Diffusion and Conformational Dynamics of Linear and Circular DNA in Crosslinked Cytoskeleton Composites

In order to carry out key cellular processes, DNA molecules must diffuse through the crowded cytoskeletal network, comprised of thin semiflexible actin filaments and thick rigid microtubules. Each of these cytoskeletal filaments can also be crosslinked to varying degrees, altering the network structure and dynamics and hence the impact on DNA diffusion and conformational dynamics. Here, we use single-molecule conformational tracking to investigate the effect of cytoskeleton crosslinking on the transport properties and time-varying conformations of diffusing DNA molecules. Specifically, we track the center-of-mass, size, and shape of single linear and circular DNA molecules diffusing in crosslinked composites of actin and microtubules. We quantify transport coefficients, anomalous diffusion scaling exponents, lengths and timescales of intramolecular fluctuations, and shape and size dynamics of DNA. We determine the role of DNA topology (circular vs linear) as well as cytoskeleton crosslinking motif on DNA dynamics and conformations. Our results shed light on how macromolecular topology and network structure impact macromolecular mobility in crowded cell-like environments.