Transcriptional Control of Motor Neuron Identity and Connectivity

描述（由申请人提供）：脊椎动物的运动行为需要在脊髓运动神经元和外周肌肉目标之间建立选择性连接。Hox转录因子的调控网络与运动分化的两个关键步骤直接相关：建立柱状身份，将运动轴突定向到特定的靶区；一列神经元的多样化形成了运动池，每个运动池针对一个肌肉。Hox蛋白参与运动神经元柱状和池状身份的分子机制尚不清楚。我们发现叉头同源结构域转录因子FoxP1在两个对hox敏感的运动神经元柱状亚型，即外侧运动柱（LMC）和神经节前柱（PGC）中选择性表达。本研究的目的是进一步阐明运动神经元中FoxP1和Hox的活动是如何协调调节的，并阐明在运动神经元身份的Hox依赖程序中至关重要的下游途径。本研究的第一个目的是探索Hox蛋白对FoxP1表达的调控，以及FoxP1在运动神经元亚型中选择性表达的机制。在这个目的中，我们将通过FoxP1在体内的过表达来确定FoxP1蛋白水平对运动神经元柱状身份的建立的影响。第二个目的是研究Foxp1基因缺失对运动神经元身份和与肌肉靶点连接的影响。我们将使用解剖学和组织学分析来研究Foxp1在肢体运动轴突投射的初始模式建立和确定突触靶点选择中的作用。在第三个目标中，将探讨FoxP1和Hox蛋白在控制运动神经元特异性基因表达中的生化相互作用。FoxP1与LMC运动神经元和特定池中表达的大部分或全部基因直接相互作用的假设将使用染色质免疫沉淀法进行检验。然后，我们将使用体外和体内试验来检查FoxP1和Hox蛋白之间相互作用的后果。总之，这些研究应该有助于更好地理解运动神经元多样性是如何产生的，并为确定神经系统其他区域神经元突触特异性的机制提供一些基本见解。

DESCRIPTION (provided by applicant): Locomotor behavior in vertebrates requires the establishment of selective connections between motor neurons in the spinal cord and muscle targets in the periphery. A regulatory network of Hox transcription factors has been directly linked with two critical steps in motor differentiation: the establishment of columnar identities which directs motor axons toward a specific target field; and the diversification of neurons within a column into motor pools, each pool targeting a single muscle. The molecular mechanisms by which Hox proteins contribute to motor neuron columnar and pool identities are not known. We have found that the forkhead homeodomain transcription factor FoxP1 is selectively expressed by two Hox-sensitive motor neuron columnar subtypes, the lateral motor column (LMC) and preganglionic column (PGC). The aim of this proposal is to further elucidate how the activities FoxP1 and Hox are coordinately regulated in motor neurons and to elucidate the downstream pathway that are critical in the Hox-dependent programs of motor neuron identity. The first aim of this proposal will explore the regulation of FoxP1 expression by Hox proteins and the mechanisms by which FoxP1 becomes selectively expressed in a subset of motor neuron subtypes. In this aim we will determine the influences of FoxP1 protein levels on the establishment of motor neuron columnar identities through overexpression of FoxP1 in vivo. In the second aim the impact of loss Foxp1 on motor neuron identity and connectivity with muscle targets will be examined. We will use anatomical and histological assays to examine the role of Foxp1 in establishing the initial patterns of motor axon projections in the limb and in defining the selection of synaptic targets. In the third aim biochemical interactions of FoxP1 and Hox proteins in the control of motor neuron-specific gene expression will be explored. The hypothesis that FoxP1 interacts directly with most or all of genes expressed in LMC motor neurons and in specific pools will be examined using chromatin immunoprecipitation assays. We will then examine the consequences of interactions between FoxP1 and Hox proteins using in vitro and in vivo assays. Together, these studies should help to provide a better understanding of how motor neuron diversity is generated and provide some of the basic insights into the mechanisms that determine the synaptic specificity of neurons in other regions of the nervous system. PUBLIC HEALTH RELEVANCE: One of the major challenges in the neural sciences is to understand how specific connections are made between neurons and their synaptic targets. The overall goal of this proposal is to elucidate the developmental programs that define the ability of motor neurons in the spinal cord to make very selective connections with muscle targets. Understanding the steps that determine the intrinsic properties of motor neurons may be essential in designing treatment strategies after spinal cord injury.