Mechanisms of photoreceptor specification and morphogenesis

光感受器规格和形态发生的机制

• 批准号: 10436881
• 负责人: James M Fadool
• 金额: $ 31.6万
• 依托单位: FLORIDA STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10436881
• 关键词: Alleles; Anatomy; Awareness; Binding; Biological Models; CRISPR/Cas technology; Cell Count; Cells; Cessation of life; Chromatin; Communities; Cone; Data; Dependence; Development; Dorsal; Embryo; Enhanced-S cone syndrome; Functional disorder; Gene Expression; Gene Transfer Techniques; Generations; Genes; Genetic; Genetic Models; Genomics; Goals; Human; In Vitro; Knowledge; Laboratories; Light; Mediating; Methods; Mitosis; Molecular; Molecular Analysis; Morphogenesis; Mosaicism; Mus; Mutate; Mutation; Neural Retina; Neurons; Pathway interactions; Pattern; Phenotype; Photoreceptors; Protocols documentation; Publishing; Regulatory Element; Reporting; Research Personnel; Resources; Retina; Retinal Cone; Rod; Role; Series; Signal Transduction; System; Testing; Transgenic Organisms; Vertebrates; Vision; Visual impairment; Work; Zebrafish; base; density; design; experimental study; falls; fovea centralis; gene regulatory network; genetic analysis; genetic manipulation; genome editing; in vivo; innovation; insight; interest; macula; member; neuroblast; novel; photoreceptor progenitor; progenitor; protein expression; protein protein interaction; relating to nervous system; retinal rods; tool; transcription factor; vision development

PROJECT SUMMARY Despite our dependency upon cone-mediated vision, and the debilitating impact of cone degeneration upon our central vision, major gaps exist in our knowledge of genes and mechanisms that regulate the number and spatial patterning of photoreceptors. Underlying this asymmetry is the fundamental choice of retinal neuroblasts to re-enter mitosis or differentiate. The larval zebrafish retina provides an unparalleled genetic model to identify fundamental mechanisms integrating photoreceptor specification and spatial patterning. The larval zebrafish retina is anatomically and functionally cone-dominated with 4 cone subtypes arranged in a highly ordered mosaic. Rods are far less numerous and distributed asymmetrically along the dorsal/ventral axis. The range of genetic tools, availability of genomic resources and access to the zebrafish embryo, permit a systematic approach to uncover the impact of genetic manipulations upon cell fate and their distribution. Our premise is based upon rigorous genetic and molecular analysis in our published studies and preliminary data showing that tbx2b/lor and six7/ljr are essential for maintaining the cone-dominated zebrafish retina. Mutations of tbx2b result in a 5-fold increase and uniform distribution of rods due to a cell fate switch of SWS1- cones into rods, a phenotype opposite to that of the rd7 mouse and enhanced-S cone syndrome in humans. Mutations of six7, an orthologue of Six3/6 result in a similar increase and uniform distribution of rods, however characterization of several alleles revealed that six7 independently suppresses mitosis of late stage progenitors, and is essential for survival of a cone subtype. Our goal is to understand the mechanisms underlying functions of tbx2b and six7 in cell fate decisions and spatial patterning in the context of know factors that regulate photoreceptor development. Aim 1 will take advantage of innovative genome editing tools, transposon-based transgenesis, and more routine methods to test the hypothesis that six7 modulates the choice of late stage photoreceptor progenitors between continued mitosis or differentiation. Aim 2 combined in vivo and in vitro approaches to test our novel hypothesis that tbx2b regulates the timing of photoreceptor determination and thereby spatial patterning through molecular interaction with known photoreceptor transcription factors. Furthermore, our unpublished data and published reports show unexpected alterations in gene expression following genetic alterations suggesting unrecognized roles for these factors in maintaining cell fate. Therefore, completion of the specific aims will significantly advance the fields knowledge of mechanisms regulating the generation of a highly specialized, cone-dominated retina.

项目摘要 尽管我们依赖于视锥细胞介导的视觉， 在我们的中心愿景退化，主要差距存在于我们的基因知识， 调节光感受器的数量和空间模式的机制。其基础 不对称性是视网膜成神经细胞重新进入有丝分裂或分化的基本选择。 斑马鱼的视网膜提供了一个无与伦比的遗传模型，以确定基本的 整合感光细胞特化和空间图案化的机制。斑马鱼的幼体 视网膜在解剖学和功能上以视锥为主，有4种视锥亚型排列在一个 高度有序的马赛克棒的数量要少得多，并且沿着沿着 背/腹轴遗传工具的范围、基因组资源的可得性和获得 斑马鱼胚胎，允许系统的方法来揭示遗传的影响， 操纵细胞命运及其分布。我们的前提是基于严格的遗传学 我们已发表的研究和初步数据中的分子分析表明，TBX 2B/LOR和 Six 7/LJR对于维持视锥细胞占优势的斑马鱼视网膜是必需的。TBX 2B突变 由于SWS 1的细胞命运转换，导致5倍的增加和杆的均匀分布。 视锥细胞转化为视杆细胞，一种与rd 7小鼠相反的表型和增强型S视锥细胞综合征 在人类身上。Six 3/6的直向同源物six 7的突变导致类似的增加和均匀的 分布的杆，然而，表征的几个等位基因显示，six 7独立 抑制晚期祖细胞的有丝分裂，并且是锥细胞亚型存活所必需的。我们 目的是了解tbx 2b和six 7在细胞命运中的作用机制 在已知调节光感受器的因素的背景下， 发展Aim 1将利用创新的基因组编辑工具， 转基因，和更常规的方法来测试的假设，six 7调节的选择， 晚期感光细胞祖细胞之间的持续有丝分裂或分化。目的2 结合体内和体外方法来测试我们的新假设，即TBX 2B调节 光感受器决定的定时以及由此通过分子的空间图案化 与已知的光感受器转录因子的相互作用。此外，我们未公布的数据 和发表的报告显示，基因表达意外的变化， 这些变化表明这些因子在维持细胞命运中的作用尚未被认识。因此，我们认为， 具体目标的完成将大大增进实地对机制的了解 调节高度特化的视锥细胞主导的视网膜的生成。