Visualizing the Formation and Collapse of DNA Toroids

Abstract

In living organisms, DNA is generally confined into very small volumes. In most viruses, positively charged multivalent ions assist the condensation of DNA into tightly packed toroidal structures. Interestingly, such cations can also induce the spontaneous formation of DNA toroids in vitro. To resolve the condensation dynamics and stability of DNA toroids, we use a combination of optical tweezers and fluorescence imaging to visualize in real-time spermine-induced (de)condensation in single DNA molecules. By actively controlling the DNA extension, we are able to follow (de)condensation under tension with high temporal and spatial resolution. We show that both processes occur in a quantized manner, caused by individual DNA loops added onto or removed from a toroidal condensate that is much smaller than previously observed in similar experiments. Finally, we present an analytical model that qualitatively captures the experimentally observed features, including an apparent force plateau.

Electrostatic interactions in biological DNA-related systems

“…Similar to unwrapping of DNA toroids, that enabled to rationalize their stability, 124 recent measurements of forceinduced stretching of chromatin fibers revealed the NCP-NCP cohesive energy of E3.4 332 and later E14 k B T, 333 depending on concentrations of mono-and divalent cations. The ionic conditions dramatically influence also the forces required to disrupt the NCP-NCP contacts in chromatin fibers (typically B2-5 pN) and to induce DNA unwrapping from NCPs (B6-20 pN).…”

“…Recent single-molecule optical tweezers manipulation experiments have enabled to decipher the physical mechanisms behind the toroidal stability and DNA condensation dynamics. 124 In particular, a step-wise unwrapping of DNA from toroids by the applied force has been detected, corresponding to multiple DNA loops released from the condensate. The number of turns released was shown to be a function of the applied tension of 1-10 pN and salt-mediated DNA-DNA attraction.…”

How a short double-stranded DNA bends

“…Although the uniform bending assumption is valid for certain situations such as DNA wound around histones and DNA collapsed into a toroid, 35 it is not generally true for linear fragment under high bending conditions. To incorporate non-uniform bending in strong coupling with the bubble in a simplest way, we assume that the DNA is bent within the ss and ds regions, respectively, with different segmental bending angles, θ s and θ d .…”

DNA intercalation optimized by two-step molecular lock mechanism

“…However, in single molecule experiments the ends of the DNA molecule are pulled apart, preventing the molecule from dropping out of solution when condensed, and preventing condensation at high pulling forces18. In addition, in single molecule experiments a much smaller concentration of ligands is required to observe significant binding, which also greatly diminishes DNA condensation19.…”