Fig. 2: PRC1 is highly dynamic with a small stably chromatin-bound fraction.

a Example cropped frames from a 67 Hz exposure SPT movie (n = 55) showing a single RING1B molecule. Tracked steps of the molecule between frames are superimposed in red. Scale bar = 1 μm. b Example tracks of individual molecules from a single 15 ms exposure RING1B SPT movie out of 55 analysed, showing a range of diffusive behaviours. Scale bar = 3 μm. c A box plot indicating the percentage of RING1B, H2B and HaloTag-NLS molecules in the bound state determined from SPT using Spot-On. Box plots represent measurements from n > 50 movies each from 2 experiments. Source data are provided as a Source Data file. d Example frames from a 2 Hz exposure SPT movie (n = 24) showing a single molecule visible for 100 s indicated by white arrowheads. Scale bar = 1 μm. e Dwell time distributions (1—cumulative distribution function, CDF) and fitted biexponential decay curves for immobile RING1B molecules for a representative set of movies from a single experiment acquired as in d. H2B is included as a photobleaching control for highly stable binding. n = 24 movies across 3 experiments. Source data are provided as a Source Data file. f Example frames from a 0.033 Hz exposure SPT movie (n = 10) showing a single molecule visible for 300 s indicated by white arrowheads. Scale bar = 1 μm. g Dwell time distributions (1—cumulative distribution function, CDF) and fitted biexponential decay curves for immobile RING1B molecules for a representative set of movies from a single experiment acquired as in f. H2B is included as a photobleaching control for highly stable binding. n = 10 movies across 2 experiments. Source data are provided as a Source Data file. h Schematic of Polycomb bodies in the nucleus (left panel). Representative maximum intensity projection of nuclear RING1B-HT-JF₅₄₉ signal acquired by spinning disk microscopy (n = 63 cells). The yellow arrowhead indicates a single Polycomb body. Scale bar = 5 μm (right panel). i FRAP recovery curves for RING1B-HaloTag in regions containing a Polycomb body (blue) or elsewhere in the nucleus (Non-Polycomb body, red). The recovered fraction was measured relative to initial fluorescence intensity and corrected using an unbleached region. The error bars denote SEM for n = 20 cells each for Polycomb body and Non-Polycomb body regions across 2 experiments. Source data are provided as a Source Data file. j A box plot illustrating the relative mean fluorescence signal per unit volume of all RING1B Polycomb bodies in each cell (Body, blue) compared to the remaining nuclear volume in the same cell (Non-Body, red), normalised to the median non-Polycomb body fluorescence. n = 63 cells across 2 experiments. Source data are provided as a Source Data file. b, j Boxes represent the interquartile range (IQR), the middle line corresponds to the median, and whiskers extend to the largest and smallest values no more than 1.5 x IQR from the box. Values outside of this range are not plotted, but are included in all analyses.