Molecular mechanisms of regulation by Ss-LrpB, a transcriptional regulator from the archaeon Sulfolobus solfataricus

Eveline Peeters, Nuno Peixeiro, Nadal Marc, Patrick Forterre, Guennadi Sezonov, David Prangishvili, Daniel Charlier

Research output: Chapter in Book/Report/Conference proceedingMeeting abstract (Book)

Abstract

Transcription in Archaea resembles closely the eukaryotic process, with a basal transcription apparatus minimally composed of TATA-binding protein (TBP), transcription factor B (TFB) and a RNA polymerase (RNAP) resembling the eukaryotic RNA polymerase II. In contrast, most archaeal transcription regulators are predicted to resemble their bacterial counterparts. Only a limited number of archaeal transcription regulators have been studied, most of which belong to the bacterial/archaeal Lrp (Leucine-responsive Regulatory Protein) family of regulators. However, regulatory mechanisms and effects of archaeal Lrp-like regulators are very poorly characterized. Here, we study the function of the Lrp-like regulator Ss-LrpB of Sulfolobus solfataricus, a hyperthermoacidophilic archaeon. Previously, we have identified four target promoters of the regulator, namely the promoters of Ss-lrpB itself (autoregulation), and of the adjacently located pyruvate ferredoxin oxidoreductase (porDAB) operon and two permease genes. In vitro binding to all promoter regions has been extensively studied and gene expression of the three non-autogenously regulated target genes is lower in an Ss-lrpB gene disruption mutant strain as compared to the isogenic WT strain, indicating that Ss-LrpB may activate transcription.
Regulatory effects exerted by Ss-LrpB were further analyzed by using a Sulfolobus in vitro transcription system. Transcription at all four target promoters was clearly stimulated upon addition of recombinant Ss-LrpB at low concentrations. However, major qualitative and quantitative differences were observed for the various target promoters in function of the Ss-LrpB concentration. The stimulatory effect was much stronger for porDAB than for the two transporter genes, and transcription increased constantly with increasing Ss-LrpB concentration. In contrast, the stimulatory effect on initiation at the permease promoters and the own promoter observed at low Ss-LrpB concentrations was reversed into a specific repression at higher Ss-LrpB concentrations. These results clearly demonstrate that Ss-LrpB is a dual regulator, able to 'switch' its function between an activator and a repressor in a concentration-dependent manner. Crucial elements determining the regulatory outcome are the binding site organization, the position of the regulatory protein relative to the promoter, cooperativity in the binding and Ss-LrpB induced DNA wrapping, as observed with the own control region. DNA wrapping was shown by a topological assay to result in a positive supercoil and is assumed to be a major determinant of repression.
Original languageEnglish
Title of host publicationMeeting Belgian Society of Microbiology
Publication statusPublished - Dec 2009

Publication series

NameMeeting Belgian Society of Microbiology

Keywords

  • archaea
  • protein-DNA interactions
  • sulfolobus
  • leucine-responsive Regulatory Protein
  • transcription regulation

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