HemN2 Regulates the Virulence of Pseudomonas donghuensis HYS through 7-Hydroxytropolone Synthesis and Oxidative Stress

Pseudomonas donghuensis HYS has lethal virulence towards Caenorhabditis elegans. Anaerobic coproporphyrinogen III oxidase (HemN) is involved in Pseudomonas heme synthesis. However, no research thus far has examined the contribution of HemN to the virulence of Pseudomonas. There are four hemN genes in P. donghuensis HYS, and we reported for the first time that the deletion of the hemN2 gene significantly reduced the virulence of P. donghuensis HYS towards C. elegans. HemN2 was negatively regulated by the Gac system and regulated bacterial virulence via 7-hydroxytropolone (7-HT) synthesis and redox levels. Our findings revealed the key role of HemN2 in bacterial virulence, which may help us to better understand the strong pathogenicity of the genus Pseudomonas. Compared to pathogens Pseudomonas aeruginosa and P. putida, P. donghuensis HYS has stronger virulence towards Caenorhabditis elegans. However, the underlying mechanisms haven't been fully understood. The heme synthesis system is essential for Pseudomonas virulence, and former studies of HemN have focused on the synthesis of heme, while the relationship between HemN and Pseudomonas virulence were barely pursued. In this study, we hypothesized that hemN2 deficiency affected 7-hydroxytropolone (7-HT) biosynthesis and redox levels, thereby reducing bacterial virulence. There are four hemN genes in P. donghuensis HYS, and we reported for the first time that deletion of hemN2 significantly reduced the virulence of HYS towards C. elegans, whereas the reduction in virulence by the other three genes was not significant. Interestingly, hemN2 deletion significantly reduced colonization of P. donghuensis HYS in the gut of C. elegans. Further studies showed that HemN2 was regulated by GacS and participated in the virulence of P. donghuensis HYS towards C. elegans by mediating the synthesis of the virulence factor 7-HT. In addition, HemN2 and GacS regulated the virulence of P. donghuensis HYS by affecting antioxidant capacity and nitrative stress. In short, the findings that HemN2 was regulated by the Gac system and that it was involved in bacterial virulence via regulating 7-HT synthesis and redox levels were reported for the first time. These insights may enlighten further understanding of HemN-based virulence in the genus Pseudomonas.