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- W2912931569 abstract "Recently proposed models on the 3D dynamical structure of chromatin mainly rely on population assays in fixed cells. We here integrate the results of these static assays with the information that can be inferred from live microscopy at the single cell level. We present a single molecule (SM) tracking study of CTCF and Cohesin, central players in the formation of Topologically Associating Domains (TADs). To study the interplay between the two proteins we perform SM tracking of Cohesin in the absence of endogenous CTCF, depleted via the auxine-inducible degron system. With SM tracking, we determine two parameters, the bound fraction of a nuclear factor and its residence time on chromatin, from which the association and dissociation rate can be inferred. The depletion of CTCF slightly affect Cohesin association rate, confirming the hypothesis that CTCF is needed to position Cohesin on chromatin but not for its loading. No significant effect is observed on Cohesin dissociation rate in the absence of CTCF. This result support a scenario in which Cohesin is stalled at CTCF sites but not actively stabilized there. To further characterize Cohesin Dynamics in presence/absence of CTCF, we analyzed the features of long trajectories of Cohesins. For each trajectory we compute the mean-square displacement and extrapolate the apparent diffusion coefficient (Dapp) and the anomalous exponent (alpha). As a reference, we also track genomic loci. When depleting CTCF, a subpopulation of Cohesins show a more dynamic behavior. Given their values of Dapp and alpha, these mobile molecules are most likely bound to chromatin, suggesting the existence of a non-stalled fraction of Cohesins. We are currently testing this hypothesis by comparing our experimental data to computational predictions in the context of the loop extrusion model." @default.
- W2912931569 created "2019-02-21" @default.
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- W2912931569 date "2019-02-01" @default.
- W2912931569 modified "2023-09-30" @default.
- W2912931569 title "Single Molecule Imaging of CTCF and Cohesin. Dissecting the Dynamic Interplay between Chromatin Loop Regulators" @default.
- W2912931569 doi "https://doi.org/10.1016/j.bpj.2018.11.160" @default.
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