FIGURE 1.

In vivo sensitivity of FNR to NO. A, dynamics of FNR-dependent in vivo transcription when E. coli cultures were exposed to NO, nitrite, or O₂. Anaerobic cultures of E. coli with a chromosomal FNR-dependent lacZ reporter were exposed to anaerobic water (filled diamonds); a mixture of the NO-releasing compounds (NOC-5 and NOC-7, 10 μm each; release of NO is indicated by the dashed line) as a source of exogenous NO (filled squares); nitrite, 10 mm, as a source of endogenous NO (open squares); or aeration (filled triangles). The cultures were sampled at the indicated times, and the relative transcription of lacZ, which is entirely dependent on FNR in this experiment, was quantified by quantitative RT-PCR. The abundance of the lacZ mRNA at t = 0 min was set at 1.0 and normalized to the housekeeping gyrA mRNA. Data from a typical experiment are shown. B, effects of increasing NO concentrations on FNR-dependent in vivo transcription. The experimental conditions were as described above except that cultures were amended with mixtures of NOC-5 and NOC-7 (0, 0.33, 0.66, 1, and 2 μm of each NOC) for 15 min at which point the cultures were sampled for the lacZ quantitative RT-PCR measurements. The concentration of NO released from the NOC mixtures after 15 min was calculated from the measured half-lives of NOC-5 and NOC-7. Data are mean values (n = 3) with standard deviations indicated.