The role of LRP and H-NS in transcription regulation: involvement of synergism, allostery and macromolecular crowding.
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.
Pul
Umit
U
Institut für Physikalische Biologie, Heinrich-Heine-Universität Düsseldorf, Universitätsstr 1, D-40225 Düsseldorf, Germany.
Wurm
Reinhild
R
Wagner
Rolf
R
D000494:Allosteric Regulation / Q000187:drug effects; D000599:Amino Acids, Diamino / Q000494:pharmacology; D001426:Bacterial Proteins / Q000378:metabolism*; D018983:DNA Footprinting; D004264:DNA Topoisomerases, Type I / Q000378:metabolism; D004269:DNA, Bacterial / Q000737:chemistry / Q000378:metabolism; D004275:DNA, Ribosomal / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism; D035021:Factor For Inversion Stimulation Protein; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D017665:Hydroxyl Radical; D051756:Leucine-Responsive Regulatory Protein / Q000378:metabolism*; D046911:Macromolecular Substances / Q000378:metabolism*; D009690:Nucleic Acid Conformation / Q000187:drug effects; D009698:Nucleoproteins / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D012407:rRNA Operon / Q000187:drug effects / Q000235:genetics
D000599:Amino Acids, Diamino; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D035021:Factor For Inversion Stimulation Protein; C501034:Fis protein, E coli; C031908:H-NS protein, bacteria; D046911:Macromolecular Substances; D009698:Nucleoproteins; D014157:Transcription Factors; D051756:Leucine-Responsive Regulatory Protein; D017665:Hydroxyl Radical; C045628:ectoine; D003850:Deoxyribonuclease I; D004264:DNA Topoisomerases, Type I