Role of Fat Cadherins in Neural Development of the Vertebrate Retina

描述（由申请人提供）：在神经回路的组装过程中，新生的神经元迁移到特定的位置，并以刻板的方式扩展过程，以联系适当的突触伙伴。这在脊椎动物视网膜中最为明显，视网膜由两个中间的突触丛状层分隔成三个核层。尽管这种组织的基本性质是知之甚少的定位神经元细胞类型和直接树突延伸的细胞机制。本研究探讨了非典型钙粘蛋白Fat3的功能，它在无毛细胞发育过程中协调这些事件。Fat3是一种异常大的钙粘蛋白分子，质量约500Kd，包含34个细胞外钙粘蛋白结构域，存在于发育中的内丛状层（IPL）中。基因敲除小鼠的分析表明，脂肪3对于指导无毛细胞树突向IPL的极化延伸以及在内核层（INL）和神经节细胞层（GCL）之间合理分布无毛细胞是必要的。在缺乏脂肪3的情况下，异位的无突细胞树突在IPL外精心制作，导致形成两个额外的突触层。在这个项目中，Fat3功能的分子和细胞机制将使用新型的Fat3敲除小鼠和条件敲除小鼠来确定。具体来说，转基因报告基因将用于鉴定表达Fat3的无毛细胞和神经节细胞的形态类别，并区分无毛细胞发育的一般功能与特定神经元回路组装的功能。另外的体外实验将剖析控制Fat3信号的调控机制，包括选择性剪接和与不同细胞质蛋白的动态相互作用。最后，将分析神经节细胞的发育，以确定Fat3是否也是这种细胞类型的形态发生和分布所必需的，并将检查中心突起，以确定Fat3是否对轴突发育是必要的。虽然重点关注的是视网膜中脂肪3的功能，但这项工作将为脂肪粘蛋白在神经发育过程中的作用以及在内耳和肾脏等其他高表达系统中的作用提供重要的见解。

DESCRIPTION (provided by applicant): During the assembly of neural circuits, newly born neurons migrate to specific locations and extend processes in stereotyped manners to contact the appropriate synaptic partners. This is most evident in the vertebrate retina which is organized into three nuclear layers separated by two intervening synaptic plexiform layers. Despite the fundamental nature of this organization little is known about the cellular mechanisms that position neuronal cell types and direct dendrite extension. This study examines the function of the atypical cadherin protein Fat3 which coordinates these events during amacrine cell development. Fat3 is an unusually large cadherin molecule with a mass of ~500Kd that contains 34 extracellular cadherin domains and is present throughout the developing inner plexiform layer (IPL). Analysis of knockout mice reveals that fat3 is necessary for directing the polarized extension of amacrine cell dendrites into the IPL as well as properly distributing amacrine cells between the inner nuclear layer (INL) and the ganglion cell layer (GCL). In the absence of fat3 ectopic amacrine cell dendrites elaborate outside of the IPL resulting in the formation of two additional synaptic layers. In this project the molecular and cellular mechanism(s) of Fat3 function will be determined using novel lines of fat3 knockout and conditional knockout mice. Specifically transgenic reporters will be used to identify morphological classes of amacrine and ganglion cells expressing Fat3 and distinguish between a general function for amacrine cell development versus a function in the assembly of specific neuronal circuits. Additional in vitro experiments will dissect the regulatory mechanisms that control Fat3 signaling including alternative splicing and dynamic interactions with different cytoplasmic proteins. Finally ganglion cell development will be analyzed to determine if Fat3 is also required for the morphogenesis and distribution of this cell type, and central projections will be examined to determine if Fat3 is necessary for axonal development. Although focused on the function of Fat3 in the retina, the proposed work will nevertheless provide important insight into the role of Fat cadherins during neurodevelopment and in other systems where they are highly expressed such as the inner ear and kidney. PUBLIC HEALTH RELEVANCE: In the vertebrate retina the organization of neurons and synaptic connections into circuits connecting the light detecting photoreceptors with retinal ganglion cells that project to the brain is essential for the formation of visual images. This project is designed to determine how retinal neurons are positioned in the retina and extend dendrites to contact the correct synaptic partners. By understanding the developmental events that build retinal circuitry we can predict how these circuits are altered in disease states, identify targets for pharmacological intervention, and contribute to our understanding of the biological basis of vision.