Nuclear FAK-mediated VSMC differentiation via epigenetic reprograming invascular diseases

通过表观遗传重编程血管疾病中核 FAK 介导的 VSMC 分化

• 批准号: 10618482
• 负责人: Steve Lim
• 金额: $ 47.54万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-16 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10618482
• 关键词: Apolipoprotein E; Area; Arterial Fatty Streak; Arteries; Atherosclerosis; Biochemical; CD47 gene; Cardiovascular system; Cell Cycle; Cell Differentiation process; Cell Nucleus; Cell Proliferation; Cells; Complex; Cyclin D1; DNA Methylation; DNA Modification Methylases; Data; Deacetylase; Disease; Epigenetic Process; Failure; Focal Adhesion Kinase 1; Gene Expression; Genes; Genetic; Genetic Transcription; Growth Factor; Histones; Hyperlipidemia; Inflammatory; Injury; Integrins; Intervention; Intervention Studies; Lead; Lesion; Mediating; Modeling; Molecular; Molecular Biology; Mus; Nuclear; Nucleosomes; Pharmacology; Phenotype; Play; Protein Tyrosine Kinase; Proteomics; Repression; Role; Signal Transduction; Smooth Muscle Actin Staining Method; Smooth Muscle Myocytes; Stimulus; Testing; Therapeutic; Thick; Ubiquitination; Vascular Diseases; Vascular Smooth Muscle; cell dedifferentiation; cell motility; cell type; chromatin remodeling; epigenetic regulation; histone modification; insight; kinase inhibitor; macrophage; mouse model; novel; oxidized lipid; prevent; promoter; response; tool; transdifferentiation; vascular injury; vascular smooth muscle cell proliferation; western diet

Project Summary Vascular smooth muscle cells (VSMCs) can dedifferentiate into a highly proliferative state with less contractile gene expression upon vessel injury or transdifferentiate into macrophage-like cells (MLCs) during atherosclerosis progression. These types of phenotypic switching are driven by multiple transcriptional and epigenetic changes. Despite vicious effects of VSMC dedifferentiation and transdifferentiation in vascular diseases, direct interventional studies that target vicious VSMC phenotype switching have been lacking. During VSMC dedifferentiation, increased secretion of matrix and growth factors alter integrin signaling and lead to aberrant focal adhesion kinase (FAK) activation, which promotes VSMC proliferation. We demonstrated that FAK is inactive and primarily localized within the nuclei of VSMCs of healthy arteries. However, vessel injury promoted FAK activation and cytoplasmic relocalization which increased cell cycling. While we also observed that FAK activation suppresses expression of VSMC contractile genes, the underlying mechanism by which FAK regulates VSMC contractile genes is not known. Our preliminary data demonstrated that inhibition of FAK catalytic activity in VSMCs induced nuclear localization of FAK and increased contractile gene transcription. Through biochemical and proteomics studies, we identified two independent epigenetic repression machineries, DNA methyltransferase 3A (DNMT3A) and the nucleosome remodeling and deacetylase (NuRD) complex, as nuclear FAK-interacting partners. Importantly, we found that FAK inhibition decreased DNMT3A and NuRD component expression, which was associated with decreased DNA methylation and increased active histone marks (H3K27ac and H4ac) in the contractile gene promoters. Using genetic FAK cytoplasmic (Cyto) restricted VSMCs, we found that FAK nuclear localization is required for reduction of DNMT3A and NuRD complex. Furthermore, FAK inhibition blocked advanced atherosclerotic lesion formation in ApoE-/- mice with decreased DNMT3A and NuRD component expression. This was associated with a thicker smooth muscle actin positive area, suggesting that FAK inhibition increased VSMC differentiation and plaque stability compared to control. Our results implicated that FAK activation upon injury or hyperlipidemia stimulation may contribute to VSMC phenotype switching potentially via epigenetic regulation. Our hypothesis is that FAK catalytic inhibition forces FAK nuclear localization and promotes VSMC differentiation via reduced expression of DNMT3A and NuRD. In Aim 1, we will elucidate the molecular mechanism of nuclear FAK-mediated VSMC phenotypic switching via epigenetic modulation of DNA methylation, and histone modification. In Aim 2, we will investigate the role of DNMT3A and the NuRD complex in VSMC dedifferentiation upon vascular injury. In Aim 3, we will evaluate the effect of FAK inhibition on blocking VSMC transdifferentiation and promoting plaque stability in early and advanced atherosclerosis. This study will provide new insights into VSMC phenotype switching via FAK-mediated epigenetic control through DNMT3A and NuRD complex stability.

项目摘要 血管平滑肌细胞（VSMC）可以将其推导成高度增殖的状态，而收缩较少 血管损伤或转分解后，基因表达在巨噬细胞样细胞（MLC）中 动脉粥样硬化进展。这些类型的表型切换是由多个转录驱动的， 表观遗传变化。尽管VSMC的降解和跨不同的血管造成了恶性影响 缺乏针对恶性VSMC表型切换的疾病，直接介入的研究。期间 VSMC去分化，基质的分泌增加和生长因子改变整合素信号传导，并导致 异常的局灶性粘附激酶（FAK）激活，促进VSMC增殖。我们证明了这一点 FAK是不活跃的，主要定位于健康动脉VSMC的核中。但是，船只受伤 促进了FAK激活和细胞质重定位，从而增加了细胞循环。虽然我们也观察到 FAK激活抑制了VSMC收缩基因的表达，这是一种基本机制 FAK调节VSMC收缩基因尚不清楚。我们的初步数据表明，抑制FAK VSMC中的催化活性诱导FAK的核定位和收缩基因转录增加。 通过生化和蛋白质组学研究，我们确定了两个独立的表观抑制 机械，DNA甲基转移酶3A（DNMT3A）和核小体重塑和脱乙酰基酶（NURD） 复杂，作为核FAK交互伙伴。重要的是，我们发现FAK抑制减少了DNMT3A 和NURD成分表达，与DNA甲基化降低相关并增加 收缩基因启动子中的活性组蛋白标记（H3K27AC和H4AC）。使用遗传FAK细胞质 （CYTO）受限制的VSMC，我们发现减少DNMT3A和 努尔德综合体。此外，FAK抑制阻塞了ApoE - / - 小鼠的高级动脉粥样硬化病变形成 DNMT3A和NURD组分表达降低。这与较厚的平滑肌有关 肌动蛋白阳性面积，表明FAK抑制增加了VSMC分化和斑块稳定性 与对照相比。我们的结果暗示FAK激活受伤或高脂血症刺激可能 通过表观遗传调节有可能导致VSMC表型切换。我们的假设是Fak 催化抑制力FAK核定位并通过降低表达促进VSMC分化 DNMT3A和NURD。在AIM 1中，我们将阐明核FAK介导的VSMC的分子机制 通过表观遗传学调节DNA甲基化和组蛋白修饰，表型转换。在AIM 2中，我们将 研究DNMT3A和NURD复合物在血管损伤后VSMC去分化中的作用。目标 3，我们将评估FAK抑制对阻断VSMC转变和促进斑块的影响 早期和晚期动脉粥样硬化的稳定性。这项研究将为VSMC表型提供新的见解 通过FAK介导的表观遗传控制通过DNMT3A和NURD复杂稳定性进行切换。