Torque modulates nucleosome stability and facilitates H2A/H2B dimer loss.

The nucleosome, the fundamental packing unit of chromatin, has a distinct chirality: 147 bp of DNA are wrapped around the core histones in a left-handed, negative superhelix. It has been suggested that this chirality has functional significance, particularly in the context of the cellular processes that generate DNA supercoiling, such as transcription and replication. However, the impact of torsion on nucleosome structure and stability is largely unknown. Here we perform a detailed investigation of single nucleosome behaviour on the high-affinity 601-positioning sequence under tension and torque using the angular optical trapping technique. We find that torque has only a moderate effect on nucleosome unwrapping. In contrast, we observe a dramatic loss of H2A/H2B dimers on nucleosome disruption under positive torque, whereas (H3/H4)2 tetramers are efficiently retained irrespective of torsion. These data indicate that torque could regulate histone exchange during transcription and replication.

Torque measurement at the single-molecule level.

Methods for exerting and measuring forces on single molecules have revolutionized the study of the physics of biology. However, it is often the case that biological processes involve rotation or torque generation, and these parameters have been more difficult to access experimentally. Recent advances in the single-molecule field have led to the development of techniques that add the capability of torque measurement. By combining force, displacement, torque, and rotational data, a more comprehensive description of the mechanics of a biomolecule can be achieved. In this review, we highlight a number of biological processes for which torque plays a key mechanical role. We describe the various techniques that have been developed to directly probe the torque experienced by a single molecule, and detail a variety of measurements made to date using these new technologies. We conclude by discussing a number of open questions and propose systems of study that would be well suited for analysis with torsional measurement techniques.

Recent advances in single molecule studies of nucleosomes.

As the fundamental packing units of DNA in eukaryotes, nucleosomes play a central role in governing DNA accessibility in a variety of cellular processes. Our understanding of the mechanisms underlying this complex regulation has been aided by unique structural and dynamic perspectives offered by single molecule techniques. Recent years have witnessed remarkable advances achieved using these techniques, including the generation of a detailed histone-DNA energy landscape, elucidation of nucleosome disassembly processes, and real-time monitoring of molecular motors interacting with nucleosomes. These and other highlights of single molecule nucleosome studies will be discussed in this review.

Underwound DNA under tension: structure, elasticity, and sequence-dependent behaviors.

DNA melting under torsion plays an important role in a wide variety of cellular processes. In the present Letter, we have investigated DNA melting at the single-molecule level using an angular optical trap. By directly measuring force, extension, torque, and angle of DNA, we determined the structural and elastic parameters of torsionally melted DNA. Our data reveal that under moderate forces, the melted DNA assumes a left-handed structure as opposed to an open bubble conformation and is highly torsionally compliant. We have also discovered that at low forces melted DNA properties are highly dependent on DNA sequence. These results provide a more comprehensive picture of the global DNA force-torque phase diagram.

Discontinuities at the DNA supercoiling transition.

While slowly turning the ends of a single molecule of DNA at constant applied force, a discontinuity was recently observed at the supercoiling transition when a small plectoneme is suddenly formed. This can be understood as an abrupt transition into a state in which stretched and plectonemic DNA coexist. We argue that there should be discontinuities in both the extension and the torque at the transition and provide experimental evidence for both. To predict the sizes of these discontinuities and how they change with the overall length of DNA, we organize a phenomenological theory for the coexisting plectonemic state in terms of four parameters. We also test supercoiling theories, including our own elastic rod simulation, finding discrepancies with experiment that can be understood in terms of the four coexisting state parameters.

Twist-stretch coupling and phase transition during DNA supercoiling.

As a single DNA molecule is positively supercoiled under constant tension, its extension initially increases due to a negative twist-stretch coupling. The subsequent attainment of an extension maximum has previously been assumed to be indicative of the onset of a phase transition from B- to scP-DNA. Here we show that an extension maximum in fact does not coincide with the onset of a phase transition. This transition is evidenced by a direct observation of a torque plateau using an angular optical trap. Instead we find that the shape of the extension curve can be well explained with a theory that incorporates both DNA twist-stretch coupling and bending fluctuations. This theory also provides a more accurate method of determining the value of the twist-stretch coupling modulus, which has possibly been underestimated in previous studies that did not take into consideration the bending fluctuations. Our study demonstrates the importance of torque detection in the correct identification of phase transitions as well as the contribution of the twist-stretch coupling and bending fluctuations to DNA extension.

Abrupt buckling transition observed during the plectoneme formation of individual DNA molecules.

The response of single DNA molecules to externally applied forces and torques was directly measured using an angular optical trap. Upon overwinding, DNA buckled abruptly as revealed by a sharp extension drop followed by a torque plateau. When the DNA was held at the buckling transition, its extension hopped rapidly between two distinct states. Furthermore, the initial plectonemic loop absorbed approximately twice as much extension as was absorbed into the plectoneme upon each additional turn. The observed extension change after buckling and the postbuckling torque support a recent DNA elasticity model.