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Iron is both essential for bacterial growth and toxic at higher concentrations; thus, iron homeostasis is tightly regulated. In Neisseria meningitidis the majority of iron-responsive gene regulation is mediated by the ferric uptake regulator protein (Fur), a protein classically defined as a transcriptional repressor. Recently, however, microarray studies have identified a number of genes in N. meningitidis that are iron and Fur activated, demonstrating a new role for Fur as a transcriptional activator. Since Fur has been shown to indirectly activate gene transcription through the repression of small regulatory RNA molecules in other organisms, we hypothesized that a similar mechanism could account for Fur-dependent, iron-activated gene transcription in N. meningitidis. In this study, we used a bioinformatics approach to screen for the presence of Fur-regulated small RNA molecules in N. meningitidis MC58. This screen identified one small RNA, herein named NrrF (for neisserial regulatory RNA responsive to iron [Fe]), which was demonstrated to be both iron responsive and Fur regulated and which has a well-conserved orthologue in N. gonorrhoeae. In addition, this screen identified a number of other likely, novel small RNA transcripts. Lastly, we utilized a new bioinformatics approach to predict regulatory targets of the NrrF small RNA. This analysis led to the identification of the sdhA and sdhC genes, which were subsequently demonstrated to be under NrrF regulation in an nrrF mutant. This study is the first report of small RNAs in N. meningitidis and the first to use a bioinformatics approach to identify, a priori, regulatory targets of a small RNA.

Our whole-genome microarray studies of Neisseria meningitidis MC58 previously identified a set of 153 genes whose transcription was activated during growth in iron. In this study, Fur-mediated regulation of the iron-activated nspA gene was confirmed, whereas iron-activated regulation of the secY gene was demonstrated to be Fur independent. Analysis of the Fur binding sequences in the nspA gene and an additional iron-activated and Fur-regulated gene identified a hexameric (G/T)ATAAT unit in the operator regions of these genes similar to that observed in Fur- and iron-repressed genes. These studies indicate that the expression of the iron-activated nspA and secY genes in N. meningitidis occur by Fur-dependent and -independent mechanisms, respectively. [ABSTRACT FROM AUTHOR]