Heterochromatin dynamics in vascular smooth muscle plasticity

PROJECT SUMMARY Vascular smooth muscle cells (SMCs) do not terminally differentiate, but retain the remarkable ability to assume a new cellular identity in response to extracellular stimuli. The ability of mature, contractile SMCs to transition to fibroblast/myofibroblast (“synthetic”), macrophage-like, or osteochondrocyte-like phenotypes allows for vascular growth and remodeling, but also contributes to cardiovascular pathologies, including atherosclerosis, restenosis, aneurysm, transplant vasculopathy, pulmonary hypertension, and others. Understanding how SMCs profoundly alter their phenotype has important implications for prevention and treatment of cardiovascular diseases. Cell type-specific gene expression is controlled by DNA distribution in accessible euchromatin and silenced heterochromatin, which is governed by epigenetic modifications of DNA and histones. Our prior work reveals that dynamic epigenetic regulation contributes to the unique plasticity of SMC, identifying TET2 and p300 as key pro-differentiation mediators. We now hypothesize that a uniquely dynamic heterochromatin state allows for SMC fate transitions. Epigenetic marks, including H3K9me3, silence unnecessary gene programs, and H3K9me3 heterochromatin helps maintain cellular identity. SUV39H1 is a histone methyltransferase that generates H3K9me3 heterochromatin marks. Our new preliminary data suggest important roles for SUV39H1 and H3K9me3 in SMC plasticity. We observed SUV39H1-dependent changes in H3K9me3 during SMC phenotypic transitions. 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 vitro. SUV39H1 knockdown induces SMC differentiation-specific genes while suppressing proliferation and migration, and inhibits expression of KLF4, and SUV39H1 represses KDM4A, an enzyme that removes H3K9me3 marks. Notably, PDGF and SUV39H1 mediate reversible H3K9me3 enrichment at key proximal promoter regions in differentiation-specific contractile genes. We will test the overarching hypothesis that unique SUV39H1-dependent heterochromatin dynamics underlie the phenotypic plasticity of vascular SMCs. In Aim 1, we will use high resolution imaging and genome- wide epigenetic analyses to determine whether SMC phenotypic switching involves extensive heterochromatin remodeling, whether H3K9me3 represents a reversible mark that exists outside of heterochromatin, and whether SMCs contain unique subtypes of heterochromatin that allow for accession of alternate gene programs. In Aim 2, we will identify mechanisms that contribute to SUV39H1 regulation and function. In Aim 3, we will study heterochromatin dynamics and SUV39H1-effects in SMC remodeling in vivo.