Molecular and General Genetics MGG

, Volume 210, Issue 1, pp 140–144 | Cite as

Positional requirements for the function of nif-specific upstream activator sequences

  • Martin Buck
  • Joanna Woodcock
  • Wendy Cannon
  • Lesley Mitchenall
  • Martin Drummond


The upstream activator sequence (UAS) found in Klebsiella pneumoniae nif promoters and required for the activation of transcription by nifA, is absent from the nifF-nifL intergenic region, but is present downstream from the nifLA transcription start at+59. To determine whether nif upstream activator sequences can function in a 3′ position, the nifH UAS was cloned downstream from the NifH transcription start, but no activation of transcription by nifA dependent upon the UAS in its 3′ location could be detected. A mild repressive effect was detectable when the nifH UAS was placed downstream of the nifH promoter, but not when the cat promoter was substituted for the nifLA promoter upstream from the motif at+59 described above. However, deletion analysis showed that the UAS motif located downstream of the nifLA promoter has a role in transcription from the nifF promoter, although it is situated at position-263 with respect to the nifF transcription start, about 100 bp further upstream than previously described occurrences of the activator sequence.

Key words

Transcriptional Activation Upstream sequences 


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  1. Ausubel FM (1984) Regulation of nitrogen fixation genes. Cell 37:5–6Google Scholar
  2. Besse J, von Wilcken-Bergmann B, Müller-Hill B (1986) Synthetic lac operator mediates repression through lac repressor when introduced upstream and downstream from lac promoter. EMBO J 5:1377–1381Google Scholar
  3. Beynon J, Cannon M, Buchanan-Wollaston V, Cannon F (1983) The nif promoters of Klebsiella pneumoniae have a characteristic primary structure. Cell 34:665–671Google Scholar
  4. Brown SE, Ausubel FM (1984) Mutations affecting regulation of the Klebsiella pneumoniae nifH (nitrogenase reductase) promoter. J Bacteriol 157:143–147Google Scholar
  5. Buchanan-Wollaston V, Cannon FC (1984) In: Veeger C, Newton WE (eds) Advances in nitrogen fixation research. Nijhoff/Junk, p 732Google Scholar
  6. Buchanan-Wollaston V, Cannon MC, Beynon JC, Cannon FC (1981a) Role of the nifA gene product in the regulation of nif expression in Klebsiella pneumoniae. Nature 294:776–776Google Scholar
  7. Buchanan-Wollaston V, Cannon MC, Cannon FC (1981b) The use of cloned nif (nitrogen fixation) DNA to investigate transcriptional regulation of nif expression in Klebsiella pneumoniae. Mol Gen Genet 184:102–106Google Scholar
  8. Buck M, Cannon W (1987) Frameshifts close to the Klebsiella pneumoniae nifH promoter prevent multicopy inhibition by hybrid nifH plasmids. Mol Gen Genet 207:492–498Google Scholar
  9. Buck M, Khan H, Dixon R (1985) Site directed mutagenesis of the Klebsiella pneumoniae nifL and nifH promoters and in vivo analysis of promoter activities. Nucleic Acids Res 13:7621–7638Google Scholar
  10. Buck M, Miller S, Drummond M, Dixon R (1986) Upstream activator sequences are present in the promoters of nitrogen fixation genes. Nature 320:374–378Google Scholar
  11. Buck M, Woodcock JM, Cannon W (1987) Mol Microbiol (in press)Google Scholar
  12. Buikema WJ, Szeto WW, Lemley PV, Orme-Johnson WH, Ausubel FM (1985) Nitrogen fixation specific regulatory genes of Klebsiella pneumoniae and Rhizobium meliloti share homology with the general nitrogen regulatory gene ntrC of K. pneumoniae. Nucleic Acids Res 13:4539–4555Google Scholar
  13. Cannon M, Hill S, Kavanagh E, Cannon F (1985) A molecular genetic study of nif expression in Klebsiella pneumoniae at the level of transcription translation and nitrogenase activity. Mol Gen Genet 198:198–206Google Scholar
  14. Casadaban MJ, Chou J, Cohen SN (1980) In vitro gene fusions that join an enzymatically active β-galactosidase segment to amino-terminal fragments of exogenous proteins: Escherichia coli plasmid vectors for the detection and cloning of translation initiation signals. J Bacteriol 143:971–980Google Scholar
  15. Drummond M, Clements J, Merrick M, Dixon R (1983) Positive control and autogenous regulation of the nifLA promoter in Klebsiella pneumoniae. Nature 301:302–307Google Scholar
  16. Drummond M, Whitty P, Wootton J (1986) Sequence and domain relationships of ntrC and nifA from Klebsiella pneumoniae: homologies to other regulatory proteins. EMBO J 5:441–447Google Scholar
  17. Gussin GN, Ronson CW, Ausubel FM (1986) Regulation of nitrogen fixation genes. Annu Rev Genet 20:567–591Google Scholar
  18. Hirschman J, Wang P-K, Sei K, Keener J, Kustu S (1985) Products of nitrogen regulatory genes ntrA and ntrC of enteric bacteria activate glnA transcription in vitro: Evidence that the ntrA product is a σ factor. Proc Natl Acad Sci USA 82:7525–7529Google Scholar
  19. Hunt TP, Magasanik B (1985) Transcription of glnA by purified Escherichia coli components: Core RNA polymerase and the products of glnF, glnG and glnL. Proc Natl Acad Sci USA 82:8453–8457Google Scholar
  20. Merrick MJ (1983) Nitrogen control of the nif regulon in Klebsiella pneumoniae: involvement of the nitrA gene and analogies between ntrC and nifA. EMBO J 2:39–44Google Scholar
  21. Merrick M, Austin S, Buck M, Dixon R, Drummond M, Holtel A, MacFarlane S (1987) In: Phosphate metabolism in microorganisms. ASM publications (in press)Google Scholar
  22. Ow DW, Ausubel FM (1983) Regulation of nitrogen metabolism by nifA gene product in Klebsiella pneumoniae. Nature 301:307–313Google Scholar
  23. Ptashne M (1986) Gene regulation by proteins acting nearby and at a distance. Nature 322:697–701Google Scholar
  24. Reitzer LJ, Magasanik B (1986) Transcription of glnA in E. coli is stimulated by activator bound to sites far from the promoter. Cell 45:785–792Google Scholar
  25. Riedel GE, Ausubel FM, Cannon FC (1979) Physical map of chromosomal nitrogen fixation (nif) genes of Klebsiella pneumoniae. Proc Natl Acad Sci USA 76:2866–2870Google Scholar
  26. Riedel GE, Brown SE, Ausubel FM (1983) Nitrogen fixation by Klebsiella pneumoniae is inhibited by certain multicopy hybrid nif plasmids. J Bacteriol 153:45–56Google Scholar
  27. Struhl K (1987) Promoters, activator proteins and the mechanism of transcriptional initiation in yeast. Cell 49:295–297Google Scholar

Copyright information

© Springer-Verlag 1987

Authors and Affiliations

  • Martin Buck
    • 1
  • Joanna Woodcock
    • 1
  • Wendy Cannon
    • 1
  • Lesley Mitchenall
    • 1
  • Martin Drummond
    • 1
  1. 1.AFRC Unit of Nitrogen FixationUniversity of SussexBrightonUK

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