Cell surface molecules that require arrangement of retinal neurons and arbors

描述（由申请人提供）：多种亚型神经元之间的有序且特定的连接是神经回路功能的基础。我们自己的工作使用视网膜作为模型来阐明特异性背后的分子和机制，重点关注突触前和突触后伙伴（特别是突触层）的匹配。然而，还有另一种顺序较少受到关注：细胞及其神经突在正交 (x-y) 平面上的排列。促成这种排列的过程包括神经元胞体的马赛克间距、树突的平铺以及单个乔木内过程的自我回避。这些过程被认为可以确保视野的均匀覆盖、精确的连接和适当的感受野大小。它们在脊椎动物中的分子基础仍然未知。 最近，我们开始分析两组我们怀疑与层流特异性有关的细胞表面蛋白：MEGF10 和 11，以及一组 22 个相关的 γ 原钙粘蛋白 (Pcdhgs)。出乎意料的是，初步结果表明两者都参与调节特定神经元及其树突在 x-y 平面上的排列。此外，两者都影响相同的细胞类型，星爆无长突细胞（SAC），但以不同的方式：MEGF10/11 调节 SAC 胞体的嵌合排列，而 Pcdhgs 是其树突自我回避所必需的。现在，我们将使用这些结果作为起点，深入了解电路组装这些常见但研究很少的方面背后的机制。首先，我们将使用体内功能获得和丧失的方法来表征 MEGF10/11 在马赛克形成中的作用。我们会问MEGF10/11是否只在发育过程中有效，它的作用是否持久，它是否可以在马赛克形成后破坏它们，以及这两种同源物是否有不同的作用。其次，我们将询问在条件性 Pcdhg 突变小鼠中观察到的自我回避缺陷是否是细胞自主的，以及它们是否反映了树突形成或细化的问题。然后，我们将使用遗传方法来减少视网膜细胞表达的 Pcdhg 同工型。因此，我们可以测试异构体多样性在此过程中的作用，并了解不同的异构体是否发挥不同的作用。第三，我们将询问除 SAC 之外的视网膜亚型是否使用 MEGF10/11 或 Pcdhgs 来图案化其体细胞或乔木。最后，我们将结合体外和体内研究来分析 MEGF10/11 和 Pcdhgs 发挥作用的信号机制。我们将询问 MEGF10/11 是否充当受体、配体或同时充当配体和受体（即同源性）。对于 Pcdhgs，我们将询问 Pcdhgs 似乎促进的最初粘附相互作用如何转化为自我回避所需的排斥性相互作用。总之，这些结果将进一步加深我们对两个知之甚少的基因家族以及一组对神经回路模式重要的知之甚少的过程的理解。 公共卫生相关性：无数神经元类型之间的特定联系是大脑功能的基础，而连接缺陷是某些精神疾病的基础。连接性的一个关键决定因素是每种特定类型的神经元及其轴突或树突的有序排列。该项目使用视网膜（中枢神经系统的一个紧凑且可访问的部分）来分析神经元表面上参与建立这些排列的两组蛋白质。

DESCRIPTION (provided by applicant): Orderly and specific connections among neurons of multiple subtypes underlie the function of neural circuits. Our own work has used retina as a model to elucidate molecules and mechanisms that underlie specificity, focusing on the matching of pre- and postsynaptic partners in particular synaptic laminae. There is another sort of order, however, that has received less attention: the arrangement of cells and their neurites in the orthogonal (x-y) plane. Processes that contribute to this arrangement include mosaic spacing of neuronal somata, tiling of dendrites, and self-avoidance of processes within a single arbor. These processes are believed to ensure uniform coverage of the visual field, precise connectivity, and appropriate receptive field size. Their molecular bases remain unknown in vertebrates. Recently, we began analyzing two sets of cell surface proteins that we suspected to be involved in laminar specificity: MEGF10 and 11, and a cluster of 22 related gamma protocadherins (Pcdhgs). Unexpectedly, preliminary results suggest that both are involved in regulating the arrangement of specific neurons and their dendrites in the x-y plane. Moreover, both affect the same cell type, starburst amacrine cells (SACs), but in different ways: MEGF10/11 regulate the mosaic arrangement of SAC somata whereas Pcdhgs are required for self-avoidance of their dendrites. We will now use these results as starting points to obtain insights into the mechanisms that underlie these common but little-studied aspects of circuit assembly. First, we will use gain- and loss-of function methods in vivo to characterize the role of MEGF10/11 in mosaic formation. We will ask whether MEGF10/11 is effective only during development, whether its effects endure, whether it can disrupt mosaics after they form, and whether the two homologues have distinct effects. Second, we will ask whether defects in self-avoidance observed in conditional Pcdhg mutant mice are cell-autonomous and whether they reflect problems in dendrite formation or refinement. We will then use genetic methods to reduce the repertoire of Pcdhg isoforms that retinal cells express. We can thereby test the role of isoform diversity in the process and learn whether different isoforms play different roles. Third, we will ask whether retinal subtypes other than SACs use MEGF10/11 or Pcdhgs to pattern their somata or arbors. Finally, we will combine studies in vitro and in vivo to initiate analyses of the signaling mechanisms by which MEGF10/11 and Pcdhgs function. We will ask whether MEGF10/11 act as receptors, as ligands, or as both ligand and receptor (that is, homophilically). For Pcdhgs, we will ask how the initially adhesive interaction that Pcdhgs appear to promote is translated into the repellent one required for self-avoidance. Together, these results will further our understanding of two poorly understood gene families and of a poorly understood set of processes important for patterning neural circuits. PUBLIC HEALTH RELEVANCE: Specific connections among myriad neuronal types underlie brain functions, and defects in connectivity underlie some psychiatric disorders. A critical determinant of connectivity is the orderly arrangement of the neurons of each particular type, and of their axonal or dendritic processes. This project uses the retina, a compact and accessible portion of the central nervous system, to analyze two groups of proteins on the neuronal surface that are involved in establishing these arrangements.