Potassium permanganate as a probe to map DNA-protein interactions in vivo.
Potassium permanganate (KMnO4) has widely been used in genomic footprinting assays to map unusual gene structures, including the melting DNA block in transcriptional elongation that results from promoter-proximal pausing of RNA polymerase (Pol) II complexes. Although it has been assumed that DNA-bound proteins do not protect underlying nucleic acids from KMnO4 modifications, we provide evidence herein that this chemical can readily be used to detect nuclear factor loading at a promoter when using optimized conditions. Moreover, by comparing parallel KMnO4 and dimethylsulfate (DMS) in vivo footprintings, we show that the utilization of KMnO4 in combination with another chemical probe maximizes the detection of factor occupancy at a DNA regulatory region, thus providing a better opportunity to define the actual profiles of DNA-protein contacts at given genomic sites in living cells.
Spicuglia
Salvatore
S
Centre d'Immunologie de Marseille-Luminy (CIML), INSERM-CNRS-Université de la Méditerranée Case 906, 13288 Marseille cedex 09, France. s.spicuglia@ncmls.kun.nl
Kumar
Sanjeev
S
Chasson
Lionel
L
Payet-Bornet
Dominique
D
Ferrier
Pierre
P
D000818:Animals; D001483:Base Sequence; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D018983:DNA Footprinting / Q000295:instrumentation*; D051379:Mice; D008969:Molecular Sequence Data; D011196:Potassium Permanganate / Q000737:chemistry* / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism*; D013463:Sulfuric Acid Esters / Q000378:metabolism; D013950:Thymus Gland / Q000378:metabolism
D011506:Proteins; D013463:Sulfuric Acid Esters; D011196:Potassium Permanganate; D004247:DNA; C007482:dimethyl sulfate