Fig. 2: PRMT1 regulates MSX1 phase separation by dimethylating MSX1.

A Representative confocal images of MSX1-mEGFP-expressing HEK293T treated with DMSO or MS023 (left), and quantitative analysis of circularity (sphericity) of MSX1-mEGFP puncta per nucleus was shown (right). Scale bars, 10 μm. DMSO: n =  115, MS023: n = 216 condensates. B Representative confocal images (left) and circularity quantification (right) of MSX1-mEGFP-expressing HEK293T transfected with NC siRNA or PRMT1 siRNA. Scale bars, 10 μm. siNC: n = 105, siPRMT1: n = 310 condensates. C Representative images of FRAP of MSX1-mEGFP- expressing HEK293T treated with DMSO or MS023. Scale bars, 10 μm. D Representative graphs of FRAP of MSX1-mEGFP-expressing HEK293T transfected with NC or PRMT1 siRNA. Scale bars, 10 μm. E FRAP quantification of MSX1-mEGFP-expressing in HEK293T treated with DMSO or MS023. n = 3 biologically independent experiments. F Quantitative of FRAP assay in HEK293T transfected with NC or PRMT1 siRNA. n = 3 biologically independent experiments. G Representative immunofluorescence images for endogenous MSX1 and PRMT1 in HEK293T. Scale bars, 10 μm. H Representative western blot images of reciprocal Co-IP analysis confirming the association between MSX1 and PRMT1 in HEK293T ectopically expressing MSX1-mEGFP and PRMT1-FLAG. I Representative western blot images of pulldown assay using MSX1-mEGFP and PRMT1-mCherry purified proteins. J, K Confocal images (J) and colocalization analysis (K) of HEK293T with ectopic overexpression of MSX1-mEGFP and PRMT1-mCherry. Scale bars, 10 μm. The white solid line indicates the nucleus. L Representative DIC and fluorescence images of condensates formed by purified proteins MSX1-mEGFP and PRMT1-mCherry. Scale bars, 10 μm. M Representative western blot images of Co-IP assays assessing the aDMA and sDMA of MSX1. N Representative western blot images of Co-IP assays assessing the aDMA of MSX1 in treated with MS023 or PRMT1 siRNA. O Representative western blot images of in vitro methylation assay using MSX1 and PRMT1 purified proteins with/without S-adenosylmethionine (SAM) treatment. Coomassie blue-stained gel showed loading controls for MSX1 and PRMT1 purified proteins. P Droplet formation (left) and turbidity measurement (right) of unmethylated and methylated MSX1 purified proteins. Scale bars, 10 μm. n = 10 samples. 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 (A, B, E, F, P). Source data are provided as a Source Data file.