Endothelial Lineage Diversity: Role of Epigenetics

该项目的科学目标是进一步了解基于染色质的途径如何促进内皮细胞(EC)基因的表达。我们研究的一个重点是负责血管内皮细胞产生NO的基因，即内皮型一氧化氮合酶(ENOS)。我们已经报道了eNOS基因的启动子在体外和体内的内皮细胞中都发生了低甲基化。相反，启动子在从不表达eNOS的细胞类型中分离出来的基因组DNA中被密集甲基化。我们还报道了包含eNOS核心启动子的核小体在表达细胞类型与非表达细胞类型中高度富含激活组蛋白修饰。该项目的总体假设是，表观遗传过程在EC基因表达的空间和时间调节中发挥主要作用。这个项目是对我们整个PPG主题的极大补充。我们将与PPG的所有成员合作，充分利用PPG核心的优势。在目标I中，我们将确定eNOS近端启动子上的细胞特异性表观遗传标记的维持对于维持内皮细胞中eNOS的结构性转录和在不表达eNOS的分化细胞类型中的转录抑制是否至关重要。在AIM II中，我们将确定表观遗传途径在EC分化和血管发育过程中对eNOS表达的时间调控的贡献。我们将使用体外和体内方法。我们在一些EC特异性基因的启动子上发现了不同的表观遗传修饰模式，这表明我们对eNOS基因的发现可能具有广泛的相关性。因此，在目标III中，我们将使用ChlP-on-Chip方法来定义表观遗传途径对EC特异性基因表达的全球调控的贡献。我们还将确定大的基因间非编码RNA，特别是在内皮细胞中丰富。我们预计我们的研究将为我们提供对 表观遗传途径对EC特异性基因表达的全球模式的贡献

The scientific goal of this project is to gain further insjght into how chromatin-based pathways contribute to endothelial cell (EC) gene expression. A focus of our studies is the gene responsible for the production of NO by vascular endothelium, namely endothelial nitric oxide synthase (eNOS). We have reported that the promoter of the eNOS gene is hypomethylated in endothelial cells both in vitro and in vivo. In contrast, the promoter was densely methylated in genomic DNA isolated from cell types that do not express eNOS. We have also reported that the nucleosomes that encompass the eNOS core promoter were highly enriched in activating histone modifications in expressing versus non-expressing cell types. The overall hypothesis of this project is that epigenetic processes play a major role in the spatial and temporal regulation of EC gene expression. This project is extremely complementary to our overall PPG theme. We will collaborate with all members of this PPG and take full advantage of the PPG Cores. In Aim I we will define whether the maintenance of cell-specific epigenetic marks at the eNOS proximal promoter is critically important for maintaining constitutive transcription of eNOS in ECs and transcriptional repression in differentiated cell types that do not express eNOS. In Aim II we will define the contribution of epigenetic pathways to the temporal regulation of eNOS expression during EC differentiation and vascular development. We will use in vitro and in vivo approaches. We have found differential patterns of epigenetic modifications at the promoters of a number of EC-specific genes suggesting that our findings with the eNOS gene may be broadly relevant. Therefore, in Aim III we will use a ChlP-on-chip approach to define the contribution of epigenetic pathways to the global regulation of EC-specific gene expression. We will also identify large-intergenic non-coding RNAs that especially enriched in ECs. We anticipate that our studies will provide new insight into the contribution of epigenetic pathways to global patterns of EC-specific gene expression