The Role of LRP and H-NS in Transcription Regulation: Involvement of Synergism, Allostery and Macromolecular Crowding

J Mol Biol. 2007 Feb 23;366(3):900-15. doi: 10.1016/j.jmb.2006.11.067. Epub 2006 Dec 2.

Abstract

LRP has recently been shown to interact with the regulatory regions of bacterial ribosomal RNA promoters. Here we study details of the LRP-rDNA interaction by gel retardation and high-resolution footprinting techniques. We show that a second regulator for rRNA transcription, H-NS, facilitates the formation of a higher-order LRP-nucleoprotein complex, probably acting transiently as a DNA chaperone. The macromolecular crowding substance ectoine stabilizes the formation of this dynamic complex, while the amino acid leucine, as a metabolic effector, has the opposite effect. DNase I and hydroxyl radical footprint experiments with LRP-DNA complexes reveal a periodic change of the target DNA structure, which implies extensive DNA wrapping reaching into the promoter core region. We show furthermore that LRP binding is able to constrain supercoils, providing a link between DNA topology and regulation. The results support the conclusion that the bacterial DNA-binding protein LRP, assisted by H-NS, forms a repressive nucleoprotein structure involved in regulation of rRNA transcription. The formation of this regulatory structure appears to be directly affected by environmental changes.

MeSH terms

  • Allosteric Regulation / drug effects
  • Amino Acids, Diamino / pharmacology
  • Bacterial Proteins / metabolism*
  • DNA Footprinting
  • DNA Topoisomerases, Type I / metabolism
  • DNA, Bacterial / chemistry
  • DNA, Bacterial / metabolism
  • DNA, Ribosomal / metabolism
  • DNA-Binding Proteins / metabolism*
  • Deoxyribonuclease I / metabolism
  • Escherichia coli Proteins / metabolism
  • Factor For Inversion Stimulation Protein
  • Gene Expression Regulation, Bacterial* / drug effects
  • Hydroxyl Radical
  • Leucine-Responsive Regulatory Protein / metabolism*
  • Macromolecular Substances / metabolism*
  • Nucleic Acid Conformation / drug effects
  • Nucleoproteins / metabolism
  • Promoter Regions, Genetic / drug effects
  • Promoter Regions, Genetic / genetics
  • Protein Binding / drug effects
  • Transcription Factors / metabolism
  • Transcription, Genetic* / drug effects
  • rRNA Operon / drug effects
  • rRNA Operon / genetics

Substances

  • Amino Acids, Diamino
  • Bacterial Proteins
  • DNA, Bacterial
  • DNA, Ribosomal
  • DNA-Binding Proteins
  • Escherichia coli Proteins
  • Factor For Inversion Stimulation Protein
  • Fis protein, E coli
  • H-NS protein, bacteria
  • Macromolecular Substances
  • Nucleoproteins
  • Transcription Factors
  • Leucine-Responsive Regulatory Protein
  • Hydroxyl Radical
  • ectoine
  • Deoxyribonuclease I
  • DNA Topoisomerases, Type I