title: Bacterial promoter repression by DNA looping without protein-protein binding competition.
abstract: The Escherichia coli lactose operon provides a paradigm for understanding gene control by DNA looping where the lac repressor (LacI) protein competes with RNA polymerase for DNA binding. Not all promoter loops involve direct competition between repressor and RNA polymerase. This raises the possibility that positioning a promoter within a tightly constrained DNA loop is repressive per se, an idea that has previously only been considered in vitro. Here, we engineer living E. coli bacteria to measure repression due to promoter positioning within such a tightly constrained DNA loop in the absence of protein-protein binding competition. We show that promoters held within such DNA loops are repressed ∼100-fold, with up to an additional ∼10-fold repression (∼1000-fold total) dependent on topological positioning of the promoter on the inner or outer face of the DNA loop. Chromatin immunoprecipitation data suggest that repression involves inhibition of both RNA polymerase initiation and elongation. These in vivo results show that gene repression can result from tightly looping promoter DNA even in the absence of direct competition between repressor and RNA polymerase binding.
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, 200 First St. SW, Rochester, MN 55905, USA.
authors.forename: Nicole A
authors.initials: NA
authors.lastname: Becker
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, 200 First St. SW, Rochester, MN 55905, USA Luther College, Departments of Biology and Chemistry, Decorah, IA 52101, USA.
authors.forename: Alexander M
authors.initials: AM
authors.lastname: Greiner
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, 200 First St. SW, Rochester, MN 55905, USA.
authors.forename: Justin P
authors.initials: JP
authors.lastname: Peters
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, 200 First St. SW, Rochester, MN 55905, USA maher@mayo.edu.
authors.forename: L James
authors.initials: LJ
authors.lastname: Maher
mesh_terms: D001667:Binding, Competitive; D004269:DNA, Bacterial / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D020868:Gene Silencing; D017930:Genes, Reporter; D007763:Lac Operon; D056968:Lac Repressors / Q000235:genetics / Q000378:metabolism; D008957:Models, Genetic; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000235:genetics
chemical_list: D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D056968:Lac Repressors; C539064:LacI protein, E coli; D012321:DNA-Directed RNA Polymerases; D001616:beta-Galactosidase