Protein-mediated loops in supercoiled DNA create large topological domains

Abstract

Supercoiling can alter the form and base pairing of the double helix and directly impact protein binding. More indirectly, changes in protein binding and the stress of supercoiling also influence the thermodynamic stability of regulatory, protein-mediated loops and shift the equilibria of fundamental DNA/chromatin transactions. For example, supercoiling affects the hierarchical organization and function of chromatin in topologically associating domains (TADs) in both eukaryotes and bacteria. On the other hand, a protein-mediated loop in DNA can constrain supercoiling within a plectonemic structure. To characterize the extent of constrained supercoiling, 400 bp, lac repressor-secured loops were formed in extensively over- or under-wound DNA under gentle tension in a magnetic tweezer. The protein-mediated loops constrained variable amounts of supercoiling that often exceeded the maximum writhe expected for a 400 bp plectoneme. Loops with such high levels of supercoiling appear to be entangled with flanking domains. Thus, loop-mediating proteins operating on supercoiled substrates can establish topological domains that may coordinate gene regulation and other DNA transactions across spans in the genome that are larger than the separation between the binding sites.

Nucleosome spacing periodically modulates nucleosome chain folding and DNA topology in circular nucleosome arrays

“…Recent chromosome conformation capture studies have revealed chromatin looping anchored by SMC proteins (condensin and cohesin) and self-association between similar epigenetically marked chromatin states as the two universal principles of structural and functional organization of eukaryotic genome (35). Remarkably, loops anchored by the SMC proteins could be strong enough to maintain a locally altered DNA supercoiling within a closed loop (36). Chromatin loops play an important functional role in mediating interactions between transcription-activating DNA elements such as enhancers and promoters (37,38).…”

Transfer-matrix calculations of the effects of tension and torque constraints on DNA–protein interactions

“…[4][5][6][7][8][9]), that generate stretching and twisting forces on the chromosomal DNA [10][11][12][13][14][15]. It is also known that chromosomes form extensive adhesion contacts with a number of nuclear membrane proteins, establishing force-transmitting links between the chro-plexes (such as eukaryotic/archaeal histones) [23,24,35]; 2) DNA-bending proteins, which sharply curve DNA at the protein binding site (like bacterial HU, IHF and Fis) [22, 25-28, 30, 32, 36]; 3) DNA-bridging proteins that cross-link DNA duplexes (for example, bacterial H-NS, human HMGA2, or any other protein that mediates DNA loops) [29,[37][38][39], and 4) DNA-stiffening proteins forming rigid nucleoprotein filaments along DNA (like archaeal TrmBL2 and Alba) [31,33,40]. Thus, the four major groups of DNA-architectural proteins form nucleoprotein complexes, which have very different 3D structures, leading to diverse responses of these proteins to force and torque constraints applied to DNA.…”

Requirements for DNA-Bridging Proteins to Act as Topological Barriers of the Bacterial Genome

“…Based on recent experimental results ( 12 , 13 , 14 , 15 , 16 ), one may wonder whether DNA loops are ipso facto independent topological domains or whether some additional, more subtle properties of DNA-bridging proteins are required to achieve this goal. We emphasize that the word “loop” is used here according to its biological meaning and not the geometrical one.…”

“…The most straightforward explanation for this result is that two bridges in tandem are required to sustain the torque arising from the difference in torsional stress in the two DNA loops. Yet, single molecule experiments have demonstrated that a single LacI bridge can sustain the torque associated with physiological values of supercoiling, that is σ ≈ −0.06 ( 16 ). In this context, we surmise that the slow relaxation observed in ( 15 ) for single DNA bridges may correspond to the slow intrication obtained with our second model for DNA-bridging proteins, whereas the more stable topological separation observed in ( 15 ) for bridges in tandem corresponds to true barriers obtained with the third model for DNA-bridging proteins.…”

“…Although this question has now been addressed for nearly two decades ( 7 , 8 , 9 ), the mechanisms underlying the formation of topological barriers remain mostly elusive. The most plausible models for the formation of topological barriers are 1) actively transcribing RNA polymerases (RNAPs), which generate both positive and negative supercoils ( 10 ) and may consequently block the displacement and dissipation of plectonemes ( 11 , 12 ); and 2) the formation of DNA loops by certain nucleoid proteins that may serve as topological barriers ( 13 , 14 , 15 , 16 ). In this work, we focus on this latter mechanism and establish under which conditions DNA-bridging proteins may serve as topological barriers.…”