biostudies-literatureKefalopoulou SEC | EU Framework Programme for Research and Innovation H2020 | H2020 Priority Excellent Science | H2020 Marie Skłodowska-Curie Actions (H2020 Excellent Science - Marie Skłodowska-Curie Actions)Swiss National Science FoundationEuropean Research CouncilDutch Research Council (NWO)Human Frontier Science Program (HFSP)1186-1198https://www.ebi.ac.uk/biostudies/studies/S-EPMC12270907biostudies-literatureUnknown27(7)The regulation of gene expression is governed at multiple levels of chromatin organization. However, how gene regulation is co-ordinated remains relatively unexplored. Here we develop Dam&ChIC, a method that enables retrospective and multifactorial chromatin profiling in single cells. Dam&ChIC employs chromatin labelling in living cells with ^m6A to acquire a past chromatin state, coupled with an antibody-mediated readout to capture the present chromatin state. Analyses of diverse factor combinations highlight its versatility and superior resolution. By tracking lamina-associated domain inheritance over the cell cycle, we showcase that Dam&ChIC provides retrospective single-cell chromatin data. When applied in random X chromosome inactivation, Dam&ChIC disentangles the temporal oNature cell biologyRetrospective and multifactorial single-cell profiling reveals sequential chromatin reorganization during X inactivation.PMC12270907184.034.019798573016.161.339NWO-CW714.016.001101053581174991LT000209/2018-LKefalopoulou SKorthout TZeller Pvan Oudenaarden AKind JRullens PMJde Vries SSde Luca KLfalseRetrospective and multifactorial single-cell profiling reveals sequential chromatin reorganization during X inactivation.The regulation of gene expression is governed at multiple levels of chromatin organization. However, how gene regulation is co-ordinated remains relatively unexplored. Here we develop Dam&ChIC, a method that enables retrospective and multifactorial chromatin profiling in single cells. Dam&ChIC employs chromatin labelling in living cells with ^m6A to acquire a past chromatin state, coupled with an antibody-mediated readout to capture the present chromatin state. Analyses of diverse factor combinations highlight its versatility and superior resolution. By tracking lamina-associated domain inheritance over the cell cycle, we showcase that Dam&ChIC provides retrospective single-cell chromatin data. When applied in random X chromosome inactivation, Dam&ChIC disentangles the temporal o2025-01-01T00:00:00Z2025 Jul2026-07-15T10:55:52.254Z2025-08-14T03:05:38.43ZS-EPMC122709074064050110.1038/s41556-025-01687-w