Animal genomes are folded into loops and topologically associating domains (TADs) by CTCF and loop extruding cohesins, but the live dynamics of loop formation and stability remain unknown. Here, we directly visualize chromatin looping at the Fbn2 TAD in mouse embryonic stem cells using super-resolution live-cell imaging and quantify looping dynamics by Bayesian inference. Unexpectedly, the Fbn2 loop is both rare and dynamic, with a looped fraction of ~3-6.5% and a median loop lifetime of ~10-30 min. Our results establish that the Fbn2 TAD is highly dynamic, where ~92% of the time cohesin-extruded loops exist within the TAD without bridging both CTCF boundaries. This suggests that single CTCF boundaries rather than the fully CTCF-CTCF looped state may be the primary regulators of functional interactions.

Our understanding of the physical principles organizing the genome in the nucleus is limited by the lack of tools to directly exert and measure forces on interphase chromosomes in vivo and probe their material nature. Here, we introduce an approach to actively manipulate a genomic locus using controlled magnetic forces inside the nucleus of a living human cell. We observed viscoelastic displacements over micrometers within minutes in response to near-piconewton forces, which are consistent with a Rouse polymer model. Our results highlight the fluidity of chromatin, with a moderate contribution of the surrounding material, revealing minor roles for cross-links and topological effects and challenging the view that interphase chromatin is a gel-like material. Our technology opens avenues for future research in areas from chromosome mechanics to genome functions.