Engrailed targets and the control of synaptic circuits in Drosophila

果蝇的纠缠目标和突触回路的控制

• 批准号: 8468764
• 负责人: JONATHAN M BLAGBURN
• 金额: $ 36.19万
• 依托单位: UNIVERSITY OF PUERTO RICO MED SCIENCES
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-01 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8468764
• 关键词: Adult; Afferent Neurons; Animals; Auditory; Auditory system; Autistic Disorder; Automobile Driving; Axon; Binding; Biological Assay; Biological Models; Brain; Branchiostoma floridae AmphiEn protein; Cell Adhesion Molecules; Cell surface; Coupling; Culicidae; Data; Databases; Dengue; Development; Disease; Disease Vectors; Drosophila genus; Drosophila melanogaster; Electrophysiology (science); Ensure; Evolution; Fiber; Gene Targeting; Genes; Goals; Health; Human; Insecta; Knock-out; Knockout Mice; Knowledge; Label; Leucine-Rich Repeat; Link; Malaria; Mammals; Measures; Methods; Midbrain structure; Modeling; Molecular; Morphology; Neuraxis; Neurons; Parkinson Disease; Partner in relationship; Pathway interactions; Pattern; Pilot Projects; Play; Positioning Attribute; Process; Protein Overexpression; Proteins; RNA Interference; Regulation; Repression; Research; Role; Specificity; Symptoms; Synapses; System; Testing; autism spectrum disorder; axon guidance; axonal guidance; axonal pathfinding; connectin; design; disorder control; dopaminergic neuron; feeding; fly; human disease; innovation; knock-down; member; nervous system disorder; neural circuit; neurobiotin; neuroglian; neuron development; neuronal survival; overexpression; synaptogenesis; transcription factor; visual stimulus

DESCRIPTION (provided by applicant): Engrailed (En) is a transcription factor first discovered in Drosophila but later found to be present in all animals, playing an important role in controllin neuronal development. Little is known about the cell surface molecules that it regulates. The long-term goal of this research is to find out how En regulates synaptic connectivity in the CNS, with a particular focus on identifying and characterizing its target genes of cell surface effector molecules. Preliminary data show that overexpression of En in Drosophila olfactory neurons alters their axonal path finding to their targets, the olfactory glomeruli. Additionally, ectopic E expression in a normally En- negative subset of auditory neurons allows them to form synaptic connections with the Giant Fiber (GF) escape neuron. The first aim is to selectively knock out En in olfactory neurons using fly lines in which RNAi is driven by the Gal4-UAS system. I will determine the effects on axonal guidance by assaying changes in the morphology of GFP-labeled olfactory axons, and alterations in the positions of immunolabeled olfactory glomeruli. The second aim will drive En RNAi in the auditory neurons and measuring their synaptic input to the GF. The third aim is to test whether increasing or knocking down expression of the En target Connectin, a member of the conserved LRR superfamily of adhesion molecules, alters axon guidance or glomerulus positioning. I will also test whether En knockout results in overexpression of this protein, as would be expected if it is a target of En repression. In the finl aim, I will ectopically express or knock down the En-binding target gene Neuroglian, a cell surface adhesion molecule homologous to vertebrate L1-CAM, then use the auditory synapse assays of connection to the GF. I will also test whether its immunostaining is altered by overexpression or knockout of En, as would be expected if it is negatively regulated by En. Relevance: En has been shown to control the survival of midbrain dopaminergic neurons, with En knockout mice showing Parkinson-like symptoms, and it has also been linked to autism spectrum disorder. Drosophila models are particularly useful for the discovery of molecular pathways that are directly relevant to human health, because most of these pathways have been conserved during evolution. All animals have En protein, so it is very likely that any molecules that are regulated by it during the process of synapse formation in Drosophila have their counterparts in humans, playing similar roles. These molecules may be of great potential importance in neurological diseases such as Parkinson's or autism. In addition, the basic knowledge gained from this project about the development of dipteran olfactory and auditory systems could be of use in designing ways to disrupt the feeding and mating patterns of mosquitoes, which are vectors of diseases such as malaria and dengue fever.

描述(申请人提供)：Engrailed(EN)是一种转录因子，最初在果蝇中发现，但后来发现在所有动物中都存在，在控制神经元发育方面发挥重要作用。人们对它所调控的细胞表面分子知之甚少。这项研究的长期目标是找出en如何调节中枢神经系统中的突触连接，重点是识别和表征其细胞表面效应的靶基因。 分子。初步数据显示，果蝇嗅觉神经元中EN的过度表达改变了它们通往目标嗅小球的轴突路径。此外，在正常EN阴性的听神经元亚群中异位E的表达使它们能够与巨纤维逃逸神经元形成突触连接。第一个目标是利用由Gal4-UAS系统驱动RNAi的飞线选择性地敲除嗅觉神经元中的EN。我将通过分析GFP标记的嗅觉轴突的形态变化和免疫标记的嗅球位置的变化来确定对轴突引导的影响。第二个目标是在听觉神经元中驱动en RNAi，并测量它们对GF的突触输入。第三个目的是测试EN靶连接素表达的增加或下调是否改变轴突导向或肾小球的位置。EN靶连接素是保守的LRR超家族黏附分子中的一员。我还将测试en基因敲除是否会导致该蛋白的过度表达，如果它是en抑制的目标，这一点是可以预料的。在FINL目标中，我将异位表达或敲除EN结合的靶基因Neuroglian，这是一种与脊椎动物L1-CAM同源的细胞表面黏附分子，然后使用与GF连接的听觉突触分析。我还将测试它的免疫染色是否会因EN的过度表达或敲除而改变，如果它被EN负调控，预计会发生变化。相关性：EN已被证明可以控制中脑多巴胺能神经元的存活，EN基因敲除的小鼠表现出帕金森样症状，它也与自闭症谱系障碍有关。果蝇模型对于发现与人类健康直接相关的分子途径特别有用，因为这些途径中的大多数在进化过程中都是保守的。所有的动物都有EN蛋白，所以很可能在果蝇突触形成过程中受EN蛋白调控的任何分子都有与人类相同的分子，扮演着类似的角色。这些分子在帕金森氏症或自闭症等神经系统疾病中可能具有巨大的潜在重要性。此外，从这个项目中获得的关于双翅目昆虫嗅觉和听觉系统发育的基本知识，可能有助于设计方法来扰乱蚊子的进食和交配模式，蚊子是疟疾和登革热等疾病的媒介。