Transcriptional control of epithelial behaviors that drive mammalian neural tubeclosure

驱动哺乳动物神经管闭合的上皮行为的转录控制

• 批准号: 9660106
• 负责人: Lee A. Niswander
• 金额: $ 9.37万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-01 至 2020-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9660106
• 关键词: Transcriptional; control; epithelial; behaviors; drive

 DESCRIPTION (provided by applicant): Failure of neural tube closure is a devastating birth defect. Research in the Niswander lab has provided significant insights into the molecular and cellular regulation of NT closure. We have created and studied mouse models with neural tube defects (NTDs) to elucidate the molecular foundations of NT closure. Moreover, we have created robust and novel technology to visualize NT closure in a living mammalian embryo. Our dynamic imaging and key genetic mutants have focused attention on the little studied but critical role for the non-neural ectoderm (NNE) in NT closure. In addition we developed methods to specifically isolate NNE cells to provide a refined and robust platform to study the biology of the NNE. Here we will build upon our unique perspective and turn our attention to spinal NTDs, to provide insight into the most common type of NTD in humans, and to two genetic pathways that are associated with spina bifida in mice and humans. Aim 1 will test the hypothesis that the two closely related Grainyhead-like (GRHL) transcription factors, GRHL2 and GRHL3, differentially control cranial and spinal NT closure through unique and differential activation of target genes, in part mediated by interaction with the JNK signaling pathway that activates the AP1 (cJUN/cFOS) transcription factor. Aim 2 will extend our live platform to test the hypothesis that GRHL-regulated NNE transcriptional programs drive NT closure by controlling cell adhesion, recycling of membrane components, cell shape changes, and/or actin dynamics. Aim 3 will combine our comprehensive molecular and cellular insights with novel unpublished analyses of hundreds of NTD samples to test the hypothesis that mutations identified in GRHL3 and the JNK pathway are causative for spinal NTDs in humans. Relevance of research to public health: The proposed experiments will lead to new cellular and molecular insights into the causes of caudal NTDs, the most common type of NTD and which leads to a profoundly important and frequently disrupted aspect of mammalian embryogenesis. Moreover, our studies will impart novel insights into the general mechanisms of embryonic tissue fusion including the face and body wall. The insights gained here may lead to therapies of general application for treatment of embryonic tissue closure defects that together represent a significant percentage of human birth defects. Abbreviations used in proposal: CDH1 Cadherin1 or E-cadherin EMT Epithelial-to-mesenchymal transition GRHL Grainyhead-like (GRH is the fly homolog) KD Knock-down mT/mG Membrane tomato/membrane GFP fluorescent reporter, GFP expression is activated by Cre NNE Non-neural or surface ectoderm NT Neural tube NTD Neural tube defect Grhl2-null: We will use Grhl21Nisw allele that we isolated in our ENU-screen and which has the same phenotype as other Grhl2 null alleles. Grhl3-Cre: We will use Grhl3-Cre which generates a null allele (obtained from S. Coughlin; Grhl3tm1(cre)Cgh).

 描述（由申请人提供）： 神经管闭合失败是一种毁灭性的出生缺陷。Niswander实验室的研究为NT闭合的分子和细胞调控提供了重要的见解。我们已经建立并研究了小鼠模型与神经管缺陷（NTDs），以阐明NT关闭的分子基础。此外，我们已经创造了强大的和新颖的技术来可视化NT关闭在活的哺乳动物胚胎。我们的动态成像和关键的遗传突变体的关注点很少研究，但关键作用的非神经外胚层（NNE）在NT关闭。此外，我们开发了特异性分离NNE细胞的方法，以提供一个精细和强大的平台来研究NNE细胞的生物学。 NNE。 在这里，我们将建立在我们独特的视角，并把我们的注意力转向脊柱NTD，提供洞察人类最常见的NTD类型，以及与小鼠和人类脊柱裂相关的两种遗传途径。目的1将测试的假设，即两个密切相关的Grainyhead样（GRHL）转录因子，GRHL 2和GRHL 3，差异控制颅和脊髓NT关闭通过独特的和差异激活的靶基因，部分介导的相互作用与JNK信号通路激活AP 1（cJUN/cFOS）转录因子。目标2将扩展我们的实时平台，以测试这一假设，即GRHL调节的NNE转录程序通过控制细胞粘附，膜成分的再循环，细胞形状变化和/或肌动蛋白动力学来驱动NT关闭。目标3将联合收割机我们全面的分子和细胞的见解与数百个NTD样本的新的未发表的分析，以测试在GRHL 3和JNK通路中鉴定的突变是人类脊髓NTD的病因的假设。研究与公共卫生的相关性：拟议的实验将导致对尾部NTD原因的新的细胞和分子见解，这是最常见的NTD类型，并导致哺乳动物胚胎发育的一个非常重要和经常中断的方面。此外，我们的研究将赋予新的见解胚胎组织融合的一般机制，包括脸和体壁。这里获得的见解可能导致治疗胚胎组织闭合缺陷的一般应用的疗法，这些缺陷共同代表了人类出生缺陷的显著百分比。提案中使用的缩写：CDH 1钙粘蛋白1或E-cadherin EMT上皮-间质转化GRHL粒头样（GRH是果蝇同源物）KD敲低mT/mG膜番茄/膜GFP荧光报告基因，GFP表达被Cre NNE激活非神经或表面外胚层NT神经管NTD神经管缺陷Grhl 2-null：我们将使用我们在ENU筛选中分离的Grhl 21 Nisw等位基因，其具有与其他Grhl 2无效等位基因相同的表型。Grhl 3-Cre：我们将使用产生无效等位基因的Grhl 3-Cre（从S. Coughlin; Grhl3tm1（cre）Cgh）。