Retinoid-Related Genes in Diaphragm and Cardiac Development

膈肌和心脏发育中的类维生素A相关基因

• 批准号: 8440700
• 负责人: Daryl Armstrong Scott
• 金额: $ 29.02万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-03-15 至 2015-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8440700
• 关键词: 8p23.1; Anterior; Binding; Binding Sites; Biological; Biological Assay; Cardiac; Cardiovascular system; Cells; Chromosomes; Congenital Abnormality; Congenital Heart Defects; Congenital diaphragmatic hernia; Defect; Development; Diagnosis; Embryo; Genes; Genetic; Genotype; Goals; Heart; Heterozygote; Human; Human Resources; In Vitro; Incidence; Individual; Knowledge; Lead; Life; Luciferases; Modeling; Molecular; Mus; Organ; Pathway interactions; Patients; Peritoneal; Phenotype; Play; Pleural; Reporter; Respiratory Diaphragm; Retinoids; Role; Severities; Signal Pathway; System; Techniques; Tretinoin; Up-Regulation; Xenopus; apoAI regulatory protein-1; base; cardiogenesis; chromatin immunoprecipitation; cohort; human data; in vivo; malformation; mouse model; novel; novel therapeutics; prevent; programs; promoter; public health relevance; transcription factor

DESCRIPTION (provided by applicant): Our long-term goal is to understand the molecular basis of congenital diaphragmatic hernia (CDH)-a common life threatening birth defect. However, our ability to identify key molecular interactions involved in CDH development is hindered by a general paucity of known CDH-related genes, an incomplete understanding of the histopathologic changes that underlie CDH development and the lack of a cell based/culture based system in which to study diaphragm development. These limitations could be minimized by studying CDH-related genes that are also involved in the development of related organs, such as the heart, that are more amenable to study. Since up to 40% of individuals with CDH also have cardiovascular malformations (CVMs), it is likely that some key developmental programs/pathways are shared between these organs. In this proposal we will identify these commonalities by studying SOX7 and GATA4-two retinoid-responsive transcription factors located together in the CDH/CVM minimal deleted region on chromosome 8p23.1-using novel mouse models, cell based systems, and unique human resources. Specific aims for this proposal include: 1) identify the histopathologic changes associated with disruption of Sox7 and Gata4 in the diaphragm, 2) define the role of Sox7 in cardiac development, 3) identify the molecular basis of SOX7-dependent up regulation of Gata4 and determine if Sox7 interacts genetically with Gata4 in diaphragm and cardiac development and 4) identify deleterious changes in retinoid-related genes that cause or predispose to the development of CDH. We have shown that a portion of Sox7 mice develop anterior CDH similar to that seen in Gata4 mice. We will first determine the histopathologic changes that predispose to CDH in this model and then determine the role of Sox7 and Gata4 in the posterior diaphragm by conditionally disrupting their expression using an Nkx3-2 Cre. To define Sox7's role in heart development we will alter Sox7 expression in differentiating P19 cells in vitro and analyze the in vivo cardiac phenotype of Sox7-/- embryos and mice in which Sox7 expression has been disrupted in the first and second heart fields using an Nkx2-5 Cre. The molecular changes that underlie these phenotypes will then be identified using standard molecular techniques. We will also determine if direct binding of SOX7 to the Gata4 promoter is responsible for SOX7-dependent up regulation of Gata4. By comparing the incidence and severity of CDH and CVMs in single and double heterozygous mice, we will also determine if Sox7 interacts genetically with Gata4 in vivo. To confirm their individual roles in human CDH, and to discover novel genotype/phenotype correlations, we will identify deleterious changes in SOX7, GATA4 and two other retinoid-related transcription factors, COUP-TFII and FOG2, in a cohort of CDH patients. In the short term, these studies will help us to understand the key molecular and biological pathway regulated by SOX7 and GATA4 in both the diaphragm and heart whose disruption leads to development of CDH and CVMs. This, in turn, may lead to the development of novel therapeutic or preventative strategies for these birth defects.

描述（由申请人提供）：我们的长期目标是了解先天性膈疝（CDH）的分子基础，这是一种常见的危及生命的出生缺陷。然而，我们识别 CDH 发育中涉及的关键分子相互作用的能力受到以下因素的阻碍：已知的 CDH 相关基因普遍缺乏、对 CDH 发育背后的组织病理学变化的不完全理解以及缺乏基于细胞/基于培养的系统研究膈肌的发育。通过研究 CDH 相关基因可以最大限度地减少这些限制，这些基因也参与相关器官的发育，例如更适合研究的心脏。由于高达 40% 的 CDH 患者还患有心血管畸形 (CVM)，因此这些器官之间可能共享一些关键的发育程序/途径。在本提案中，我们将通过研究 SOX7 和 GATA4（位于染色体 8p23.1 上 CDH/CVM 最小缺失区域的两个类维生素A反应转录因子）来确定这些共性，使用新型小鼠模型、基于细胞的系统和独特的人力资源。该提案的具体目标包括：1) 确定与膈肌中 Sox7 和 Gata4 破坏相关的组织病理学变化，2) 确定 Sox7 在心脏发育中的作用，3) 确定 SOX7 依赖性 Gata4 上调和确定 Sox7 在膈肌和心脏发育中是否与 Gata4 存在遗传相互作用，4) 识别类视黄醇相关基因中导致或易导致 CDH 发展的有害变化。我们已经证明，部分 Sox7 小鼠会产生与 Gata4 小鼠相似的前部 CDH。我们将首先确定该模型中易患 CDH 的组织病理学变化，然后通过使用 Nkx3-2 Cre 有条件地破坏 Sox7 和 Gata4 的表达来确定 Sox7 和 Gata4 在后膈肌中的作用。为了确定 Sox7 在心脏发育中的作用，我们将改变体外分化 P19 细胞中的 Sox7 表达，并分析 Sox7-/- 胚胎和小鼠的体内心脏表型，其中使用 Nkx2 在第一和第二心脏区域中 Sox7 表达被破坏。 -5 克里。然后将使用标准分子技术来鉴定这些表型背后的分子变化。我们还将确定 SOX7 与 Gata4 启动子的直接结合是否负责 SOX7 依赖性 Gata4 的上调。通过比较单杂合子和双杂合子小鼠中 CDH 和 CVM 的发生率和严重程度，我们还将确定 Sox7 是否在体内与 Gata4 存在遗传相互作用。为了确认它们在人类 CDH 中的个体作用，并发现新的基因型/表型相关性，我们将在一组 CDH 患者中鉴定 SOX7、GATA4 和其他两种视黄醇相关转录因子 COUP-TFII 和 FOG2 的有害变化。短期内，这些研究将帮助我们了解隔膜和心脏中 SOX7 和 GATA4 调节的关键分子和生物途径，这些途径的破坏会导致 CDH 和 CVM 的发展。反过来，这可能会导致针对这些出生缺陷的新型治疗或预防策略的开发。