Characterization of the Neisseria gonorrhoeae Iron and Fur regulatory network

Abstract

The Neisseria gonorrhoeae ferric uptake regulator (Fur) protein controls expression of iron homeostasis genes in response to intracellular iron levels. In this study, using transcriptome sequencing (RNA-seq) analysis of an N. gonorrhoeae fur strain, we defined the gonococcal Fur and iron regulons and characterized Fur-controlled expression of an ArsR-like DNA binding protein. We observed that 158 genes (8% of the genome) showed differential expression in response to iron in an N. gonorrhoeae wildtype or fur strain, while 54 genes exhibited differential expression in response to Fur. The Fur regulon was extended to additional regulators, including NrrF and 13 other small RNAs (sRNAs), and two transcriptional factors. One transcriptional factor, coding for an ArsR-like regulator (ArsR), exhibited increased expression under iron-replete conditions in the wild-type strain but showed decreased expression across iron conditions in the fur strain, an effect that was reversed in a fur-complemented strain. Fur was shown to bind to the promoter region of the arsR gene downstream of a predicted σ70 promoter region. Electrophoretic mobility shift assay (EMSA) analysis confirmed binding of the ArsR protein to the norB promoter region, and sequence analysis identified two additional putative targets, NGO1411 and NGO1646. A gonococcal arsR strain demonstrated decreased survival in human endocervical epithelial cells compared to that of the wild-type and arsR-complemented strains, suggesting that the ArsR regulon includes genes required for survival in host cells. Collectively, these results demonstrate that the N. gonorrhoeae Fur functions as a global regulatory protein to repress or activate expression of a large repertoire of genes, including additional transcriptional regulatory proteins.