Epigenetic Control of Smooth Muscle Cell Lineage and Phenotypic Switching

描述（由申请人提供）：已知平滑肌细胞（SMC）分化状态的改变控制或“表型转换”在许多主要人类疾病的发生和/或进展中起关键作用，包括动脉粥样硬化、哮喘和血管成形术后再狭窄。然而，控制SMC表型转换的机制尚不清楚。本研究的重点是确定染色质组蛋白模式的机制，这是高等真核细胞的一个关键表观遗传控制，调节SMC在发育和疾病中的分化。重要的是，在目前的资助期内，我们完成了一系列开创性的研究，表明胚胎干细胞（ESC）的SMC发育与SMC标记基因位点上独特的组蛋白修饰模式相关，该模式将SMC与非SMC区分开来，并使这些位点允许转录激活。相反，SMC在血管损伤或PDGF BB治疗时的表型转换与许多SMC选择性组蛋白修饰的丧失以及与转录沉默/染色质浓缩相关的组蛋白变化的获得有关。然而，SMC标记基因位点上的H3K4去甲基化（一种出现在ESC向SMC发展过程中的组蛋白变化）在所有SMC表型转换模型中完全不变，这表明它可能相对“固定”，并有助于在可逆表型转换过程中保存SMC“谱系记忆”。综上所述，研究结果表明，组蛋白修饰存在一种独特的模式，将SMC与非SMC区分开来，这些SMC和基因位点特异性的表观遗传修饰可能在发育和疾病中调节SMC分化标记基因的表达中发挥关键作用。这个项目的重点是验证这样一个假设，即从胚胎干细胞发育SMC与SMC标记（和调节）基因位点上获得一种独特的组蛋白修饰模式有关，这些组蛋白修饰在决定这些基因位点转录激活的许可性以及在可逆表型转换期间提供“SMC谱系记忆”方面起着关键作用。我们将通过解决以下两个具体目标来解决这个假设。目的1是确定SMC选择性/特异性染色质修饰在多能干细胞谱系发育过程中调控SMC分化标记和调控基因表达的机制。这将包括首次直接测试特定组蛋白修饰在控制SMC谱系/分化中的作用，以及这些组蛋白修饰在发育过程中如何获得的研究。目的2是确定表观遗传修饰在体内血管损伤或PDGF BB治疗培养SMC时介导血管SMC可逆表型转换中的作用。研究将确定控制SMC分化的基本机制，并可能导致治疗SMC表型转换起主要作用的疾病的新疗法。

DESCRIPTION (provided by applicant): Altered control of the differentiated state of the smooth muscle cell (SMC) or "phenotypic switching" is known to play a critical role in the development and/or progression of a number of major human diseases including atherosclerosis, asthma, and post-angioplasty restenosis. However, the mechanisms that control SMC phenotypic switching are poorly understood. The focus of this proposal is to determine mechanisms by which histone patterning of chromatin, a key epigenetic control in higher eukaryotic cells, regulates SMC differentiation in development and disease. Of major significance, during the current funding period, we completed a series of pioneering studies showing that development of SMC from embryonic stem cells (ESC) is associated with acquisition of a unique pattern of histone modifications at SMC marker gene loci that distinguish them from non-SMC, and make these loci permissive for transcriptional activation. In contrast, phenotypic switching of SMC in response to vascular injury, or PDGF BB treatment, was associated with loss of many of these SMC-selective histone modifications, as well as acquisition of histone changes associated with transcriptional silencing/chromatin condensation. However, H3K4 demethylation at SMC marker gene loci, a histone change that appears during development of SMC from ESC, was completely unchanged in all models of SMC phenotypic switching examined, suggesting that it may be relatively "fixed", and serve to preserve SMC "lineage memory" during reversible phenotypic switching. Taken together, results indicate that there is a distinct pattern of histone modifications that distinguishes SMC from non-SMC, and that these SMC- and gene locus-specific epigenetic modifications are likely to play a key role in regulating SMC differentiation marker gene expression in development and disease. The focus of this project is to test the hypothesis that development of SMC from embryonic stem cells is associated with acquisition of a unique pattern of histone modifications at SMC marker (and regulatory) gene loci and that these histone modifications play a key role in determining the permissiveness of these gene loci for transcriptional activation as well as in providing "SMC lineage memory" during reversible phenotypic switching. We will address this hypothesis by addressing the following two specific aims. Aim 1 is to determine mechanisms by which SMC selective/specific chromatin modifications regulate expression of SMC differentiation marker and regulatory genes during development of SMC lineages from multipotential stem cells. This will include the first studies to directly test the role of specific histone modifications in control of SMC lineage/differentiation, and how these histone modifications are acquired during development. Aim 2 is to define the role of epigenetic modifications in mediating reversible phenotypic switching of vascular SMC in response to vascular injury in vivo or treatment of cultured SMC with PDGF BB. Studies will define fundamental mechanisms that control differentiation of SMC, and are likely to lead to novel therapies for treatment of diseases in which SMC phenotypic switching plays a major role.