A complex mechanism involving LysR and TetR/ AcrR that regulates iron scavenger biosynthesis in Pseudomonas donghuensis HYS

Abstract

7-Hydroxytropolone (7-HT) is a symmetrical seven-membered heteroatomic ring with a carboxyl group and two hydroxyl groups and was recently reported to be an iron scavenger of Pseudomonas donghuensis HYS. Cluster 1 includes 12 genes related to the synthesis of 7-HT; among these genes, those for two regulators, Orf1 and Orf12, were predicted to regulate 7-HT biosynthesis and to be LysRtype transcriptional regulators (LTTRs) and TetR/AcrR family transcriptional regulators, respectively. Data from real-time quantitative PCR and β-galactosidase and classical siderophore assays indicated that the transcription levels of orf1 and orf12, as well as those of crucial genes orf6 to orf9, were repressed under high-iron conditions. The deletion of orf1 and orf12 led to an absence of 7-HT and a decrease in orf6-orf9 expression. Orf1 and Orf12 were essential for the production of 7-HT through orf6-orf9. These two regulators are regulated by the Gac/Rsm system; Orf1 facilitates the expression of Orf12, and Orf12 concomitantly stimulates the expression of orf6-orf9 to synthesize 7-HT. The overexpression of Orf12 decreased 7-HT yields, possibly through decreased orf6-orf9 expression. This work thus outlines a complex mechanism regulating the biosynthesis of the iron scavenger 7-HT in P. donghuensis HYS. The synergy between Orf1 and Orf12 ensures that 7-HT acts as an iron chelator despite being toxic to bacteria and provides new ideas for the novel regulation of dual-functional secondary metabolism and research on 7-HT and its derivates in other bacteria.