Abstract Body:
Introduction: The phenotypic plasticity of vascular smooth muscle cells (VSMCs) is central to growth and remodeling processes, but also contributes to various cardiovascular pathologies, including intimal hyperplasia. We tested the hypothesis that SUV39H1, a histone methyltransferase that generates H3K9me3 repressive heterochromatin-associated epigenetic marks regulates VSMC plasticity. Methods: We employed in vivo ligation and in vitro knockdown, qPCR, western, chromatin immunoprecipitation (ChIP) assays, RNA-Seq and immunofluorescent staining to assess the role of SUV39H1 in heterochromatin dynamics and SMC plasticity. Results: We find that SUV39H1 expression is regulated during SMC phenotypic switching, with low level expression in healthy arteries, but robust induction of SUV39H1 and H3K9me3 following vascular injury, or PDGF treatment in cultured SMCs. Conversely, expression of H3K9me3 demethylase KDM4A, an enzyme that removes H3K9me3 marks, was high in healthy media but decreased over time during progression of intimal hyperplasia. We further find that SUV39H1 knockdown induces SMC differentiation-specific genes and contractility while suppressing proliferation and migration, and inhibits mRNA stability and expression of KLF4, a master dedifferentiation factor. PDGF-BB induced SUV39H1 expression and SUV39H1-dependent H3K9me3 modification of contractile gene promoters in hCASMC. SUV39H1 knockdown reduced the repressive H3K9me3 and 5mC marks but increased the H3K27Ac mark at these promoters. SUV39H1 knockdown also increased KDM4A expression and binding to contractile promoters, suggesting an opposing regulatory relationship between the writer and eraser of H3K9me3 in phenotypic switching. Immunostaining with IMARIS 3D rendering noted H3K9me3 marks were primarily localized to the nuclear periphery with isolated interior puncta in control VSMCs. Notably, PDGF-BB treatment induced a SUV39H1-dependent relocalization of these marks toward the nuclear interior. Conclusion: SUV39H1 is an epigenetic regulator that alters chromatin state and stabilizes KLF4 expression, promoting VSMC dedifferentiation. We report dynamic SUV39H1-dependent regulation of multiple chromatin marks at contractile gene promoters, and opposing regulation of the enzymes that write (SUV39H1) and erase (KDM4A) H3K9me3 heterochromatin marks during VSMC phenotypic switching, suggesting that coordinated regulation of histone and DNA methylation contribute to VSMC phenotypic plasticity.