Transcriptional Control of Motor Neuron Identity and Connectivity

描述(申请人提供)：脊椎动物的运动行为需要在脊髓中的运动神经元和周围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，将运动轴突引向特定的靶区；以及柱状神经元多样化成运动池，每个运动池针对一块肌肉。HOX蛋白对运动神经元的柱状和池特性的贡献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选择性地表达于两种对HOX敏感的运动神经元柱状亚型，即外侧运动柱(LMC)和节前柱(PGC)。这一建议的目的是进一步阐明运动神经元中FoxP1和Hox活性是如何协调调节的，并阐明在依赖Hox的运动神经元识别程序中至关重要的下游通路。这项建议的第一个目的是探索HOX蛋白对FoxP1表达的调节，以及FoxP1在运动神经元亚型中选择性表达的机制。为此，我们将通过体内FoxP1的过表达来确定FoxP1蛋白水平对运动神经元柱状身份建立的影响。在第二个目标中，我们将研究Foxp1的缺失对运动神经元的同一性和与肌肉靶标的连接的影响。我们将使用解剖学和组织学分析来研究Foxp1在建立肢体运动轴突投射的初始模式和确定突触靶点的选择中的作用。在第三个目标中，将探索FoxP1和Hox蛋白在控制运动神经元特异性基因表达方面的生化相互作用。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通过染色质免疫沉淀分析进行验证。然后，我们将使用体外和体内分析来研究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，并为决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的见解。 公共卫生相关性：神经科学中的主要挑战之一是了解神经元和它们的突触目标之间是如何建立特定联系的。这项提案的总体目标是阐明定义脊髓运动神经元与肌肉靶标进行非常选择性连接的能力的发育程序。了解决定运动神经元内在特性的步骤可能对设计脊髓损伤后的治疗策略至关重要。

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.