Characterization of the Escherichia coli K-12 ydhYVWXUT operon

regulation by FNR, NarL and NarP

Jonathan D. Partridge, Douglas F. Browning, Meng Xu, Louise J. Newnham, Colin Scott, Ruth E. Roberts, Robert K. Poole, Jeffrey Green*

*Corresponding author for this work

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

In Escherichia coli K-12 the expression of many genes is controlled by the oxygen-responsive transcription factor FNR and the nitrate- and nitrite-responsive two-component systems NarXL and NarPQ. Here, the ydhY gene is shown to be the first gene, of a six-gene operon (ydhYVWXUT) that encodes proteins predicted to be components of an oxidoreductase. Mapping the ydhY-T transcript start and site-directed mutagenesis confirmed that the ydhY-T genes are transcribed from an FNR-dependent class II promoter and showed that the FNR site is centred at -42.5. In the presence of nitrate or nitrite, NarXL and NarPQ repressed ydhY-T expression. Analysis of the DNA sequence of the ydhY promoter region (PydhY) revealed the presence of four heptameric sequences similar to NarL/P binding sites centred at -42, -16, +6 and +15. The latter heptamers are arranged as a 7-2-7 inverted repeat, which is required for recognition by NarP. Accordingly, NarP protected the 7-2-7 region in DNase I footprints, and mutation of either heptamer +6 or heptamer +15 impaired nitrite-mediated regression, whereas mutation of heptamer -42 and heptamer -16 did not affect the response to nitrite. The NarL protein also protected the 7-2-7 region, but in contrast to NarP, the NarL footprint extended further upstream to encompass the -16 heptamer. The extended NarL footprint was consistent with the presence of multiple NarL-PydhY complexes in gel retardation assays. Mutation of heptamer -42, which is located within the FNR binding site, or heptamer + 6 (but not heptamers -16 or +15) impaired nitrate-mediated repression. Thus, although the region of the ydhY-T promoter containing the -16 and +15 heptamers was recognized by NarL in vitro, mutation of these heptamers did not affect NarL-mediated repression in vivo.

Original languageEnglish
Pages (from-to)608-618
Number of pages11
JournalMicrobiology
Volume154
Issue number2
DOIs
Publication statusPublished - Feb 2008
Externally publishedYes

    Fingerprint

Cite this

Partridge, J. D., Browning, D. F., Xu, M., Newnham, L. J., Scott, C., Roberts, R. E., ... Green, J. (2008). Characterization of the Escherichia coli K-12 ydhYVWXUT operon: regulation by FNR, NarL and NarP. Microbiology, 154(2), 608-618. https://doi.org/10.1099/mic.0.2007/012146-0