Abstract
Short-range DNA looping has been proposed to affect promoter activity in many bacterial species and operator configurations, but only few examples have been experimentally investigated in molecular detail. Here we present evidence for a metal-responsive DNA condensation mechanism controlled by the Helicobacter pylori ferric uptake regulator (Fur), an orthologue of the widespread Fur family of prokaryotic metal-dependent regulators. H. pylori Fur represses the transcription of the essential arsRS acid acclimation operon through iron-responsive oligomerization and DNA compaction, encasing the arsR transcriptional start site in a repressive macromolecular complex. A second metal-dependent regulator NikR functions as nickel-dependent anti-repressor at this promoter, antagonizing the binding of Fur to the operator elements responsible for the DNA condensation. The results allow unifying H. pylori metal ion homeostasis and acid acclimation in a mechanistically coherent model, and demonstrate, for the first time, the existence of a selective metal-responsive DNA compaction mechanism controlling bacterial transcriptional regulation.
Highlights
Short-range DNA looping has been proposed to affect promoter activity in many bacterial species and operator configurations, but only few examples have been experimentally investigated in molecular detail
To investigate how metal homeostasis regulation could transcriptionally feed into the ArsR regulon, we performed primer extension analyses of the native ParsR promoter using total RNA extracted from exponential phase H. pylori cultures and we compared the responses elicited by treatment with different metal ions (Fe2 þ and Ni2 þ ) or iron chelator (Dipy), under different genetic backgrounds (Fig. 1)
ParsR was constitutively de-repressed in a fur knockout strain, suggesting a prototypical holo-ferric uptake regulator (Fur) repression in which the iron ion acts as co-repressor (FeOFF)
Summary
Short-range DNA looping has been proposed to affect promoter activity in many bacterial species and operator configurations, but only few examples have been experimentally investigated in molecular detail. Fur proteins belong to an ubiquitously conserved superfamily of prokaryotic regulators involved in the homeostasis of different metal ions and oxidative stress responses[11] Because of their ability to oligomerize in a metal-dependent fashion[12,13] and to bind promoters at multiple sites[14,15,16], they represent ideal candidates for the investigation of metal-dependent DNAe. In addition, topological modifications of the DNA induced by Fur binding, such as bending and wrapping, have been reported in vitro using various footprinting and microscopy techniques, including atomic force microscopy (AFM)[17,18]. We use the thoroughly characterized H. pylori Fur regulator as a model to explore the metal-dependent short-scale DNA compaction mechanisms involved in transcriptional regulation To this aim, we investigate the Fur-regulated arsR promoter by a combination of DNase I footprinting, AFM and promoter functional analysis. Evidence is presented that this promoter architecture allows for iron- and
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