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（由申请人提供）：视网膜神经元在视网膜上均匀间隔，这种模式称为马赛克。甚至在开发过程中也会通过接触介导的排斥力在相同类型的神经元之间进行的接触介导的排斥。尚不清楚同型神经元彼此识别的分子机制，因此彼此避免。这里的目的是了解如何启动，接收和转化为调整细胞位置的信号。中心假设是跨膜蛋白MEGF10和MEGF11构成了一种受体配体系统：1）通过结合细胞 - 细胞接触来赋予同型识别； 2）触发产生相互细胞抑制的细胞内信号通路，从而产生镶嵌间距。这项工作的基本原理是，它将通过揭示第一个识别的识别分子集（即MEGF10/11）位置神经元的第一个机械解释。这样揭示的机制有望提供有关视网膜神经元如何识别和避免彼此的一般见解，开辟了理解镶嵌物以及其他影响视觉功能的神经元模式事件的道路。为此，提出了以下特定目标：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通过与自身的结合并充当受体和配体来介导这些相互作用。为了检验该假设，每个分子的结合特异性将通过生化确定，并使用MEGF10和MEGF11突变小鼠在体内确认其受体/配体功能。 2）确定细胞中如何报告识别信号。初步数据表明，MEGF10需要传递识别信号。使用生化和体内遗传实验，此目标将检验以下假设：MEGF10细胞内域中的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镶嵌间距。该目标将检验识别会改变树突行为的假设。具体而言，有人提出识别会导致同型树突抑制作用，神经元通过该抑制作用，使他们能够避开邻居。在MEGF10中，识别将被遗传废除； MEGF11双重突变小鼠和树突状排斥将通过视网膜外植体的实时成像进行评估。总之，这三个目标中提出的实验将首次揭示。1）当相同类型接触的细胞接触时，相互结合的细胞表面分子； 2）这些分子如何触发排斥以指定神经元位置。这种方法具有创新性，因为它采用了新颖的工具和方法来实现镶嵌图案中同型识别的首次分子研究。贡献将是重要的，因为确定神经元精确位置的分子事件对于视网膜和整个神经系统中的电路功能都很重要。