**5. Conclusions**

Figure 7 summarizes the effects of external factors and regulatory proteins on *isc-suf* expression and also indicates the influence of the P1, P5, and the P4 promoter region on the activity of P2, which is the main promoter for *iscRS* expression but also regulates other genes of iron metabolism. Our study revealed IscR as the main regulator for iron- and tBOOH-dependent expression of the *isc-suf* operon. IscR (under iron repletion and iron depletion) and Irr (under iron depletion) also repressed P3 activity by direct binding. The presence of the upstream region, including P1, allowed higher transcription rates of *isc-suf* than P2 alone, and both antisense promoters stimulated P2 activity. The influence of Fur/Mur was observed for P3, but the effects were small and most likely indirect. Thus, multiple promoters and multiple regulators were involved in adjusting *isc-suf* expression to environmental conditions, and by regulating *iscR* expression indirectly, other genes of the iron metabolism that were controlled by IscR were also affected.

**Supplementary Materials:** The following are available online at http://www.mdpi.com/2076-2607/7/12/671/s1.

**Author Contributions:** X.N. contributed to acquisition, analysis, interpretation of the data, preparation of figures, and writing of the part of the manuscript. B.R. contributed to the design of the study. G.K. managed the project, contributed to the conception and design of the study, to the interpretation of the data, and writing of the manuscript.

**Funding:** The work was supported by Deutsche Forschungsgemeinschaft [grant Kl563/34-1], and Xin Nie was supported by a fellowship of the Chinese Scholarship Council (CSC).

**Acknowledgments:** We thank Katrin Werler (former employee of the Institute of Microbiology and Molecular Biology, University of Giessen) for constructing some of the reporter fusions and the RirA single mutants; Kerstin Haberzettl (Institute of Microbiology and Molecular Biology, University of Giessen) for excellent technical assistance; and Jochen Blom (Institute of Bioinformatics and Systems Biology, University of Giessen) for his kind help with the synteny analysis.

**Conflicts of Interest:** The authors declare no conflict of interest.
