Transcription-Coupled Structural Dynamics of Topologically Associating Domains Regulate Replication Origin Efficiency

Metazoan cells only utilize a small subset of the potential DNA replication origins to duplicate the whole genome in each cell cycle. Origin choice is linked to cell growth, differentiation, and replication stress. Despite various genetic and epigenetic signatures have been suggested to define active origins, it remains elusive how the selection of origins is determined. The classic Rosette model proposes that the origins clustered at the center of a chromatin domain are preferentially and simultaneously fired, but direct imaging evidence has been lacking due to insufficient spatial resolution. Here, we applied dual-color stochastic optical reconstruction microscopy (STORM) super-resolution imaging to map the spatial distribution of origins within individual topologically associating domains (TADs). We found that replication generally initiates at the spatial boundary of a TAD at the G1/S transition, in contrary to the Rosette model. Intriguingly, while both active and dormant origins are distributed homogeneously in the TAD during the G1 phase, active origins relocate to the TAD periphery at the G1/S transition. We proved that such origin relocalization is dependent on both transcription and CTCF-mediated chromatin structure. Further, we observed that the replication machinery protein PCNA forms immobile clusters around the TADs at the G1/S transition, which explains why origins at the TAD periphery are preferentially fired. Thus, we propose a “Chromatin Re-organization Induced Selective Initiation” (CRISI) model that the transcription-coupled chromatin structural re-organization determines the selection of replication origins, which transcends the scope of specific genetic and epigenetic signatures for origin efficiency. Our in situ super-resolution imaging unveiled coordination among DNA replication, transcription, and chromatin organization inside individual TADs, providing new insights into the biological functions of sub-domain chromatin structural dynamics.