The Role of MicroRNAs in Axon Pathfinding

项目摘要 在发育中的神经系统中，适当的轴突生长和寻路受到多种因素的调节。 引导分子、它们的受体和细胞内信号通路。生物多样性的时空调控 引导受体的表达促进神经元对引导线索的不同反应以形成 准确的神经元布线。在脊椎动物中，引导受体Robo1的水平在杂交前和 在交叉后连合轴突(CA)的高位，一种模式发挥着‘分子开关’调节 对缝隙排斥力敏感，并引导CA中线交叉。然而，罚款背后的机制是- 对Robo1表达的时空调节在很大程度上仍不清楚。MicroRNAs(MiRNAs) 通过与靶mRNAs的3‘非翻译区(3’UTR)特异性结合来调节基因表达，从而 抑制翻译和/或诱导mRNA降解。我们的初步研究表明，Robo1 在发育中的脊髓中，3‘非编码区是蛋白质表达调控所必需的。GGA-MIR-92，一种高度 保守的miRNA，在发育中的鸡脊髓中差异表达，并调节缝隙 抑制连合神经元cRobo1的表达从而控制CA投射的敏感性 和中线交叉。成熟的gga-miR-92与MMU-miR-92a和MMU-miR-92b具有相同的序列 除MMU-miR-92b 5‘端的第10个碱基外。MMU-miR-92b也能抑制mRobo1的表达 通过瞄准它的3‘UTR。因此，我们假设MMU-miR-92a和MMU-miR-92b与gga-miR-92一样， 特异性调节连合神经元mRobo1的表达以控制Sit/Robo1介导的CA 指导小鼠脊髓发育。我们将研究MMU-miR-92a和MMU-miR- 92B在(1)抑制发育中的连合神经元中mRobo1的表达(目标1)和(2) Sit/mRobo1介导的脊髓CA引导(目标2)。对这项建议的研究将增进我们的理解 Sit/Robo介导的轴突引导的分子机制。

Project Summary In the developing nervous system, proper axon outgrowth and pathfinding are regulated by multiple guidance molecules, their receptors and intracellular signaling pathways. Spatiotemporal regulation of guidance receptor expression facilitates differential responses of neurons to guidance cues in order to form accurate neuronal wiring. In vertebrates, levels of the guidance receptor Robo1 are low in precrossing and high in postcrossing commissural axons (CAs), a pattern functioning as a ‘molecular switch’ to regulate sensitivity to Slit repulsion and guide CA midline crossing. However, the mechanism underlying the fine- tuned spatiotemporal regulation of Robo1 expression remains largely unknown. MicroRNAs (miRNAs) regulate gene expression by binding specifically to the 3’untranslated region (3’UTR) of target mRNAs, thus repressing translation and/or inducing mRNA degradation. Our preliminary studies indicate that the Robo1 3’UTR is required for regulation of protein expression in the developing spinal cord. Gga-miR-92, a highly conserved miRNA, is differentially expressed in the developing chicken spinal cord, and regulates Slit sensitivity via suppression of cRobo1 expression in commissural neurons, thereby controlling CA projection and midline crossing. The mature gga-miR-92 has the same sequence as mmu-miR-92a and mmu-miR-92b except the tenth base at the 5’ end of mmu-miR-92b. Mmu-miR-92b can also repress mRobo1 expression by targeting its 3’UTR. Therefore, we hypothesize that mmu-miR-92a and mmu-miR-92b, like gga-miR-92, specifically regulate mRobo1 expression in commissural neurons to control Slit/Robo1-mediated CA guidance in the developing mouse spinal cord. We will examine the role of mmu-miR-92a and mmu-miR- 92b in (1) suppressing mRobo1 expression in developing commissural neurons (Aim 1) and (2) Slit/mRobo1-mediated spinal CA guidance (Aim 2). Studies in this proposal will advance our understanding of molecular mechanisms underlying Slit/Robo-mediated axon guidance.