Quantitation of interactions between two DNA loops demonstrates loop domain insulation in E. coli cells

• Published in: Proceedings of the National Academy of Sciences - PNAS 2014-10, Vol.111 (42), p.E4449-E4457
• Main Authors: Priest, David G, Kumar, Sandip, Yan, Yan, Dunlap, David D, Dodd, Ian B, Shearwin, Keith E
• Format: Article
• Language: English
• Subjects: Binding Sites; Biological Sciences; Cells; Deoxyribonucleic acid; DNA; DNA, Bacterial - chemistry; DNA, Bacterial - genetics; DNA, Superhelical - chemistry; E coli; Escherichia coli; Escherichia coli - genetics; Eukaryotes; Gene Expression Regulation, Bacterial; Genes; Genes, Reporter; Lac Operon; Lac Repressors; Models, Statistical; Monte Carlo Method; Nucleic Acid Conformation; Operator Regions, Genetic; PNAS Plus; Promoter Regions, Genetic; Repressor Proteins - chemistry; Stress, Mechanical
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1410764111

Significance Genes are frequently regulated by interactions between proteins that bind to the DNA near the gene and proteins that bind to DNA sites located far away, with the intervening DNA looped out. In eukaryotic genomes, genes and their distant sites are intermingled in complex ways and it is not understood how the correct connections are formed. Using two pairs of DNA-looping sites in bacterial cells, we tested the idea that one DNA loop can either assist or interfere with the formation of another DNA loop. By measuring the strength of these interactions between loops, we showed that this mechanism is capable of directing a distant site to the correct gene and preventing it contacting the wrong gene. Eukaryotic gene regulation involves complex patterns of long-range DNA-looping interactions between enhancers and promoters, but how these specific interactions are achieved is poorly understood. Models that posit other DNA loops—that aid or inhibit enhancer–promoter contact—are difficult to test or quantitate rigorously in eukaryotic cells. Here, we use the well-characterized DNA-looping proteins Lac repressor and phage λ CI to measure interactions between pairs of long DNA loops in E. coli cells in the three possible topological arrangements. We find that side-by-side loops do not affect each other. Nested loops assist each other’s formation consistent with their distance-shortening effect. In contrast, alternating loops, where one looping element is placed within the other DNA loop, inhibit each other’s formation, thus providing clear support for the loop domain model for insulation. Modeling shows that combining loop assistance and loop interference can provide strong specificity in long-range interactions.