1887

Abstract

In 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 gene is shown to be the first gene of a six-gene operon () that encodes proteins predicted to be components of an oxidoreductase. Mapping the transcript start and site-directed mutagenesis confirmed that the 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 expression. Analysis of the DNA sequence of the promoter region (P) 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 repression, 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–P 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 promoter containing the −16 and +15 heptamers was recognized by NarL , mutation of these heptamers did not affect NarL-mediated repression .

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2008-02-01
2019-10-14
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