Estimation of DNA Loop Interactions Supports the Loop Domain Model of Insulator Action

Abstract

Long range regulation of transcription by regulatory proteins that bind to DNA elements involves DNA-looping interactions. The high specificity of these interactions is not well understood but may result from the “loop domain model”. According to this model, insulators form DNA loops and partition DNA into separate topological domains. Interactions between DNA elements within domains are enhanced while those between domains are attenuated. To quantify the specificity of DNA-loop interactions, DNA constructs were designed to form separate or overlapping loops mediated by lac and lambda repressors. DNA constructs with identical loop segments were used in both in vivo studies in E. coli and in vitro single molecule tethered particle motion (TPM) experiments. In the presence of lac and/or lambda repressors, the probabilities of loop formation ‘within’ or ‘between’ topological domains established by another loop were quantified. Fitting the probabilities to a statistical model produced an estimate of the assistance or interference that formation of one loop provided to a second loop forming within or across the boundaries of the first loop. Between domains loop interference was calculated to be 10 fold in vivo 2.5 fold in vitro. In contrast within domains, assistance was calculated to be 3 fold in vivo and 12.6 fold in vitro. Supercoiling within bacteria might be the reason for the difference between in vivo and in vitro estimates. Modelling these data shows that a combination of interference between domains and assistance within domains can generate highly specific long range interactions.