Hand2 Function and Regulation During Craniofacial Development

Hand2 颅面发育过程中的功能和调节

• 批准号: 7731205
• 负责人: David E. Clouthier
• 金额: $ 37.04万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-20 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7731205
• 关键词: Address; BHLH Protein; Biological Models; Branchial arch structure; Cartilage; Cell Death; Cells; Cephalic; Chromosome Mapping; Cleft Palate; Congenital Abnormality; Connective Tissue; Coupling; Data; Defect; Development; Developmental Process; Distal; Down-Regulation; Embryo; Embryonic Development; Endothelin-1; Enhancers; Event; Face; Fibrinogen; First Pharyngeal Arch; Future; Gap Junctions; Gene Expression; Genes; Genetic; Goals; Heart; Human; Human Development; Jaw; Knock-out; Knockout Mice; Knowledge; Lead; Limb Development; Live Birth; Mammals; Mandible; Maps; Micrognathism; Molecular; Molecular Analysis; Morphogenesis; Mouse Strains; Mus; Mutant Strains Mice; Neck; Neural Crest; Neural Crest Cell; Osteogenesis; Pathway interactions; Pattern; Phenotype; Process; Regulation; Role; Signal Pathway; Signal Transduction; Signaling Molecule; Societies; Structure; Syndrome; System; Systems Biology; Testing; Tissue Engineering; Tissues; Transcription Coactivator; Transcription Repressor/Corepressor; Transgenes; Zebrafish; base; bone; craniofacial; daughter cell; developmental genetics; fascinate; gene repression; malformation; middle ear; mutant; novel; prevent; public health relevance; social; transcription factor

DESCRIPTION (provided by applicant): The goal of this project is to define the function of the basic helix-loop-helix transcription factor Hand2 during mammalian lower jaw development. Our hypothesis is that Hand2 contributes to the development of most lower jaw structures by establishing a distal morphogenetic domain in the mandibular arch. Within this boundary, Hand2 confines genes involved in mandibular pharyngeal arch development while also repressing the expression of genes normally observed outside this domain. Further, we hypothesize that Hand2 expression and function is at least partially regulated by Twist1 through both transcriptional repression and genetic interactions. In both mouse and zebrafish, Hand2 is expressed in cranial neural crest cell (NCC)-derived cells within the mandibular pharyngeal arch, from which bone and cartilage of the lower jaw arise. Targeted inactivation of the Hand2 gene in mice results in early embryonic lethality (by E10.5). However, hand2-mutant zebrafish are viable for five days; in these mutants, extensive malformations in craniofacial cartilages are accompanied by positive and negative changes in gene expression. We have shown in term mouse embryos that Hand2-daughter cells compose most structures derived from the mandibular arch. In our Preliminary Data, we show that loss of Hand2 in NCCs resultst in craniofacial defects indicative of NCC mispatterning. In this proposal, we will address the role of Hand2 in facial morphogenesis in three Specific Aims. In Aim 1, we will define the cellular and molecular changes in facial development following conditional inactivation of the Hand2 gene in cranial neural crest cells. In Aim 2, we will use hand2 zebrafish mutants to examine whether Hand2 acts as both a transcriptional activator and repressor and use Hand2 conditional mouse mutants to identify targets of Hand2 action. In Aim 3, we will define the role of Twist1 in the Hand2 mandibular arch domain and determine whether Twist1 and Hand2 interact genetically within this domain. By coupling our cellular and molecular analysis of Hand2 conditional knockout mice with our functional analysis of Hand2 action in mouse and zebrafish, we expect to uncover novel regulatory mechanisms governing fate and identity of NCCs that ultimately lead to facial development and whose disruption can lead to human facial birth defects, including micrognathia. PUBLIC HEALTH RELEVANCE: Craniofacial birth defect syndromes occur in 1 out of every 250 live births and represent a large financial and social burden within our society. While numerous mouse developmental genetics studies have elucidated the basis of some of these syndromes, the cause of many more remains unknown. The phenotype of the hand2 mutant zebrafish indicates that Hand2 may be involved in establishing or maintaining a developmental domain necessary for mandibular arch development, though early lethality prevents analysis of the role of Hand2 in later arch patterning events and bone formation. Our study will directly address the function and regulation of Hand2 during lower jaw development using a combination of model systems. Together, our studies will help define how regional developmental domains necessary for facial development are established and how different domains may antagonize each other to produce a final facial plan. Such knowledge can be subsequently combined using a systems biology approach to build a more comprehensive "gene map" involved in facial formation, which could lead to significant advances in future tissue engineering approaches to treat human craniofacial anomalies.

描述（由申请人提供）：该项目的目标是确定基本螺旋-环-螺旋转录因子Hand2在哺乳动物下颌发育过程中的功能。我们的假设是，Hand2 通过在下颌弓中建立远端形态发生域来促进大多数下颌结构的发育。在这个边界内，Hand2 限制了参与下颌咽弓发育的基因，同时也抑制了通常在该域之外观察到的基因的表达。此外，我们假设 Hand2 的表达和功能至少部分受到 Twist1 通过转录抑制和遗传相互作用的调节。在小鼠和斑马鱼中，Hand2 在下颌咽弓内的颅神经嵴细胞 (NCC) 衍生细胞中表达，下颌骨和软骨均由咽弓产生。小鼠 Hand2 基因的靶向失活会导致早期胚胎死亡（E10.5）。然而，hand2 突变斑马鱼可以存活五天；在这些突变体中，颅面软骨的广泛畸形伴随着基因表达的正向和负向变化。我们在足月小鼠胚胎中证明，Hand2 子细胞构成了下颌弓的大部分结构。在我们的初步数据中，我们表明 NCC 中 Hand2 的缺失会导致颅面缺陷，表明 NCC 模式错误。在本提案中，我们将在三个具体目标中讨论 Hand2 在面部形态发生中的作用。在目标 1 中，我们将定义颅神经嵴细胞中 Hand2 基因条件失活后面部发育的细胞和分子变化。在目标 2 中，我们将使用 hand2 斑马鱼突变体来检查 Hand2 是否同时充当转录激活子和阻遏子，并使用 Hand2 条件小鼠突变体来识别 Hand2 作用的目标。在目标 3 中，我们将定义 Twist1 在 Hand2 下颌弓结构域中的作用，并确定 Twist1 和 Hand2 是否在该结构域内发生遗传相互作用。通过将我们对 Hand2 条件敲除小鼠的细胞和分子分析与我们对小鼠和斑马鱼中 Hand2 作用的功能分析结合起来，我们期望发现控制 NCC 命运和身份的新调控机制，最终导致面部发育，其破坏可能导致人类面部出生缺陷，包括小颌畸形。公共健康相关性：每 250 名活产儿中就有 1 人患有颅面出生缺陷综合症，对我们的社会造成了巨大的经济和社会负担。虽然大量的小鼠发育遗传学研究已经阐明了其中一些综合征的基础，但还有更多综合征的原因仍然未知。 hand2突变斑马鱼的表型表明Hand2可能参与建立或维持下颌牙弓发育所需的发育域，尽管早期致死性阻碍了对Hand2在后来的牙弓图案化事件和骨形成中的作用的分析。我们的研究将使用模型系统的组合直接解决下颌发育过程中 Hand2 的功能和调节。总之，我们的研究将有助于确定如何建立面部发育所需的区域发育域，以及不同域如何相互对抗以产生最终的面部计划。随后可以使用系统生物学方法将这些知识结合起来，构建涉及面部形成的更全面的“基因图谱”，这可能会导致未来治疗人类颅面异常的组织工程方法取得重大进展。