Losick R., Sonenshein A. L. Ribosomal RNA synthesis is turned off during sporulation of Bacillus subtilis. J Mol Biol. 1971 Apr 14;57(1):59–70. doi: 10.1016/0022-2836(71)90119-7. [DOI] [PubMed] [Google Scholar]
  4. Kennel D. Titration of the gene sites on DNA by DNA-RNA hybridization. II. The Escherichia coli chromosome. J Mol Biol. 1968 May 28;34(1):85–103. doi: 10.1016/0022-2836(68)90236-2. [DOI] [PubMed] [Google Scholar]
  5. Losick R., Shorenstein R. G., Sonenshein A. L. Structural alteration of RNA polymerase during sporulation. Nature. 1970 Aug 29;227(5261):910–913. doi: 10.1038/227910a0. [DOI] [PubMed] [Google Scholar]
  6. Losick R., Sonenshein A. L. Change in the template specificity of RNA polymerase during sporulation of Bacillus subtilis. Nature. 1969 Oct 4;224(5214):35–37. doi: 10.1038/224035a0. [DOI] [PubMed] [Google Scholar]
  7. Margulies L., Remeza V., Rudner R. Asymmetric template function of microbial deoxyribonucleic acids: transcription of ribosomal and soluble ribonucleic acids. J Bacteriol. 1970 Sep;103(3):560–568. doi: 10.1128/jb.103.3.560-568.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Oishi M. The transcribing strands of bacillus subtilis DNA for ribosomal and transfer RNA. Proc Natl Acad Sci U S A. 1969 Jan;62(1):256–262. doi: 10.1073/pnas.62.1.256. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Pettijohn D. E., Clarkson K., Kossman C. R., Stonington O. G. Synthesis of ribosomal RNA on a protein-DNA complex isolated from bacteria: a comparison of ribosomal RNA synthesis in vitro and in vivo. J Mol Biol. 1970 Sep 14;52(2):281–300. doi: 10.1016/0022-2836(70)90031-8. [DOI] [PubMed] [Google Scholar]
  10. Rudner R., Karkas J. D., Chargaff E. Separation of B. subtilis DNA into complementary strands, I. Biological properties. Proc Natl Acad Sci U S A. 1968 Jun;60(2):630–635. doi: 10.1073/pnas.60.2.630. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. SAITO H., MIURA K. I. PREPARATION OF TRANSFORMING DEOXYRIBONUCLEIC ACID BY PHENOL TREATMENT. Biochim Biophys Acta. 1963 Aug 20;72:619–629. [PubMed] [Google Scholar]
  12. Salser W., Janin J., Levinthal C. Measurement of the unstable RNA in exponentially growing cultures of Bacillus subtilis and Escherichia coli. J Mol Biol. 1968 Jan 28;31(2):237–266. doi: 10.1016/0022-2836(68)90442-7. [DOI] [PubMed] [Google Scholar]
  13. Solymosy F., Fedorcsák I., Gulyás A., Farkas G. L., Ehrenberg L. A new method based on the use of diethyl pyrocarbonate as a nuclease inhibitor for the extraction of undegraded nucleic acid from plant tissues. Eur J Biochem. 1968 Sep 24;5(4):520–527. doi: 10.1111/j.1432-1033.1968.tb00401.x. [DOI] [PubMed] [Google Scholar]
  14. Sonenshein A. L., Losick R. RNA polymerase mutants blocked in sporulation. Nature. 1970 Aug 29;227(5261):906–909. doi: 10.1038/227906a0. [DOI] [PubMed] [Google Scholar]
  15. Travers A. A., Kamen R. I., Schleif R. F. Factor necessary for ribosomal RNA synthesis. Nature. 1970 Nov 21;228(5273):748–751. doi: 10.1038/228748a0. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES