Nascent CUT&Tag captures transcription factor binding after chromatin duplication

Abstract DNA replication strips off all chromatin proteins, which must be reassembled behind the replication fork. To track chromatin reassembly on newly synthesized DNA, we developed Nascent CUT&Tag, a chromatin profiling method that directly measures transcription factor (TF) binding on nascent chromatin. We tracked the recovery of GAGA factor (GAF) and Pleiohomeotic (PHO) in Drosophila Kc167 cells. Whereas both TFs are evicted following passage of the replication fork, GAF recovers on newly synthesized DNA over a minutes-to-hours range, whereas PHO generally requires hours to fully reestablish binding. Early recovering GAF peaks are characterized by shorter GAF motifs and are associated with functions related to cell cycle progression. Conversely, late recovering peaks are characterized by longer, degenerate GAF motifs and are associated with developmental functions. GAF recovery on newly synthesized DNA requires chromatin remodeling by Brahma Associated Factor (BAF), implying that nucleosome turnover is critical to fully reestablish GAF binding. Competing Interest Statement The authors have declared no competing interest. Footnotes This version now includes PHO nascent chromatin data.