Linking motorneuron fate and connectivity in Drosophila

描述(申请人提供)：我们的长期目标是在果蝇中使用遗传学方法来了解运动神经元命运指定和分化的分子机制。在这项建议中，我们将重点放在运动神经元命运与轴突指导之间的联系机制上。在对影响运动神经元发育的突变体的大规模筛选中，我们发现了FOXO和MMP2，这两种基因都在运动神经元亚群中表达，是正确引导运动轴突所必需的。我们专注于这两个进化上保守的蛋白质，因为它们为我们阐明运动神经元的不同行为是如何发育调节的提供了新的切入点。我们的目标是完成这些基因的分子、遗传和表型分析，以便了解观察到的突变表型。FOXO是一种转录因子，以其在胰岛素信号通路中的作用而闻名，在胰岛素信号通路中，FOXO受细胞外信号调节。我们的初步数据表明，FoxO在运动神经元簇中特异性地表达，以响应靶点衍生的信号。详细的表达和表型分析将阐明FoxO在运动神经元发育中的作用。此外，我们将通过分子和遗传上位性实验确定调控运动神经元FoxO表达的信号通路(S)。基质金属蛋白酶(MMPs)是一大家族。跨膜和分泌型的蛋白水解酶几乎可以分解细胞外基质的每一个组成部分。MMP2在固定的运动神经元群体中表达，是正确的运动轴突导引所必需的。我们将通过分析MMP2突变胚胎中运动轴突起的生长来表征MMP2在有丝分裂后神经元中的表达模式，并阐明其在运动轴突起引导中的作用。此外，我们将通过分析MMP2和已知调节这一关键路径步骤的引导分子之间的遗传相互作用来确定MMP2是否对运动轴突脱散是必需的。这些研究将促进我们对不同运动神经元群体中的分子如何协调建立适当的神经肌肉连接模式的理解。运动神经元分化是神经元发育过程中的重要事件，在人类发育和疾病过程中会受到干扰。此外，由于FOXO和MMP2都是进化上保守的蛋白质，在具有强烈临床意义的通路中发挥作用，这些研究应该具有广泛的生物学和医学意义。

DESCRIPTION (provided by applicant): Our long term aim is to use a genetic approach in Drosophila to understand molecular mechanisms of motorneuron fate specification and differentiation. In this proposal, we focus on mechanisms linking motorneuron fate to axon guidance. In a large-scale screen for mutants affecting motorneuron development, we have identified foxO and Mmp2, both of which are expressed in motorneuron subsets and necessary for proper motor axon guidance. We focus on these two evolutionarily-conserved proteins as they provide us with novel entry points for elucidating how distinct behaviors of motorneurons are developmentally regulated. Our goals here are to complete molecular, genetic, and phenotypic analyses of these genes in order to understand the observed mutant phenotypes. FoxO is a transcription factor best known for its role in the insulin signaling pathway where it is regulated by extracellular signals. Our preliminary data suggest that FoxO is expressed specifically in clusters of motorneurons in response to a target-derived signal. Detailed expression and phenotypic analyses will elucidate the role of FoxO in motorneuron development. Furthermore, we will identify the signaling pathway(s) regulating FoxO expression in motorneurons through molecular and genetic epistasis experiments. Matrix metalloproteinases (Mmps) comprise a large family of. transmembrane and secreted proteases that together cleave nearly every component of the ECM. Mmp2 is expressed in stereotyped populations of motorneurons and is necessary for proper motor axon guidance. We will characterize the expression pattern of Mmp2 in post-mitotic neurons and elucidate its role in motor axon guidance by analyzing motor axon outgrowth in Mmp2 mutant embryos. Additionally, we will establish whether Mmp2 is necessary for motor axon defasciculation by analyzing genetic interactions between Mmp2 and guidance molecules known to regulate this key pathfinding step. These studies will advance our understanding of how molecules acting in distinct motorneuron populations coordinate to establish proper patterns of neuromuscular connectivity. Motorneuron differentiation is an essential event in neuronal development and can be disrupted in human development and disease. Furthermore, since both foxO and Mmp2 are evolutionarily-conserved proteins acting in pathways of intense clinical interest, these studies should have broad biological and medical significance.