Molecular mechanisms of motor neuron development

描述（由申请人提供）：本提案的主要目标是了解运动神经元分化和规范的分子机制。类维甲酸（RA）信号是脊髓运动神经元分化过程中同步神经发生和运动神经元分化通路所必需的，这些事件是由RA应答基因GDE2介导的。GDE2编码具有胞外甘油磷酸二酯磷酸二酯酶（GDPD）结构域的六跨膜蛋白。GDPD活性是GDE2协调细胞周期退出和运动神经元规范的能力所必需的，揭示了GDPD代谢和运动神经元分化之间的新联系。GDE2的独特拓扑结构（GDPD结构域位于细胞外）和缺乏功能先例强烈预测了参与运动神经元分化的新分子网络的发现。为了确定这样的网络，使用无偏筛选来分离与GDE2相互作用的蛋白质。确定了两种不同信号通路的组成部分。在本研究中，我们将结合体外结构-功能分析和体内功能损失和增益研究来研究这些信号通路如何与GDE2整合以促进运动神经元分化。虽然GDE2的表达始于分化细胞，但GDE2的表达在终末分化的运动神经元中保持不变；这表明GDE2可能对后期运动神经元功能或存活具有重要作用。为了确定GDE2在运动神经元发育的不同阶段的功能，我们将使用小鼠遗传学来清除分化和有丝分裂后运动神经元中的GDE2。由此产生的胚胎将在运动神经元分化、运动神经元规格、运动轴突目标识别和运动神经元存活方面进行缺陷分析。这些实验将确定两种不同的信号通路如何与GDPD依赖的神经元分化模式协同作用，并进一步推进目前对参与细胞分化的调节网络的理解。此外，这些研究将探讨GDE2在多种对神经元功能和存活至关重要的细胞过程中的潜在作用。

DESCRIPTION (provided by applicant): The major goals of this proposal are to understand the molecular mechanisms of motor neuron differentiation and specification. Retinoid (RA) signals are necessary for synchronizing neurogenic and motor neuron specification pathways during spinal motor neuron differentiation and these events are mediated by the RA-responsive gene, GDE2. GDE2 encodes a six transmembrane protein with an extracellular glycerophosphodiester phosphodiesterase (GDPD) domain. GDPD activity is required for GDE2's ability to coordinate cell-cycle exit and motor neuron specification, revealing a novel link between GDPD metabolism and motor neuron differentiation. The distinctive topology of GDE2 where the GDPD domain is extracellular and the lack of functional precedent strongly predicts the discovery of new molecular networks involved in motor neuron differentiation. To identify such networks, unbiased screens were used to isolate proteins that interact with GDE2. Components of two different signaling pathways were identified. In this proposal, in vitro structure-function analyses will be combined with in vivo loss- and gain- of function studies to investigate how these signaling pathways integrate with GDE2 to promote motor neuron differentiation. While onset of GDE2 expression occurs in differentiating cells, GDE2 expression is maintained in terminally differentiated motor neurons; suggesting that GDE2 may have additional roles critical for later motor neuron function or survival. To define the function of GDE2 at different stages of motor neuron development, mouse genetics will be used to ablate GDE2 in differentiating and postmitotic motor neurons. Resultant embryos will be analyzed for defects in motor neuron differentiation, motor neuron specification, motor axon target recognition and motor neuron survival. These experiments will determine how two different signaling pathways synergize with GDPD- dependent modes of neuronal differentiation and further advance current understanding of the regulatory networks involved in cell differentiation. In addition, these studies will investigate potential roles for GDE2 in diverse cellular processes critical for neuronal function and survival.