Transcriptional regulation of mesoderm development

中胚层发育的转录调控

• 批准号: 7303726
• 负责人: ANN K CORSI
• 金额: $ 20.21万
• 依托单位: CATHOLIC UNIVERSITY OF AMERICA
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7303726
• 关键词: Alleles; Animal Model; Animals; Antithymoglobulin; BHLH Protein; Binding; Binding Sites; Biological Models; Boxing; Caenorhabditis elegans; Characteristics; Chotzen Syndrome; Code; Data; Defect; Deletion Mutation; Development; Digit structure; Disease; Drosophila held out wings protein; E protein; Education; Elements; Embryo; Embryonic Development; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic; Genetic Models; Germ Layers; Goals; Helix-Turn-Helix Motifs; Homologous Gene; Human; Individual; Introns; K22 Award; Learning; Limb structure; Link; Location; Mesoderm; Mesoderm Cell; Modeling; Molecular; Mutate; Mutation; Nematoda; Nucleotides; Organism; Pattern; Phenotype; Play; Promoter Regions; Protein Binding; Proteins; Regulation; Reporter; Reporter Genes; Role; Site-Directed Mutagenesis; Students; Syndrome; System; Testing; Tissue-Specific Gene Expression; Tissues; Transcriptional Regulation; Work; cell type; craniofacial; design; dimer; human disease; in vivo; insight; mutant; null mutation; preference; promoter; protein function; research study; tool; transcription factor; twist protein

DESCRIPTION (provided by applicant): Transcription factors play an important role in tissue-specific gene regulation during development. Twist is a basic helix-loop-helix (bHLH) transcription factor that functions in the mesoderm, or middle embryonic germ layer. Humans with Twist mutations have craniofacial and digit defects characteristic of Saethre-Chotzen syndrome. Caenorhabditis elegans is an excellent model organism for elucidating Twist function due to its simplicity, availability of molecular tools, and powerful genetics. The information learned in C. elegans about CeTwist function will be relevant to humans and human disease because CeTwist shares homology with human Twist and multiple CeTwist target genes identified to date are homologous to genes that also cause craniofacial disorders when mutated in humans. bHLH proteins function as dimers. The only dimer partner identified for CeTwist is CeE/DA, which is the homolog of vertebrate E proteins and Drosophila Daughterless (DA). The work in this proposal is designed to uncover the unique functions of CeTwist homodimers and CeTwist/CeE/DA heterodimers in the mesoderm. In the first specific aim, we will use gfp reporters to examine the mesoderm development and CeTwist target gene expression both in CeE/DA mutant animals as well as in animals expressing physically-linked CeTwist dimers that are predicted to behave as forced homodimers in vivo. These experiments are expected to reveal the role that CeE/DA and CeTwist/CeE/DA heterodimers play in the mesoderm as well as to confirm a predicted role for CeTwist homodimers. In the second specific aim, we will examine a promoter paradigm that we recently developed for understanding elements that regulate tissue-specific expression of a model CeTwist target gene, arg-1. bHLH proteins bind to E boxes that are defined as CANNTG, with N being any nucleotide. The minimal arg-1 promoter has 3 E boxes that we have shown by site-directed mutagenesis are uniquely required for gene expression in multiple tissues. The contribution of each E box as well as the intervening sequences between the E boxes will be examined using gfp reporters. Then, the E boxes will be tested in vivo for expression in the presence of excess CeTwist or CeTwist and CeE/DA to determine whether homodimers or heterodimers are working through the individual E boxes. Because the projects described here are suitable for students, this work will also have a profound impact on the education of undergraduate and graduate students. The goal of this work is to examine CeTwist function and target gene regulation. Due to the relatedness between the human and C. elegans Twist proteins, information we learn in our simple model system will be relevant to understanding human Twist and provide insight into diseases caused by defective Twist. The goal of this project is to provide insight into the function of proteins that are defective in human syndromes characterized by craniofacial and limb defects. We have developed a system for examining how these proteins are regulating gene expression in the roundworm, which is a simple model genetic organism. We expect because roundworm and human proteins are related, the information we learn will be relevant to humans and human disease.

描述（由申请人提供）：转录因子在发育期间的组织特异性基因调控中发挥重要作用。Twist是一种碱性螺旋-环-螺旋（bHLH）转录因子，在中胚层或中胚层中发挥作用。具有Twist突变的人具有Saethre-Chotzen综合征特征的颅面和手指缺陷。秀丽隐杆线虫是一个很好的模式生物，阐明扭曲功能，由于其简单，分子工具的可用性，和强大的遗传学。在C.关于CeTwist功能的研究将与人类和人类疾病相关，因为CeTwist与人类Twist具有同源性，并且迄今为止鉴定的多个CeTwist靶基因与在人类中突变时也引起颅面病症的基因同源。bHLH蛋白作为二聚体发挥功能。CeTwist的唯一二聚体伴侣是CeE/DA，它是脊椎动物E蛋白和果蝇无女儿（Drosophila Daughterless，DA）的同系物。这项工作旨在揭示CeTwist同源二聚体和CeTwist/CeE/DA异源二聚体在中胚层中的独特功能。在第一个具体目标中，我们将使用gfp报告基因来检查CeE/DA突变动物以及表达物理连接的CeTwist二聚体的动物中的中胚层发育和CeTwist靶基因表达，所述CeTwist二聚体被预测在体内表现为强制同源二聚体。这些实验预期揭示CeE/DA和CeTwist/CeE/DA异二聚体在中胚层中发挥的作用，以及确认CeTwist同二聚体的预测作用。在第二个具体的目标，我们将研究启动子范例，我们最近开发的理解元素，调节组织特异性表达的模型CeTwist靶基因，arg-1。bHLH蛋白与定义为CANNTG的E盒结合，其中N是任何核苷酸。最小的arg-1启动子有3个E盒，我们已经通过定点诱变显示，这是在多种组织中基因表达所必需的。将使用gfp报告基因检查每个E盒以及E盒之间的间插序列的贡献。然后，在过量CeTwist或CeTwist和CeE/DA存在下，体内测试E盒的表达，以确定是否同源二聚体或异源二聚体通过单个E盒起作用。由于这里描述的项目适合学生，这项工作也将对本科生和研究生的教育产生深远的影响。这项工作的目标是检查CeTwist功能和靶基因调控。由于人类与C.因此，我们在简单的模型系统中了解到的信息将与理解人类Twist相关，并提供对缺陷Twist引起的疾病的深入了解。该项目的目标是深入了解以颅面和肢体缺陷为特征的人类综合征中有缺陷的蛋白质的功能。我们已经开发了一个系统，用于检查这些蛋白质如何调节蛔虫中的基因表达，蛔虫是一种简单的模式遗传生物。我们期望，因为蛔虫和人类蛋白质是相关的，我们了解的信息将与人类和人类疾病有关。