Independent and cooperative regulation of staphylopine biosynthesis and trafficking by Fur and Zur.

Staphylococcus aureus expresses the Cnt system implicated in the active transport of trace metals by synthesizing (CntKLM) and exporting (CntE) staphylopine, a metallophore chelating metals and then taken up by an ABC-transporter (CntABCDF). This machinery is encoded in the cntKLMABCDFE operon, preceded by a non-coding region (PcntK) and containing an internal promoter region (PcntA). PcntK comprises a Fur box followed by a Zur box, a sRNA transcription start and a repeated region, while PcntA comprises a Fur box that overlaps a Zur box. We found that PcntK promoter activity is attenuated by the repeated sequence and strictly controlled by Fur or Zur binding to its respective target sequences. Interestingly, we discovered a cooperative regulation of the PcntA activity by both Fur and Zur binding to the Fur/Zur box, by identifying a tripartite complex with DNA. Repression of PcntA is less sensitive to metal concentration and therefore loosely repressed as compared to PcntK activity. Furthermore, the Cnt system is essential for the optimal import of zinc, thereby linking regulation and function of Cnt. Overall, our results highlight the need for fine and differential tuning of staphylopine biosynthesis and trafficking in order to efficiently respond to metal starvation and optimize metal recovery.