Molecular control of neuronal position during retinal development

视网膜发育过程中神经元位置的分子控制

• 批准号: 8765567
• 负责人: Jeremy N Kay
• 金额: $ 39.29万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2019-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8765567
• 关键词: Behavior; Binding; Biochemical; Biological Assay; Cell surface; Cells; Code; Cytoplasmic Tail; Data; Dendrites; Development; Disease; Dissection; Enzyme-Linked Immunosorbent Assay; Event; Gene Delivery; Genetic; Goals; Homo; ITAM; Image; In Situ; Individual; Integral Membrane Protein; Knowledge; Learning; Life; Ligand Binding; Ligands; Location; Mediating; Methods; Mole the mammal; Molecular; Molecular Genetics; Mus; Mutant Strains Mice; Natural regeneration; Nervous system structure; Neurodegenerative Disorders; Neurons; Pattern; Phosphorylation; Phosphotyrosine; Positioning Attribute; Public Health; Recruitment Activity; Reporting; Research; Retina; Retinal; Retinal Diseases; Retinal blind spot; Signal Pathway; Signal Transduction; Specificity; Surface; System; Testing; Time; To specify; Touch sensation; Translating; Vision; Work; base; cell type; design; genetic manipulation; human SYK protein; in utero; in vivo; innovation; insight; neural circuit; neuronal patterning; novel; prevent; public health relevance; receptor; regenerative; regenerative therapy; research study; response; retinal neuron; src-Family Kinases; tool; two-photon; vision science

DESCRIPTION (provided by applicant): Retinal neurons are evenly spaced across the retina, a pattern known as a mosaic. Even spacing arises during development through contact-mediated repulsion that occurs specifically between neurons of the same type. The molecular mechanisms that allow homotypic neurons to recognize each other, and consequently to avoid each other, are not known. The objective here is to learn how homotypic recognition signals are initiated, received, and translated into signals that adjust cell position. The central hypothesis s that the transmembrane proteins MEGF10 and MEGF11 constitute a receptor-ligand system that: 1) confers homotypic recognition through binding upon cell-cell contact; and 2) triggers intracellular signaling pathways that produce mutual cell-cell repulsion, thereby creating mosaic spacing. The rationale for this work is that it will provide the first mechanistic explanation of mosaic formation, by revealing how the first identified set of recognition molecules (i.e. MEGF10/11) positions neurons. The mechanisms thus revealed are expected to provide general insight into how retinal neurons recognize and avoid each other, opening the way to understanding both mosaics as well as other neuronal patterning events that influence visual function. To this end, the following Specific Aims are proposed: 1) Determine the intercellular molecular interactions that initiate recognition signals. Preliminary data suggest that MEGF10 and 11 mediate these interactions by binding to themselves and acting as both receptors and ligands. To test this hypothesis the binding specificity of each molecule will be determined biochemically, and their receptor/ligand function will be confirmed in vivo using Megf10 and Megf11 mutant mice. 2) Determine how recognition signals are reported in the cell. Preliminary data show that MEGF10 is required to transduce recognition signals. Using biochemical and in vivo genetic experiments, this Aim will test the hypothesis that ITAM phosphotyrosine motifs in the MEGF10 intracellular domain mediate these recognition signals. 3) Determine how recognition signals alter cellular behavior to produce mosaic spacing. This aim will test the hypothesis that recognition alters the behavior of dendrites. Specifically, it is proposed that recognition causes homotypic dendritic repulsion, through which neurons stake out unique territories that allow them to avoid their neighbors. Recognition will be abrogated genetically in Megf10; Megf11 double mutant mice and dendritic repulsion will be assessed by live imaging of retinal explants. Together, the experiments proposed in these three Aims are expected to reveal for the first time 1) the cell-surface molecules that bind to each other when cells of the same type touch; and 2) how these molecules trigger repulsion in order to specify neuronal position. The approach is innovative because it deploys novel tools and methods to enable the first molecular studies of homotypic recognition in mosaic patterning. The contribution will be significant because molecular events that determine the precise locations of neurons are important for circuit function, both in the retina and throughout the nervous system.

描述(申请人提供)：视网膜神经元在整个视网膜上均匀分布，这种图案被称为马赛克。在发育过程中，通过接触调节的排斥力产生均匀的间隔，这种排斥力特别发生在相同类型的神经元之间。使同型神经元相互识别并因此避开彼此的分子机制尚不清楚。这里的目标是了解同型识别信号是如何启动、接收和转换成调整细胞位置的信号的。S的中心假说认为，跨膜蛋白MEGF10和MEGF11构成了一个受体-配体系统，该系统：1)通过细胞-细胞接触结合实现同型识别；2)触发细胞内信号通路，产生细胞-细胞相互排斥，从而产生镶嵌间距。这项工作的基本原理是，它将通过揭示第一组识别分子(即MEGF10/11)如何定位神经元，提供第一个马赛克形成的机制解释。由此揭示的机制有望提供对视网膜神经元如何识别和避免彼此的总体洞察，为理解马赛克以及其他影响视觉功能的神经元模式事件开辟了道路。为此，提出了以下具体目标：1)确定启动识别信号的细胞间分子相互作用。初步数据表明，MEGF10和11通过与自身结合并同时作为受体和配体来调节这些相互作用。为了验证这一假设，将通过生化方法确定每个分子的结合特异性，并使用MEGF10和Megf11突变小鼠在体内证实它们的受体/配体功能。2)确定识别信号在单元中的报告方式。初步数据显示，需要MEGF10来转换识别信号。利用生化和体内遗传学实验，这一目标将检验这一假设，即MEGF10细胞内结构域中的ITAM磷酸酪氨酸基序介导了这些识别信号。3)确定识别信号如何改变细胞行为以产生马赛克间距。这一目标将检验识别改变树突行为的假设。具体地说，有人提出，识别会引起同型树突排斥，通过这种排斥，神经元可以划定独特的区域，使它们能够避开邻居。在MEGF10中，识别将在基因上被取消；Megf11双突变小鼠和树突排斥将通过视网膜外植体的活体成像进行评估。总之，这三个目标中提出的实验有望首次揭示：1)当相同类型的细胞接触时，细胞表面分子相互结合；2)这些分子如何触发斥力，以确定神经元的位置。这种方法是创新的，因为它部署了新的工具和方法，使马赛克图案中同型识别的第一次分子研究成为可能。这一贡献将是重大的，因为决定神经元准确位置的分子事件对视网膜和整个神经系统的电路功能都很重要。