PRMT1-methylated MSX1 phase separates to control palate development

Little is known about the regulation and function of liquid‒liquid phase separation (LLPS) in craniofacial developmental disorders, where Msh homeobox 1 (MSX1) and protein arginine methyltransferase 1 (PRMT1) are two important regulators. MSX1 mutations are associated with human cleft palate, the most common craniofacial birth defect. Here, we show that MSX1 LLPS, regulated by PRMT1-catalyzed methylation, is a vertebrate-conserved mechanism underlying the proliferation of embryonic palatal mesenchymal cells (EPMs) and palatal fusion. MSX1 LLPS is triggered by its N-terminal intrinsically disordered protein region (IDR) and regulated by PRMT1-catalyzed methylation, specifically asymmetric dimethylation of arginine (R). Biochemically, PRMT1 methylates MSX1 by dimethylating the residues R150 and R157 in the MSX1 IDR. Physiochemically, the R157 to serine (R157S) mutant, R150S mutant, and hypomethylated MSX1 under PRMT1 deficiency consistently cause abnormal MSX1 LLPS to form less dynamic gel-like condensates. PRMT1 and its methylation target R residues in MSX1 are conserved in vertebrates, from zebrafish to humans. Phenotypically, less dynamic gel-like condensates formed by abnormal MSX1 LLPS cause EPM proliferation defects and cleft palate.In this study, rna-seq was used to reveal the differences between the effects of Msx1 and its mutants on hepm cells. Our findings provide a new paradigm for further exploration of LLPS underlying genetic disorders for future innovation of genetic disorder treatment by direct modulation of LLPS.