Molecules and Cells

Korean Society for Molecular and Cellular Biology (한국분자세포생물학회)
권호 표지

Effects of Base Changes at the Transcription Start Site on Stringent Control of rnpB in Escherichia coli

  • Choi, Hyun-Sook (Department of Chemistry and Center for Molecular Design and Synthesis, Korea Advanced Institute of Science and Technology) ;
  • Park, Jeong Won (Department of Chemistry and Center for Molecular Design and Synthesis, Korea Advanced Institute of Science and Technology) ;
  • Hong, Soon Kang (Department of Fire Service Administration, Chodang University) ;
  • Lee, Kangseok (Department of Life Science, Chung-Ang University) ;
  • Lee, Younghoon (Department of Chemistry and Center for Molecular Design and Synthesis, Korea Advanced Institute of Science and Technology)
  • Received : 2008.02.01
  • Accepted : 2008.03.10
  • Published : 2008.08.31
⟨ Previous Next ⟩

Abstract

The GC-rich discriminator sequence between the -10 region and the transcription start site of the rnpB promoter is responsible for stringent control of M1 RNA synthesis. The rnpB promoter also contains a G nucleotide at the previously identified transcription start site. In this study, we examined by mutagenesis of G to A whether this +1G nucleotide is involved in the stringent response. We found that the change did not alter the stringent response. Since the +1 mutation might alter transcription initiation, we compared the transcription start sites of the wt and mutant promoters by primer extension analysis. Surprisingly, we found that wild type rnpB transcription starts at both the +1G position (70%) and the -1C position (30%), and that the +1A mutation led to transcription initiation exclusively at the -1C position. We also generated two transversion mutations at the -1 position, both of which led to transcription starting exclusively at that position. The -1G mutant promoter gave a stringent signal similar to the wild-type, whereas the -1A mutant generated a significantly less stringent signal. Base on these results, we propose that a short sequence, up to 7 bp downstream of the -10 region, is involved in the stringent response of the rnpB promoter.

Keywords

Acknowledgement

Supported by : Korea Science and Engineering Foundation

References

  1. Cashel, M., Gentry, D.R., Hernandez, J.V., and Vinella, D. (1996). The stringent response in Escherichia coli and Satmonellar. In Cellular and Molecular Biology, F.C. Neidhardt, ed. (Washington, DC., USA: Am. Soc. Microbiol.), pp. 1458-1496.
  2. Choi, H.S., Kim, K.S., Park, J.W., Jung, Y.H., and Lee, Y. (2005). Effects of FIS protein rnpB transcription in Escherichia coli. Mol. Cells 19, 239-245.
  3. Guerrier-Takada, C., Gardiner, K., Marsh, T., Pace, N., and Altman, S. (1983). The RNA moiety of ribonuclease P is the catalytic subunit of the enzyme. Cell 35, 849-857. https://doi.org/10.1016/0092-8674(83)90117-4
  4. Harley, C.B., and Reynolds, R.P. (1987). Analysis of E. coli promoter sequences. Nucleic Acids Res. 15, 2343-2361. https://doi.org/10.1093/nar/15.5.2343
  5. Hawley, D.K., and McClure, W.R. (1983). Compilation and analysis of Escherichia coli promoter DNA sequences. Nucleic Acids Res. 11, 2237-2255. https://doi.org/10.1093/nar/11.8.2237
  6. Hershberg, R., Bejerano, G., Santos-Zavaleta, A., and Margalit, H. (2001). PromEC: An updated database of Escherichia coli mRNA promoters with experimentally identified transcriptional start sites. Nucleic Acids Res. 29, 277. https://doi.org/10.1093/nar/29.1.277
  7. Jeong, W., and Kang, C. (1994). Start site selection at lacUV5 promoter affected by the sequence context around the initiation sites. Nucleic Acids Res. 22, 4667-4672. https://doi.org/10.1093/nar/22.22.4667
  8. Josaitis, C.A., Gaal, T., and Gourse, R.L. (1995). Stringent control and growth-rate-dependent control have nonidentical promoter sequence requirements. Proc. Natl. Acad. Sci. USA 92, 1117-1121.
  9. Jung, Y.H., and Lee, Y. (1997). Escherichia coli rnpB promoter mutants altered in stringent response. Biochem. Biophys. Res. Commun. 230, 582-586. https://doi.org/10.1006/bbrc.1996.6005
  10. Kim, S., Kim, H., Park, I., and Lee, Y. (1996). Mutational analysis of RNA structures and sequences postulated to affect 3' processing of M1 RNA, the RNA component of Escherichia coli RNase P. J. Biol. Chem. 271, 19330-19337. https://doi.org/10.1074/jbc.271.32.19330
  11. Kim, K.S., Sim, S., Ko, J.H., and Lee, Y. (2005). Processing of M1 RNA at the 3′ end protects its primary transcript from degradation. J. Biol. Chem. 280, 34667-34674. https://doi.org/10.1074/jbc.M505005200
  12. Lamond, A.I., and Travers, A.A. (1985). Genetically separable functional elements mediate the optimal expression and stringent regulation of a bacterial tRNA gene. Cell 40, 319-326. https://doi.org/10.1016/0092-8674(85)90146-1
  13. Lee, S.J., Jung, Y.H., Park, C.-U., and Lee, Y. (1991). Stringent control of Escherichia coli rnpB gene transcription in vivo in multicopy plasmids. Mol. Cells 1, 415-420.
  14. Liu, J., and Turnbough, C.L., Jr. (1994). Effects of transcriptional start site sequence and position on nucleotide-sensitive selection of alternative start sites at the éóê` promoter in Escherichia coli. J. Bacteriol. 176, 2938-2945. https://doi.org/10.1128/jb.176.10.2938-2945.1994
  15. Maitra, U., Cohen, S.N., and Hurwitz, J. (1966). Specificity of initiation and synthesis of RNA from DNA templates. Cold Spring Harb. Symp. Quant. Biol. 31, 113-122.
  16. Metzger, S., Dror, I.B., Aizenman, E., Schreiber, G., Toone, M., Friesen, J. D., Cashel, M., and Glaser, G. (1988). The nucleotide sequence and characterization of the refA gene of bëJ Escherichia coli. J. Biol. Chem. 263, 15699-15704.
  17. Park, J.W., Jung, Y., Lee, S.J., Jin, D.J., and Lee, Y. (2002). Alteration of stringent response of the Escherichia coli rnpB promoter by mutations in the -35 region. Biochem. Biophys. Res. Commun. 290, 1183-1187. https://doi.org/10.1006/bbrc.2001.6331
  18. Robertson, H.D., Altman, S., and Smith, J.D. (1972). Purification and properties of a specific Escherichia coli ribonuclease which cleaves a tyrosine transfer ribonucleic acid presursor. J. Biol. Chem. 247, 5243-5251.
  19. Sung, Y.H., Kim, H.J., and Lee, H.W. (2007). Identification of a novel Rb-regulated gene associated with the cell cycle. Mol. Cells 24, 409-415.
  20. Travers, A.A. (1980a). Promoter sequence for stringent control of bacterial ribonucleic acid synthesis. J. Bacteriol. 141, 973-976.
  21. Travers, A.A. (1980b). A tRNATyr promoter with an altered in vitro response to ppgpp. J. Mol. Biol. 141, 91-97. https://doi.org/10.1016/S0022-2836(80)80030-1
  22. Zacharias, M., Goringer, H.U., and Wagner, R. (1989). Influence of the GCGC discriminator motif introduced into the ribosomal RNA P2- and ttac promoter on growth-rate control and stringent sensitivity. EMBO J. 8, 3357-3363.