Topology-dependent anomalous dynamics of ring and linear DNA are sensitive to cytoskeleton crosslinking

Abstract

Cytoskeletal crowding plays a key role in the diffusion of DNA molecules through the cell, acting as a barrier to effective intracellular transport and conformational stability required for such processes as transfection, viral infection, and gene therapy. Here we elucidate the transport properties and conformational dynamics of linear and ring DNA molecules diffusing through entangled and crosslinked composite networks of actin and microtubules. We couple single-molecule conformational tracking with differential dynamic microscopy to reveal that ring and linear DNA exhibit surprisingly distinct transport properties that are influenced differently by cytoskeleton crosslinking. Ring DNA coils are swollen and undergo heterogeneous and biphasic subdiffusion that is hindered by crosslinking. Conversely, crosslinking actually facilitates the single-mode subdiffusion that compacted linear chains exhibit. Our collective results demonstrate that transient threading by cytoskeleton filaments plays a key role in the dynamics of ring DNA, whereas the mobility of the cytoskeleton dictates transport of linear DNA.

Anomalous and heterogeneous DNA transport in biomimetic cytoskeleton networks

“…Ring DNA assumes a greater range of conformational states than linear DNA, as demonstrated by an increased FWHM (Fig 5). There is ample literature, both experimental 26,43,44,48 and theoretical, [45][46][47][48] suggesting that ring polymers entangled by linear chains assume folded, amoeba-like, or threaded conformations not accessible to linear molecules, leading to anomalous subdiffusion and coil swelling, in alignment with our results.…”

“…All three networks, characterized previously, are considered isotropic with no phase separation between proteins. 26,27,87 The mesh size of all composite networks is ξ ≈ 0.81 μm. 27…”

“…Finally, we compute the non-Gaussianity parameter, β NG t = have been described and validated previously. 25,26,30,32,88…”

“…34,35 This diffusive process, known as constraint release, [52][53][54] is extremely slow compared to reptation and is likely largely eliminated if the linear chains are crosslinked. 26 However, experimental evidence of these predicted diffusive processes, especially for biologically relevant polymers and systems, is sparse. 26,47,48,55 The most well understood characteristic of diffusive motion in homogenous media is the linear growth of the mean-squared displacement (MSD) with time, i.e.…”

“…26 However, experimental evidence of these predicted diffusive processes, especially for biologically relevant polymers and systems, is sparse. 26,47,48,55 The most well understood characteristic of diffusive motion in homogenous media is the linear growth of the mean-squared displacement (MSD) with time, i.e. MSD ∝ t α where α = 1.…”