Average local nucleosome motion remains nearly constant during interphase in living human cells

Dynamic chromatin behavior, which is related to chromatin accessibility, plays a critical role in various genome functions such as RNA transcription and DNA replication/repair. Previous studies using highly synchronized cells showed that average local chromatin motion, captured by single-nucleosome imaging on a subsecond time scale, remained nearly constant throughout G1, S, and G2 phases in living human cells. Here, we combined single-nucleosome imaging with Fucci cell-cycle probes to test this finding in asynchronous living human cells. Using HeLa and HCT116 cells expressing H2B-HaloTag and Fucci probes, we found that local nucleosome motion remained similar on average throughout interphase. Consistently, H3.3-Halo-labeled nucleosomes, which are enriched in Hi-C A compartments (euchromatin), also showed near-constant motion throughout interphase. Transcription inhibition increased nucleosome motion throughout interphase. Local nucleosome motion also increased following cellular perturbations, such as replication stress or DNA damage. Our findings suggest that near-constant chromatin motion supports housekeeping functions under similar physical conditions during interphase. They also suggest that cells can transiently change chromatin motion to perform ad hoc tasks in response to intra- and extracellular signals, such as DNA damage.