scWGS analysis of HCT116 exposed to mild replication stress passing through a single-cell cycle with and without BLM helicase

The BLM helicase is a critical genome maintenance factor involved in diverse cellular processes including DNA replication, repair, transcription, and chromosome segregation. During mitosis, BLM is known to cooperate with the PICH helicase and topoisomerases TOP2A and TOP3A at ultrafine DNA bridges (UFBs)—non-chromatinized DNA structures that link sister chromatids, but the molecular mechanisms of UFB resolution remain incompletely defined. Here, we endogenously tagged BLM and PICH with fluorescent proteins and auxin-inducible degrons to generate a live-cell imaging system that enables temporal tracking of UFB dynamics in the presence or absence of BLM. Time-resolved lattice light sheet microscopy revealed the dynamic localization patterns of BLM and PICH throughout the cell cycle. While BLM cycles between PML bodies and DNA repair foci in interphase, it dissociates from chromatin at mitotic entry, and re-associates during anaphase to UFBs as well as to CENP-B-positive mitotic foci. Acute BLM depletion during mitosis increased the fraction of unresolved UFBs, micronuclei containing acentric fragments, binucleation, and resulted in subtle karyotypic abnormalities detected by single-cell whole-genome sequencing. These findings highlight a mitosis-specific role for BLM in UFB resolution and underscore its function in preserving genomic stability.