A forward genetics approach to identify novel core regulators of epithelial patte

一种识别上皮细胞新型核心调节因子的正向遗传学方法

基本信息

批准号：
8189594
负责人：
MARK q MARTINDALE
金额：
$ 20.51万
依托单位：
UNIVERSITY OF HAWAII AT MANOA
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-07-01 至 2013-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8189594
关键词：
Affect Alleles Anemone Animals Apical Automobile Driving Biological Biological Models Biological Process Candidate Disease Gene Cell Polarity Cell Proliferation Cells Chromosome Mapping Cleft Palate Cnidaria Congenital Abnormality Crystalline Lens DNA Data Defect Detection Development Embryo Employee Strikes Epithelial Ethylnitrosourea Event Future Gastrointestinal tract structure Generations Genes Genetic Genetic Polymorphism Genetic Screening Genome Genomics Goals Growth Health Heart Human Human Development Induced Mutation Introns Invertebrates Investigation Lead Lesion Limb Bud Limb structure Link Lung Maps Marine Invertebrates Modeling Molecular Molecular Target Morphogenesis Morphology Mutagens Mutation Natural regeneration Neural tube Oral cavity Organ Pancreas Pathway interactions Pattern Phase Phenotype Positioning Attribute Primordium Process Protocols documentation Regenerative Medicine Research Research Personnel Resolution Scheme Screening procedure Sea Anemones Shapes Spinal Dysraphism Structure Synteny System Technology Time Tissues Vertebrates Work base cell motility constriction cost effective feeding genetic pedigree improved insight interest model development molecular marker mutant next generation novel positional cloning regenerative tool vertebrate genome

项目摘要

DESCRIPTION (provided by applicant): This proposal aims to develop a forward genetic approach to identify core genetic components that regulate epithelial patterning and morphogenesis in the novel invertebrate cnidarian model system Nematostella vectensis. Epithelial morphogenesis is a critical component that underlies formation and proper development of structures ranging from the neural tube to internal organs. Invertebrate model systems are ideal tools for cost- effective forward approaches to identify genes that regulate similar biological processes to those in vertebrates. Nematostella is a unique system that has simple epithelial body plan, a genome more similar to vertebrates in gene content and organization than current invertebrate genetic systems, and is highly regenerative. During development of Nematostella, tentacle formation occurs around the mouth of the animal and is remarkably similar to the patterning and morphological changes observed in development of epithelial structures, such as the neural tube, limb buds, and organs, in vertebrate embryos. Though reverse genetic approaches are now well established allowing for investigations of candidate genes during Nematostella development, a forward non-biased approach to identify novel genes and pathways for universal biological processes is lacking. Development of forward approaches to study tentacle formation in this novel system will transform and enhance our understanding of epithelial patterning and morphogenesis. We have developed a protocol to induce mutations via treatment with the common mutagen ENU and describe a standard three generation crossing scheme to screen for recessive mutations that affect tentacle formation. We have generated a preliminary physical map, which we propose to expand in order to perform bulk segregant analysis to map mutant genomic regions linked to tentacle development phenotypes. Next-generation sequencing technology will be used to sequence the genomic region of interest in pooled mutant animals. The sequence data will be compared to the parental sequences and the sequenced Nematostella genome to identify the mutation(s) that lead to the observed tentacle phenotype. Mutations identified in this study will be used for future characterization and investigations focused on understanding the core components that control the initiation and execution of epithelial patterning and morphogenesis during animal development and regeneration. Upon completion of this study, we will have accomplished two goals that will promote our ability to investigate human health and development in invertebrate model systems. (1) We will have identified novel and core regulators of epithelial patterning and morphogenesis; and (2) We will have carried out the first genetic screen in a non-bilaterian marine invertebrate that has striking molecular similarity to vertebrates, thus providing a blueprint for future researchers to exploit this novel model system to improve our understanding of animal development as it relates to human health and regenerative medicine. PUBLIC HEALTH RELEVANCE: This work is relevant to human health because it identifies core genes for future study that regulate patterning, growth, and morphological (shape) changes in sheets of cells called epithelial tissue. Morphological changes and controlled growth of epithelial tissue occurs in the development of nearly all body structures (neural tube, limbs and organs etc.) and defects in this process often lead to birth defects such as spina bifida or cleft palate. Identifying and studying genes that control epithelial tissue patterning, growth, and morphology will enhance our ability to develop detection and treatments for a wide range of birth defects.

描述（由申请人提供）：该提案旨在开发一种前向遗传学方法，以识别新型无脊椎动物Cnidarian模型系统Nematostella vectensis中调节上皮模式和形态发生的核心遗传成分。上皮形态发生是一个关键组成部分，其基础是从神经管到内部器官的结构的形成和正确发展。无脊椎动物模型系统是具有具有成本效益的远期方法的理想工具，可以识别调节与脊椎动物中类似生物学过程的基因。 Nematostella是一个独特的系统，具有简单的上皮身体计划，一种基因组，与当前的无脊椎动物遗传系统相比，基因组和组织中更类似于脊椎动物，并且具有高度再生。在黑头杆菌的发育过程中，触手的形成发生在动物的口周围，并且与脊椎动物胚胎中上皮结构（例如神经管，肢体芽和器官）中观察到的图案和形态学变化非常相似。尽管现在已经建立了反向遗传方法，允许在nematostella发育过程中研究候选基因，但缺乏一种鉴定通用生物学过程的新型基因和途径的前瞻性非偏差方法。在这种新型系统中研究触手形成的正向方法的发展将改变和增强我们对上皮形式和形态发生的理解。我们已经开发了一种方案，可以通过使用共同诱变剂ENU处理诱导突变，并描述了标准的三代交叉方案，以筛选影响触手形成的隐性突变。我们已经生成了一个初步的物理图，我们建议它扩展以进行大量分析分析以绘制与触手发育表型相关的突变基因组区域。下一代测序技术将用于对合并突变动物感兴趣的基因组区域进行测序。将序列数据与父母序列和测序的Nematostella基因组进行比较，以鉴定导致观察到的触手表型的突变。本研究中确定的突变将用于未来的表征和研究，重点是理解控制动物发育和再生期间上皮形式和形态发生的启动和执行的核心成分。这项研究完成后，我们将实现两个目标，这些目标将促进我们调查无脊椎动物模型系统中人类健康和发展的能力。（1）我们将确定上皮模式和形态发生的新颖和核心调节剂；（2）我们将在非二脑海洋无脊椎动物中进行第一个遗传筛选，该遗传筛查与脊椎动物具有惊人的分子相似性，从而为未来的研究人员提供了蓝图，以利用这种新颖的模型系统来利用这一新型模型，以提高我们对动物发展的理解，因为它与人类健康和再生医学有关。公共卫生相关性：这项工作与人类健康相关，因为它确定了未来研究的核心基因，该研究调节了称为上皮组织的细胞板的模式，生长和形态（形状）变化。上皮组织的形态变化和控制的生长发生在几乎所有人体结构（神经管，四肢和器官等）的发展中，并且在此过程中的缺陷通常会导致诸如脊柱裂或left裂等出生缺陷。识别和研究控制上皮组织模式，生长和形态的基因将增强我们为广泛的出生缺陷开发检测和治疗的能力。