CELL INTERACTIONS IN MOTOR NEURON DIFFERENTIATION

在发育中的脊髓中，本体感觉神经元和运动神经元之间形成的连接的特异性依赖于一系列严格控制的细胞相互作用，最终导致传入感觉传入识别特定的运动神经元树突。新的证据表明，由感觉神经元和运动神经元的功能亚群表达的转录因子指导选择性脑脊髓炎的发生 感觉-运动连接。一个重要的悬而未决的问题是控制运动和感觉轴突生长和连接的转录因子的相关靶标的一致性。这项建议的主要目的是确定脊髓运动和感觉神经元身份的转录调控与控制轴突生长的关键细胞表面识别分子之间的联系，以及该神经回路中的靶标特异性。本课程将讨论运动神经元和感觉神经元发育的三个具体方面。 位于外侧运动柱内的运动神经元在LIM同源结构域转录因子的控制下，沿着肢体背腹轴以不同的方式投射轴突。我们将使用小鼠遗传方法来探索LIM同源结构域蛋白通过调节运动轴突和肢体间充质细胞上Eph激酶和ephins的表达来指导运动轴突导引的可能性。 Ets类转录因子调节运动神经元和感觉神经元的运动轨迹和终末轴突分支，遗传学研究表明Ets基因调节运动神经元中信号素的表达。我们将利用小鼠遗传学方法绘制特定信号素及其主要受体丛状蛋白在运动神经元和感觉神经元分化中的分布并分析其功能，重点关注这些基因在感觉神经轴突生长和连接中的功能。 识别LIM同源结构域和ETS基因的相关靶点的一个困难是可以从小鼠胚胎获得的原代神经元的数量有限。为了解决这个问题，我们将利用从小鼠ES细胞产生基本上无限数量的运动神经元的能力，以及发现ES细胞来源的运动神经元缺乏许多LIM同源结构域和ETS蛋白的表达，这些蛋白调节运动轴突生长和连接。 因此，ES细胞来源的运动神经元将被用作基于DNA的微阵列筛选的细胞系统，以定义和研究由这些转录因子激活或抑制的额外靶基因的功能。

The specificity of connections formed between proprioceptive sensory neurons and motor neurons in the developing spinal cord is dependent on a series of tightly controlled cellular interactions that culminate in the recognition of specific motor neuron dendrites by incoming sensory afferents. Emerging evidence indicates that transcription factors expressed by functional subsets of sensory and motor neurons direct the development of selective sensory-motor connections. One important unresolved issue is the identity of the relevant targets of the transcription factors that control motor and sensory axon growth and connectivity. The main goal of this proposal is to define the link between the transcriptional regulation of spinal motor and sensory neuron identity and the key cell surface recognition molecules that control axonal growth and target specificity in this neural circuit. Three specific aspects of motor and sensory neuron development will be addressed. Motor neurons located within lateral motor column project their axons differentially along the dorsoventral axis of the limb, under the control of LIM homeodomain transcription factors. We will use mouse genetic methods to explore the possibility that LIM homeodomain proteins direct motor axon guidance by regulating the expression of Eph kinases and ephrins on motor axons and limb mesenchymal cells. ETS class transcription factors regulate the trajectory and terminal axonal branching of both motor and sensory neurons, and genetic studies have revealed that ETS genes regulate semaphorin expression in motor neurons. We will use mouse genetic approaches to map the distribution, and analyze the function, of specific semaphorins and their major receptors, the plexins, in the differentiation of motor and sensory neurons, focusing on the function of these genes in sensory axonal growth and connectivity. One difficulty in identifying relevant targets of LIM homeodomain and ETS genes has been the limitation in number of primary neurons that can be obtained from mouse embryos. To overcome this problem, we will take advantage of the ability to generate motor neurons in essentially unlimited numbers from mouse ES cells, and the finding that ES cell-derived motor neurons lack expression of many of the LIM homeodomain and ETS proteins that regulate motor axonal growth and connectivity. ES cell derived motor neurons will therefore be used as a cellular system for DNA-based microarray screens to define and study the function of additional target genes that are activated or repressed by these transcription factors.