Supplementary Figure 2: Evidence for phase-separation potential of MegaTrans components.
From: Phase separation of ligand-activated enhancers licenses cooperative chromosomal enhancer assembly
a. Computational analysis of amino acid sequences of indicated proteins using Predictor of Natural Disordered Region (PONDR). Score of >0.5 is predicted to be disordered domain b. Droplet formation by purified recombinant GATA3-MBP and ERα-MBP. The protein concentration in each case was 7 µM in 25 mM Tris buffer, pH 7.5 containing 5% PEG 8000 and 200 mM NaCl. MBP-alone do not form droplet under similar conditions c. Distribution of aspect ratios of GAvTA3-MBP and ERα-MBP protein droplets. Boxes represent interquartile ranges (IQRs). Whisker represents points in lower and upper quartiles within 1.5 IQR from lower and upper edges of IQR. d. GATA3-MBP and ERα-MBP concentrate in single droplets indicating miscibility. Plots represent the fluorescent intensity profiles of the two proteins across within a GATA3-ERα droplet formed in 25 mM Tris buffer (pH 7.5) containing 500 mM NaCl. e, f, g. Representative example of optodroplet assay in MCF7 cells expressing (e) mCherry-Cry2 (f) GATA3-IDR-mCherry-Cry2 (g) ERα-IDR-mCherry-Cry2. Cry2 was activated with 488 nm laser light for the indicated period of time.
