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Transcript analysis of nrrF, a Fur repressed sRNA of Neisseria gonorrhoeae.

ABSTRACT: Like most microorganisms, Neisseria gonorrhoeae alters gene expression in response to iron availability. The ferric uptake regulator Fur has been shown to be involved in controlling this response, but the extent of this involvement remains unknown. It is known that in addition to working directly to repress gene expression, Fur may also work indirectly by controlling additional regulatory elements. Using in silico analysis, we identified a putative small RNA (sRNA) homolog of the meningococcal nrrF locus, and demonstrate that this sRNA is iron-repressible, suggesting that this is the gonococcal analog of the rhyB locus in Escherichia coli. Quantitative real-time RT-PCR analysis indicates that this transcript may also be temporally regulated. Transcript analysis identified the 5' start of the transcript, using a single reaction, fluorescent-based, primer extension assay. This protocol allows for the rapid identification of transcriptional start sites of RNA transcripts, and could be used for high-throughput transcript mapping.

SUBMITTER: Ducey TF

PROVIDER: S-EPMC4890603 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature

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Transcript analysis of nrrF, a Fur repressed sRNA of Neisseria gonorrhoeae.

Ducey Thomas F TF Jackson Lydgia L Orvis Joshua J Dyer David W DW

Microbial pathogenesis 20090106 3

Like most microorganisms, Neisseria gonorrhoeae alters gene expression in response to iron availability. The ferric uptake regulator Fur has been shown to be involved in controlling this response, but the extent of this involvement remains unknown. It is known that in addition to working directly to repress gene expression, Fur may also work indirectly by controlling additional regulatory elements. Using in silico analysis, we identified a putative small RNA (sRNA) homolog of the meningococcal n ...[more]

PMID: 19162160

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Project description: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 wild-type 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 ?(70) 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.Gene regulation in bacteria in response to environmental stimuli, including iron, is of paramount importance to both bacterial replication and, in the case of pathogenic bacteria, successful infection. Bacterial DNA binding proteins are a common mechanism utilized by pathogens to control gene expression under various environmental conditions. Here, we show that the DNA binding protein Fur, expressed by the human pathogen Neisseria gonorrhoeae, controls the expression of a large repertoire of genes and extends this regulon by controlling expression of additional DNA binding proteins. One of these proteins, an ArsR-like regulator, was required for N. gonorrhoeae survival within host cells. These results show that the Fur regulon extends to additional regulatory proteins, which together contribute to gonococcal mechanisms of pathogenesis.

Dataset Information

Transcript analysis of nrrF, a Fur repressed sRNA of Neisseria gonorrhoeae.

Publications

Transcript analysis of nrrF, a Fur repressed sRNA of Neisseria gonorrhoeae.

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