SUV39H1 mediated regulation of KLF4 and KDM4A coordinate smooth muscle cell phenotypic plasticity

Abstract

BackgroundReversible DNA methylation contributes to the phenotypic plasticity of vascular smooth muscle cells (VSMCs). This plasticity contributes to vascular growth remodeling, but also underlies cardiovascular pathologies, including intimal hyperplasia. We investigated the role of SUV39H1, a histone methyltransferase that generates the H3K9me3 repressive epigenetic mark, in VSMC plasticity MethodsWe applied knockdown, qPCR, western blotting, chromatin immunoprecipitation (ChIP) assays, and RNA-Seq in human coronary artery SMCs (hCASMCs), and murine carotid ligation to determine the role of SUV39H1 in VSMC plasticity. ResultsExpression of SUV39H1 and the H3K9me3 mark it generates increase, whereas the cognate H3K9me3 demethylase KDM4A decreases, over time during the progression of murine intimal hyperplasia following carotid artery ligation, with marked elevation of SUV39H1 and H3K9me in the neointima. SUV39H1 knockdown induced contractile genes and contractility while decreasing migration and proliferation in hCASMCs. Transcriptomic analysis confirmed that SUV39H1 promotes SMC dedifferentiation. SUV39H1 knockdown revealed that SUV39H1 promotes KLF4 upregulation by increasing KLF4 mRNA stability. 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 activating H3K27Ac mark at these promoters. SUV39H1 knockdown also increased expression of KDM4A and its binding to contractile promoters, suggesting an opposing regulatory relationship between the writer and eraser of H3K9me3. ConclusionsWe identify SUV39H1 as an epigenetic regulator that promotes VSMC dedifferentiation by stabilizing KLF4 expression and by altering chromatin state. We report SUV39H1-dependent dynamic regulation of the repressive H3K9me3 mark at contractile gene promoters, and opposing regulation of the enzymes that write (SUV39H1) and erase (KDM4A) these marks during VSMC phenotypic switching. These studies suggest that coordinate regulation of both histone and DNA methylation contribute to VSMC phenotypic plasticity. HighlightsO_LIThe histone methyl transferase SUV39H1 is differentially regulated in VSMC phenotypic switching in culture and in vascular remodeling. SUV39H1 and the H3K9me3 mark it generates increase, whereas the cognate H3K9me3 eraser KDM4A decreases, during the progression of murine intimal hyperplasia. C_LIO_LISUV39H1 promotes PDGF-induced VSMC de-differentiation, stabilizing KLF4 mRNA. SUV39H1 opposes VSMC contractility and promotes proliferation and migration. C_LIO_LIPDGF-BB induces SUV39H1-dependent H3K9me3 marks at VSMC contractile gene promoters, and SUV39H1 loss of function alters multiple marks that govern chromatin accessibility at these promoters, with opposing effects on the repressive H3K9me3 and 5mC and activating H3K27Ac marks. C_LIO_LISUV39H1 generates H3K9me3 marks that are associated with silenced heterochromatin. We note dynamic regulation of this mark at VSMC contractile gene promoters, and opposing regulation of the enzymes that write (SUV39H1) and erase (KDM4A) these marks during VSMC phenotypic switching. C_LI