Fig. 1: MSX1 undergoes phase separation via an IDR at its N-terminus.
From: PRMT1-methylated MSX1 phase separates to control palate development
A Order/disorder region prediction of MSX1 by the protein sequence and disorder prediction (PONDR) algorithm. B, C Representative immunofluorescence images for endogenous MSX1 in HEK293T (B) and HEPM (C). Scale bars, 10 μm. D, E Representative confocal images of HEK293T (D) and HEPM (E) with ectopic overexpression of MSX1-mEGFP. Scale bars, 10 μm. F Representative fluorescence images of MSX1-mEGFP expressing HEK293T and HEPM treated with dimethyl sulfoxide (DMSO) or 5% 1,6-Hex (left), and quantification of liquid droplets formed in the indicated cells. Scale bars, 10 μm. HEK293T: n = 19. HEPM: −1,6-hex, n = 17; +1,6-hex, n = 13. G Representative graphs and quantitative analysis for the FRAP assay of MSX1-mEGFP condensates in HEK293T and HEPM. Scale bars, 10 μm. n = 3 biologically independent experiments. H Time-lapse micrographs of MSX1-mEGFP droplet fusion in HEPM. Scale bars, 10 μm. I Representative images of droplet formation of MSX1-FL-mEGFP and MSX1 IDR-mEGFP upon 3C cleavage. Scale bars, 10 μm. J Phase diagram of MSX1-mEGFP at different protein concentrations as a function of salt concentration, achieved by scoring for the existence of droplets or fibrillar hydrogels. Scale bars, 10 μm. K Representative images for the turbidity assay of MSX1-mEGFP protein solution with increasing protein concentration. L Representative images and quantification of MSX1-mEGFP and MSX1 protein solution treated with DMSO or 5% 1, 6-Hex. Scale bars, 10 μm. MSX1-mEGFP: n = 3, MSX1: n = 4 biologically independent experiments. M Representative droplet micrographs (top) and FRAP quantification (bottom) of MSX1-mEGFP protein in vitro. Scale bars, 10 μm. n = 4 biologically independent experiments. N Time-lapse micrographs of MSX1-mEGFP droplet fusion in vitro. Scale bars, 5 μm. O Schematic of MSX1 domains and truncated mutants. P Phase separation assay of truncated mutants of MSX1 in cells. Scale bars, 10 μm. Q Representative images of droplet formation for MSX1-FL-mEGFP or MSX1-ΔIDR-mEGFP at different protein concentrations in 25–200 mM NaCl. Scale bars, 10 μm. R Statistical analysis of the droplet formation of MSX1-FL-mEGFP or MSX1-ΔIDR-mEGFP. n = 3 biologically independent experiments. All data in this figure are represented as mean ± SD from at least three biologically independent experiments with similar results. Two-tailed Student’s t-test for (F, L). Source data are provided as a Source Data file.
