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Diffusion of DNA-Binding Species in the Nucleus: A Transient Anomalous Subdiffusion Model.

ABSTRACT: Single-particle tracking experiments have measured escape times of DNA-binding species diffusing in living cells: CRISPR-Cas9, TetR, and LacI. The observed distribution is a truncated power law. Working backward from the experimental results, the observed distribution appears inconsistent with a Gaussian distribution of binding energies. Working forward, the observed distribution leads to transient anomalous subdiffusion, in which diffusion is anomalous at short times and normal at long times, here only mildly anomalous. Monte Carlo simulations are used to characterize the time-dependent diffusion coefficient D(t) in terms of the anomalous exponent α, the crossover time t_cross, and the limits D(0) and D(∞) and to relate these quantities to the escape time distribution. The simplest interpretations identify the escape time as the actual binding time to DNA or the period of one-dimensional diffusion on DNA in the standard model combining one-dimensional and three-dimensional search, but a more complicated interpretation may be required. The model has several implications for cell biophysics. 1) The initial anomalous regime represents the search of the DNA-binding species for its target DNA sequence. 2) Non-target DNA sites have a significant effect on search kinetics. False positives in bioinformatic searches of the genome are potentially rate-determining in vivo. For simple binding, the search would be speeded if false-positive sequences were eliminated from the genome. 3) Both binding and obstruction affect diffusion. Obstruction ought to be measured directly, using as the primary probe the DNA-binding species with the binding site inactivated and eGFP as a calibration standard among laboratories and cell types. 4) Overexpression of the DNA-binding species reduces anomalous subdiffusion because the deepest binding sites are occupied and unavailable. 5) The model provides a coarse-grained phenomenological description of diffusion of a DNA-binding species, useful in larger-scale modeling of kinetics, FCS, and FRAP.

SUBMITTER: Saxton MJ

PROVIDER: S-EPMC7203007 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Neural tissue is a hierarchical multiscale system with intracellular and extracellular diffusion compartments at different length scales. The normal diffusion of bulk water in tissues is not able to detect the specific features of a complex system, providing nonlocal, diffusion measurement averaged on a 10-20 μm length scale. Being able to probe tissues with sub-micrometric diffusion length and quantify new local parameters, transient anomalous diffusion (tAD) would dramatically increase the diagnostic potential of diffusion MRI (DMRI) in detecting collective and sub-micro architectural changes of human tissues due to pathological damage. In DMRI, the use of tAD parameters quantified using specific DMRI acquisition protocols and their interpretation has often aroused skepticism. Although the derived formulas may accurately fit experimental diffusion-weighted data, the relationships between the postulated dynamical feature and the underlying geometrical structure remains elusive, or at most only suggestive. This work aimed to elucidate and validate the image contrast and information that can be obtained using the tAD model in white matter (WM) through a direct comparison between different diffusion metrics and histology. Towards this goal, we compared tAD metrics extracted from pure subdiffusion (α-imaging) and super-pseudodiffusion (γ-imaging) in excised mouse spinal cord WM, together with T2 and T2* relaxometry, conventional (normal diffusion-based) diffusion tensor imaging (DTI) and q-space imaging (QSI), with morphologic measures obtained by optical microscopy, to determine which structural and topological characteristics of myelinated axons influenced tAD contrast. Axon diameter (AxDiam), the standard deviation of diameters (SD ax.diam ), axonal density (AxDens) and effective local density (ELD) were extracted from optical images in several WM tracts. Among all the diffusion parameters obtained at 9.4 T, γ-metrics confirmed a strong dependence on magnetic in-homogeneities quantified by R2* = 1/T2* and showed the strongest associations with AxDiam and ELD. On the other hand, α-metrics showed strong associations with SD ax.diam and was significantly related to AxDens, suggesting its ability to quantify local heterogeneity degree in neural tissue. These results elucidate the biophysical mechanism underpinning tAD parameters and show the clinical potential of tAD-imaging, considering that both physiologic and pathologic neurodegeneration translate into alterations of WM morphometry and topology.

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Diffusion of DNA-Binding Species in the Nucleus: A Transient Anomalous Subdiffusion Model.

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