Protein Arginine Methylation in Vascular Smooth Muscle Cell Phenotypic Modulation and Calcification

Abstract Vascular calcification in blood vessels, stiffens artery and predicts adverse cardiovascular mortality and morbidity. No therapies developed so far directly targeting vascular calcification. Strong evidence has now determined the osteogenic differentiation of vascular smooth muscle cells (VSMC) into “bone-like” cells is critical for the development of vascular calcification. We and others have demonstrated SMC-derived Runx2 (Runt- related transcriptional factor 2) is essential in regulating osteogenic differentiation of VSMC, which induces vascular calcification in atherosclerosis, diabetes and kidney disease. Using single cell RNA sequencing (scRNA-seq) analysis, we discovered a novel Runx2 suppressor and its impact on SMC phenotypic switch in atherosclerosis. Specifically, we determined that the protein arginine methyltransferase 1 (PRMT1) plays a critical role in inhibiting Runx2 and modulating SMC phenotypic switch. PRMT1 is an emerging regulator in human pathological processes, however, its function in vascular calcification and atherosclerosis is entirely unknown. It is thus incumbent upon us to provide additional evidence as to how PRMT1 acts as a new Runx2 suppressor in the modulation of vascular phenotypic switch and calcification. Prompted by the intriguing observations of an inverse correlation between upregulation of Runx2 and marked downregulation of PRMT1 in the calcified atherosclerotic lesions in human and mice, we carried out functional studies using the PRMT1 gain- and loss-of-function VSMC and our novel SMC-specific PRMT1 transgenic mice. Our preliminary studies demonstrated a causative role of PRMT1 in regulating Runx2, SMC phenotypic switch and calcification in vitro; and SMC-specific transgenic PRMT1 inhibited aortic Runx2 and the development of atherosclerosis in vivo in the ApoE-/- mice. Further evidence implicates that PRMT1-directed regulation of Runx2 is mediated through methylation of Runx2. Unbiased proteomics analysis of Runx2 interactome uncovered the interaction of Runx2 with serum response factor (SRF), an essential transcriptional regulator for contractile SMC marker genes, which dysregulates SRF-dependent expression of SMC marker genes. Based on these new and exciting findings, we hypothesize that PRMT1 is a key Runx2 suppressor, which regulates VSMC Runx2 and governs VSMC phenotypic switch and calcification in atherosclerosis. Utilizing the new SMC-specific PRMT1 transgenic mouse model and comprehensive multi-Omics approaches, the proposal will uncover a novel regulatory paradigm highlighting the PRMT1/Runx2 signaling axis in modulating VSMC phenotypic switch and calcification.