Extended Data Figure 8. MeCP2 Mini forms droplets in vitro and partitions into heterochromatin condensates in mESCs.

a. Schematic of MeCP2 protein with a minimal MeCP2 protein (Mini) (Tillotson et al., Nature 2017) that retains the MBD and NID displayed below.

b. Droplet experiments examining ability of Mini MeCP2 to form droplets. MeCP2-GFP WT and Mini at 4 μM were added to droplet formation buffers with 125 mM NaCl and 10% PEG-8000.

c. Droplet areas for experiments in Extended Data Fig. 8b. Fields per condition n=12.

d. MeCP2-GFP condensed fraction for experiments in Extended Data Fig. 8b. Mean±SD. Fields per condition n=12.

e. Droplet experiments examining ability of MeCP2 WT and Mini to form droplets with HP1α and DNA. MeCP2-GFP WT or Mini at 7.5 μM was mixed with 7.5 μM HP1α-mCherry and 160 nM DNA in droplet formation buffers with 150 mM NaCl and 10% PEG-8000.

f. HP1α-mCherry partition ratios in MeCP2-GFP WT and Mini droplets for experiments in Extended Data Fig. 8e. Fields per condition n=15.

g. DNA-Cy5 partition ratios in MeCP2-GFP WT and Mini droplets for experiments in Extended Data Fig. 8e. Fields per condition n=15.

h. Droplet experiments examining ability of MeCP2 WT and Mini condensates to enrich TBLR1-CTD. MeCP2-GFP WT or Mini at 4 μM was mixed with TBLR1-CTD-mCherry at 10 μM in droplet formation buffers with 125 mM NaCl and 10% PEG-8000.

i. TBLR1-CTD-mCherry partition ratios in MeCP2-GFP WT and Mini droplets for experiments in Extended Data Fig. 8h. Fields per condition n=12.

j. Live-cell microscopy of endogenous-tagged MeCP2-GFP WT and Mini proteins with Hoechst staining in mESCs.

k. Partition ratios of MeCP2-GFP proteins at heterochromatin condensates for experiments in Extended Data Fig. 8j. Mean±SD, n=10 cells per condition. Two-tailed Student’s t-test: p=0.1161, t=1.6509, df=18.