Regulation of Venous and Lymphatic Identity by Chromatin-Remodeling

描述(申请人提供)：表观遗传调节剂在各种发育过程中发挥重要作用，但对它们如何调节血管规格知之甚少。申请人实验室的长期目标是了解依赖于ATP的染色质重塑复合体如何在表观遗传学上调节与发育和出生后血管病理有关的基因转录，例如因错误指定血管和淋巴管而引起的疾病。这项建议的目的是了解染色质重塑酶BRG1和CHD4在胚胎发育过程中如何影响静脉和淋巴的特性。中心假说是BRG1和CHD4通过调节核受体COUP-TFII的血管表达而影响静脉和淋巴的同一性。这一假设是基于申请人实验室产生的初步数据以及血管BRG1、CHD4和COUP-TFII突变小鼠胚胎之间的表型相似性。COUP-TFII在静脉和淋巴指定中发挥重要作用，但目前尚不清楚是什么促进了其血管表达。因此，识别调控其表达的表观遗传学机制将有助于阐明COUP-TFII是如何影响血管发育的。两个特定的目标将被用来检验中心假设。首先，将使用一系列遗传工具来识别BRG1和CHD4对静脉和淋巴特性的时间/空间影响。突变胚胎中的静脉和淋巴标记物将在整个发育过程中进行分析，以准确确定BRG1和CHD4影响这些血管的时间和地点。其次，将检测BRG1和CHD4对静脉和淋巴管COUP-TFII表达的表观遗传学影响。BRG1和CHD4突变体中COUP-TFII的表达将与目标1中定义的血管和淋巴指定表型相关。此外，原代和培养的内皮细胞将被用来验证BRG1和CHD4通过COUP-TFII启动子上的核小体重塑和组蛋白乙酰化直接促进血管COUP-TFII表达的假设。这项拟议的研究具有重要意义，因为它将提供有关染色质重塑酶如何影响COUP-TFII表达的新信息，并将作为开发药物策略的第一步，以操纵COUP-TFII的表达，从而积极或消极地影响血管发育。这项研究具有创新性，因为它强调血管发育的表观遗传调控，这与处理这一主题的传统方法不同。

DESCRIPTION (provided by applicant): Epigenetic regulators play important roles in a variety of developmental processes, but little is understood about how they modulate vascular specification. A long-term goal of the applicant's laboratory is to understand how ATP-dependent chromatin-remodeling complexes epigenetically regulate transcription of genes that con- tribute to developmental and postnatal vascular pathologies, such as those that arise from misspecification of blood and lymphatic vessels. The objective of this proposal is to understand how the chromatin-remodeling enzymes BRG1 and CHD4 influence venous and lymphatic identity during embryonic development. The cen- tral hypothesis is that BRG1 and CHD4 affect venous and lymphatic identity by modulating vascular expression of the nuclear receptor Coup-TFII. This hypothesis is based on preliminary data produced in the applicant's laboratory and on phenotypic similarities between vascular Brg1, Chd4, and Coup-TFII mutant mouse em- bryos. Coup-TFII plays important roles in venous and lymphatic specification but nothing is known about what promotes its vascular expression. Therefore identification of epigenetic mechanisms for regulating its expres- sion would elucidate how COUP-TFII influences vessel development. Two specific aims will be employed to test the central hypothesis. First, the temporal/spatial impact of BRG1 and CHD4 on venous and lymphatic identity will be discerned using an array of genetic tools. Venous and lymphatic markers in mutant embryos will be analyzed throughout development to determine precisely when and where BRG1 and CHD4 affect these vessels. Secondly, the epigenetic impact of BRG1 and CHD4 on Coup-TFII expression in veins and lymphatics will be examined. COUP-TFII expression in Brg1 and Chd4 mutants will be correlated with the ve- nous and lymphatic specification phenotypes defined in Aim 1. In addition, primary and cultured endothelial cells will be used to test the hypothesis that BRG1 and CHD4 directly promote vascular Coup-TFII expression through nucleosome remodeling and histone acetylation at the Coup-TFII promoter. The proposed research is significant because it will provide novel information about how chromatin-remodeling enzymes affect Coup-TFII expression and will serve as a first step toward developing pharmacological strategies to manipulate Coup-TFII expression in order to positively or negatively influence vascular development. This research is innovative be- cause it emphasizes epigenetic regulation of vascular development, which deviates from traditional ways of approaching this subject.