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Transcriptional and mutational analyses of the rpoN operon in Caulobacter crescentus.

ABSTRACT: The alternative sigma factor sigma54 is required for the biogenesis of both the flagellum and the stalk in Caulobacter crescentus. The DNA sequence downstream of the sigma54 gene (rpoN) has been determined, revealing three open reading frames (ORFs) encoding peptides of 203, 208, and 159 amino acids. ORF208 and ORF159 are homologous to ORFs found downstream of rpoN in other microorganisms. The organization of this region in C. crescentus is similar to that in other bacteria, with the exception of an additional ORF, ORF203, immediately downstream from rpoN. There is a single temporally regulated promoter that drives the expression of both rpoN and ORF203. Promoter probe analysis indicates the presence of another promoter downstream from ORF203 which exhibits a temporal control that is different from that of the rpoN promoter. Mutational analysis was used to address the function of the proteins encoded by these three downstream ORFs. The mutations have no effect on the transcription of previously known sigma54-dependent flagellar promoters except for a slight effect of an ORF159 mutation on transcription of fljK.

SUBMITTER: Janakiraman RS

PROVIDER: S-EPMC179373 | biostudies-other | 1997 Aug

REPOSITORIES: biostudies-other

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Transcriptional and mutational analyses of the rpoN operon in Caulobacter crescentus.

Janakiraman R S RS Brun Y V YV

Journal of bacteriology 19970801 16

The alternative sigma factor sigma54 is required for the biogenesis of both the flagellum and the stalk in Caulobacter crescentus. The DNA sequence downstream of the sigma54 gene (rpoN) has been determined, revealing three open reading frames (ORFs) encoding peptides of 203, 208, and 159 amino acids. ORF208 and ORF159 are homologous to ORFs found downstream of rpoN in other microorganisms. The organization of this region in C. crescentus is similar to that in other bacteria, with the exception o ...[more]

PMID: 9260957

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Project description:BackgroundIn the alpha subclass of proteobacteria iron homeostasis is controlled by diverse iron responsive regulators. Caulobacter crescentus, an important freshwater α-proteobacterium, uses the ferric uptake repressor (Fur) for such purpose. However, the impact of the iron availability on the C. crescentus transcriptome and an overall perspective of the regulatory networks involved remain unknown.ResultsIn this work we report the identification of iron-responsive and Fur-regulated genes in C. crescentus using microarray-based global transcriptional analyses. We identified 42 genes that were strongly upregulated both by mutation of fur and by iron limitation condition. Among them, there are genes involved in iron uptake (four TonB-dependent receptor gene clusters, and feoAB), riboflavin biosynthesis and genes encoding hypothetical proteins. Most of these genes are associated with predicted Fur binding sites, implicating them as direct targets of Fur-mediated repression. These data were validated by β-galactosidase and EMSA assays for two operons encoding putative transporters. The role of Fur as a positive regulator is also evident, given that 27 genes were downregulated both by mutation of fur and under low-iron condition. As expected, this group includes many genes involved in energy metabolism, mostly iron-using enzymes. Surprisingly, included in this group are also TonB-dependent receptors genes and the genes fixK, fixT and ftrB encoding an oxygen signaling network required for growth during hypoxia. Bioinformatics analyses suggest that positive regulation by Fur is mainly indirect. In addition to the Fur modulon, iron limitation altered expression of 113 more genes, including induction of genes involved in Fe-S cluster assembly, oxidative stress and heat shock response, as well as repression of genes implicated in amino acid metabolism, chemotaxis and motility.ConclusionsUsing a global transcriptional approach, we determined the C. crescentus iron stimulon. Many but not all of iron responsive genes were directly or indirectly controlled by Fur. The iron limitation stimulon overlaps with other regulatory systems, such as the RpoH and FixK regulons. Altogether, our results showed that adaptation of C. crescentus to iron limitation not only involves increasing the transcription of iron-acquisition systems and decreasing the production of iron-using proteins, but also includes novel genes and regulatory mechanisms.

Dataset Information

Transcriptional and mutational analyses of the rpoN operon in Caulobacter crescentus.

Publications

Transcriptional and mutational analyses of the rpoN operon in Caulobacter crescentus.

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