Effects of nucleoid proteins on DNA repression loop formation in Escherichia coli.
The intrinsic stiffness of DNA limits its ability to be bent and twisted over short lengths, but such deformations are required for gene regulation. One classic paradigm is DNA looping in the regulation of the Escherichia coli lac operon. Lac repressor protein binds simultaneously to two operator sequences flanking the lac promoter. Analysis of the length dependence of looping-dependent repression of the lac operon provides insight into DNA deformation energetics within cells. The apparent flexibility of DNA is greater in vivo than in vitro, possibly because of host proteins that bind DNA and induce sites of flexure. Here we test DNA looping in bacterial strains lacking the nucleoid proteins HU, IHF or H-NS. We confirm that deletion of HU inhibits looping and that quantitative modeling suggests residual looping in the induced operon. Deletion of IHF has little effect. Remarkably, DNA looping is strongly enhanced in the absence of H-NS, and an explanatory model is proposed. Chloroquine titration, psoralen crosslinking and supercoiling-sensitive reporter assays show that the effects of nucleoid proteins on looping are not correlated with their effects on either total or unrestrained supercoiling. These results suggest that host nucleoid proteins can directly facilitate or inhibit DNA looping in bacteria.
Becker
Nicole A
NA
Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, 200 First St SW, Rochester, MN 55905, USA.
Kahn
Jason D
JD
Maher
L James
LJ
D001426:Bacterial Proteins / Q000235:genetics / Q000502:physiology; D004269:DNA, Bacterial / Q000737:chemistry*; D004278:DNA, Superhelical / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000502:physiology*; D017353:Gene Deletion; D034982:Integration Host Factors / Q000235:genetics / Q000502:physiology; D007763:Lac Operon; D008957:Models, Genetic; D009690:Nucleic Acid Conformation
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C031908:H-NS protein, bacteria; D034982:Integration Host Factors; C116822:hns protein, E coli; C440502:integration host factor, E coli