Protein/DNA interactions in complex DNA topologies

Authors: Agnes Noy Thana Sutthibutpong Sarah A Harris

Publish Date: 2016/08/08

Volume: 8, Issue: 3, Pages: 233-243

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Abstract

DNA supercoiling results in compacted DNA structures that can bring distal sites into close proximity It also changes the local structure of the DNA which can in turn influence the way it is recognised by drugs other nucleic acids and proteins Here we discuss how DNA supercoiling and the formation of complex DNA topologies can affect the thermodynamics of DNA recognition We then speculate on the implications for transcriptional control and the threedimensional organisation of the genetic material using examples from our own simulations and from the literature We introduce and discuss the concept of coupling between the multiple lengthscales associated with hierarchical nuclear structural organisation through DNA supercoiling and topologyWe propose that supercoiling provides a mechanism to amplify information and to communicate activity in the genome across multiple levels of nuclear organisation In support of this view we combine the physical insight provided by computer simulations that show in atomic detail how DNA responds to topological stress with experimental evidence for the importance of supercoiling Supercoiling effects DNA interactions with other molecules across a spectrum of lengthscales from the size of the counterion species in the environment to up to nuclear domains formed by clusters of DNAbinding proteins We conclude that an understanding of the physical interactions that couple chromosome organisation to gene regulation will require a multiscale approach starting from sequencedependent DNA mechanics through the organisation of DNA domains by architectural DNAbinding proteins up to the global organisation of the whole chromosomeIn this review we first describe the origins of supercoiling in DNA and then the multiple levels of hierarchical DNA structural organisation from a DNA mechanics point of view The effect of the counterion environment and DNAbinding proteins on supercoiling locally and globally are then discussed in the context of each level of nuclear organisation The background concepts in DNA supercoiling and topology such as definitions of twist and writhe and superhelical density can be found in Bates and Maxwell Bates and Maxwell 2005 and a detailed description of the evidence for the importance of DNA supercoiling in mediating protein–DNA interactions can be found in a previous review Fogg et al 2012DNA supercoiling is a cellular strategy for packing the genetic material efficiently into a small nuclear space but it is also implicated in genetic control see for examples Fogg et al 2012 Gilbert and Allan 2014 Koster et al 2010 Lavelle 2014 The physical mechanisms that couple supercoiling to levels of gene transcription are less well understood than the biochemical signalling pathways mediated by DNAbinding proteins such as transcription factors activators and repressors because of the experimental difficulties associated with measuring supercoiling in active DNA The origins of DNA supercoiling can be considered to consist of static supercoils which change slowly and are externally regulated by the cell and dynamic supercoils which are introduced transiently by DNA processing machines such as RNA polymeraseIn prokaryotes the genome is maintained in a negatively supercoiled state by DNA gyrase which uses chemical energy to maintain an average torsional stress of around σ = −006 within the DNA Collin et al 2011 In eukaryotes the lefthanded wrapping of the DNA in the nucleosome constrains the DNA to be negatively writhed this writhe is converted into supercoiling if the histone complex is displaced Teves and Henikoff 2014 Although the higher order organisation of chromatin remains poorly understood levels of superhelical density in the range σ = −009 and −006 range have been suggested for chromatin fibres depending upon the precise organisation of the nucleosome units Norouzi and Zhurkin 2015 Magnetic tweezer experiments that subjected nucleosome arrays to torque have demonstrated that chromatin can absorb a large amount of supercoiling without undergoing a substantial change in length This ability of chromatin to act as a “topological buffer” that shields regions of the genome from changes in supercoiling could be explained by a model in which chromatin adopts multiple conformational states Bancaud et al 2006Dynamic supercoiling is introduced by transcription as separation of the doublehelical strands creates positive and negative supercoiling ahead and behind the polymerase complex respectively Liu and Wang 1987 If the ends of the DNA are restrained then this dynamic supercoiling will be stored within this section of the genome Transcription is stalled whenever too much positive supercoiling builds up ahead of the RNA polymerase and its associated machinery while a gene is being read Chong et al 2014 While the presence of topoisomerases which relax supercoiled DNA implies that the cell has mechanisms in place to dissipate dynamic supercoiling Baranello et al 2016 experiments that measured the levels of negative supercoiling in the DNA using intercalating agents detected superhelical stresses from transcription equivalent to σ = −007 over length scales of between 1 and 15 kb from the polymerase enzyme Kouzine et al 2013 as well as a large negative supercoiling gradient between the replication origin and the terminus during stationary growth phase in Escherichia coli Lal et al 2016There is growing evidence that dynamic supercoiling confers regulatory information over long distances through the genome In general genes that are AT rich tend to be downregulated by increased negative supercoiling whereas GCrich genes have a propensity to be upregulated for more details see the review by Fogg et al 2012 Negative supercoiling destabilises the doublehelical structure of the DNA which facilitates melting and therefore affects the delicate balance between DNA opening and reannealing required for successful transcription This provides a mechanism to couple together the transcriptional activity of successive genes without the requirement for DNAbinding proteins Transmission of information through the DNA itself provides a particularly efficient mechanism for cooperative gene expression as it does not rely either on protein production or the location of a specific binding site by protein diffusion A striking demonstration of the importance of coupled gene expression comes from an analysis of the E coli genome leading to the suggestion that genes are positioned and orientated to ensure that the influence of supercoiling from transcription of neighbouring genes is maintained through evolution Sobetzko 2016 Studies in eukaryotes have additionally suggested that the generation of short divergent RNA transcripts could have a regulatory function by underwinding the promoters and facilitating transcription Naughton et al 2013aThe amount of superhelical stress that builds up in the DNA from transcription the lengthscale associated with this mechanical perturbation and its timescale depends upon a complex interplay between a number of physical effects the efficiency of supercoil removal by topoisomerases the mechanical response of the DNA itself and the presence of DNAbinding proteins which may act as mechanical clamps imposing a given global topology Static and dynamic supercoiling can also be coupled For example negative supercoiling can be generated by unwinding DNA from histones which can then facilitate promotor melting and passage of RNA polymerase along the DNA Kouzine et al 2014 The connection between DNA–protein interactions and static and dynamic supercoiling introduces a coupling between the multiple levels of structural organisation within the genome which is discussed in the following section

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