library_construction_protocol: After 48 hours, cells were collected by replacing the culture media with trypsin for <5 minutes, then resuspending the detached cells in growth media and centrifuging for 5 minutes at 300 g. The cells were first washed once in PBS, then crosslinked for 10 minutes at room temperature in PBS with 1% (v/v) formaldehyde. The formaldehyde was quenched by adding glycine to a final concentration of 0.125 M, and was then washed twice in PBS. Cell pellets containing an equal number of cells in each sample were snap frozen with liquid nitrogen and stored at -80°C. The cell pellets were thawed on ice and lysed in lysis buffer (100 mM NaCl, 0.5% (v/v) SDS, 5 mM EDTA, 50 mM Tris-HCl pH 8 and 1X protease inhibitor cocktail (Sigma #4693132001)) on ice, then centrifuged for 6 minutes at 300 g. The resulting chromatin pellet was resuspended in ChIP buffer (a 2:1 ratio of lysis buffer and dilution buffer (100 mM NaCl, 5% (v/v) Triton X-100, 5mM EDTA, 100mM Tris-HCl pH 8.5 and 1X protease inhibitor cocktail (Sigma #4693132001))). For every 1x107 cells, chromatin from 5x106 cross-linked NTERA-2 cells were added. All samples were then sonicated at 4°C in a Bioruptor set to high, with 30 cycles of 30 s on and 30 s off. The sonicated samples were diluted to 1x107 cells per mL, and 1mL per antibody was used for ChIP, with 1% kept as input. Antibodies were added to each sample as specified and incubated overnight at 4 °C rotating at low speed. After the antibody incubation, the lysate was centrifuged at max speed for 30 minutes at 4 °C. The pellet was discarded and 40 µL of Dynabeads Protein G (Thermo #10003D), washed 5 times in ChIP buffer, were added to the supernatant of each sample and incubated for 2-4 hours at 4°C rotating at low speed. The beads were then washed 3 times in mixed micelle buffer (150 mM NaCl, 20 mM Tris-HCl pH 8, 5 mM EDTA, 5.2% (w/v) sucrose and 1% (v/v) Triton X-100), twice in buffer 500 (0.1% (w/v) sodium deoxycholate, 1 mM EDTA, 50 mM HEPES pH 7.5 and 1% (v/v) Triton X-100), twice in LiCl/detergent wash buffer (0.5% (w/v) sodium deoxycholate, 1 mM EDTA, 250 mM LiCl, 0.5% (v/v) NP-40, 10 mM Tris-HCl pH 8) and once in TE (10 mM Tris-HCl pH 8 and 1 mM EDTA). The beads were then resuspended in 100 µL elution buffer (1% (v/v) SDS and100 mM NaHCO3) and incubated at 65°C for 1 hour with shaking. The eluate was removed from the beads, and 10 µL of the input was combined with 90 µL of elution buffer, then all samples were reverse-crosslinked by shaking at 65 °C overnight. 1 µL of RNase A (Thermo #EN0531) was then added to all samples and incubated shaking at 37 °C for 1 hour, followed by 1 µL of Proteinase K (NEB #P8107S) with 2 hours of shaking at 55 °C. DNA was purified from these samples using the Qiagen MinElute PCR purification kit (Qiagen #28006) according to the instructions of the manufacturer. The purified DNA was dried overnight at room temperature in a biosafety hood before shipment to Azenta for Illumina sequencing with 20 million paired-end reads of 150bp per library. The antibodies used for ChIP included: anti-SUZ12 (Cell Signalling 3737, 5 µL), anti-H3K27me3 (Cell Signalling 35861SF, 3 µL) anti-Jarid2 (Cell Signalling 13594S, 5 µL), and anti-MTF2 (Proteintech 16208-1-AP, 5 µL). All libraries were constructed by Azenta. In brief, the starting material was DNA (100 - 1000ng) that passed quality control. DNA underwent end repair, dA-tailing, adapter ligation and purification. The purified DNA underwent size selection before PCR amplification for library construction. The preliminary quantification and dilution of the library was performed using Qubit3.0, and then Agilent 2100 was used to determine the insert size and nucleic acid concentration of the resulting library sample. The effective concentration of each sample library in the library mixture was determined by qPCR before sequencing to ensure the accuracy of the sample concentration and the reliability of the sequencing data. ChIP-Seq (with spike-in normalisation)