Mechanisms of Morphogen Secretion in Visual System Development and Disease

视觉系统发育和疾病中形态原分泌的机制

基本信息

批准号：
8621495
负责人：
Samuel M Kunes
金额：
$ 42.25万
依托单位：
HARVARD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-01-01 至 2018-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8621495
关键词：
Address Adult Animals Apical Area Axon Biochemical Biochemical Pathway Biogenesis Biological Biology Brain Bypass C-terminal Caliber Cell Communication Cells Child Cholesterol Cleaved cell Complex Defect Development Disease Drosophila genus Environment Equilibrium Erinaceidae Eye Fatty acid glycerol esters Genes Genetic Screening Golgi Apparatus Human Milk Image In Vitro Intervention Kinetics Life Lipids Malignant Neoplasms Mediating Methods Modeling Modification Molecular Movement N-terminal Natural regeneration Neurons Organelles Pathology Pathway interactions Photoreceptors Process Protein Export Pathway Protein Family Proteins Reporting Retina Role Shapes Signal Transduction Signaling Molecule Sorting - Cell Movement Synapses System Time Tissues Touch sensation Travel Variant Visual system structure autism spectrum disorder compound eye developmental disease genetic analysis in vivo in vivo imaging morphogens mutant neural circuit neuron development novel polypeptide protein function protein transport public health relevance smoothened signaling pathway transmission process vision development

项目摘要

PROJECT SUMMARY/ABSTRACT A number of developmental disorders and pathological conditions arise from defects in conserved cell-cell communication pathways, such as those mediated by the Hedgehog family of proteins. Recent studies have documented the important role of protein export mechanisms in shaping the activity of Hedgehog and other signaling molecules in development and disease. Hedgehog can be released apically or basally, in large multimeric or small monomeric forms. It can travel over many cell diameters in long cellular extensions, such as cytonemes and axons, prior to release. The mechanisms underlying Hedgehog export and transmission are complex and unresolved, and yet touch on many cellular mechanisms that go awry with pathological consequences. This proposal addresses this less well-studied area of Hedgehog biology. We will address Hedgehog's export mechanisms, and especially focus on a novel mechanism for axon transport that carries Hedgehog to axon termini on organelles known as lipid droplets. We will thus identify new targets for controlling this pathway in development, regeneration and disease. The Drosophila visual system offers an excellent opportunity to unravel the complexity of Hedgehog export and transmission. In this system, apical Hedgehog secretion propagates the temporal wave of ommatidial development that gives rise to the compound eye; basal targeting, transport and release from photoreceptor axon termini triggers the differentiation of post-synaptic neurons in the brain. Partitioning Hedgehog for release at the opposite poles of a photoreceptor neuron is critical to the coordinated development of these neurons, which assemble into a precise neural circuit. Hedgehog is composed of N-terminal and C-terminal domains that dissociate by self-catalyzed proteolytic cleavage. The N-terminal product, HhNp, harbors all known signaling activities. We recently discovered that polarized HhNp export involves an unusual choice of export pathway that is directed by the C-terminal self- cleavage product HhC. HhNp, when associated with HhC, is incorporated into lipid droplets. These droplets are basally targeted, bypass the Golgi apparatus, and travel by fast axon transport to the brain. Some nascent Hedgehog polypeptide is cleaved near its C-terminus, which removes an axonal targeting signal. HhNp associated with this shortened HhC is secreted apically and remains in the retina. This binary choice of export pathways balances Hedgehog activity between the developing eye and brain. This proposal aims to resolve the molecular details of this novel export mechanism. We will define the cis- acting Hedgehog polypeptide sequences involved in export pathway decisions. Since nothing is known about a role of lipid droplets as axon transport carriers, we will conduct a general and unbiased genetic screen for components of the underlying molecular machineries, and use biochemical and cell biological methods to investigate their functions. Finally, we will employ a new method for live imaging of axon transport in differentiating photoreceptor neurons to analyze the kinetics of Hedgehog and lipid droplet axon transport and characterize newly identified export machinery components. Investigating this novel pathway will uncover a new set of targets for interventions into developmental anomalies and pathologies involving the Hedgehog pathway, protein trafficking and axon transport.

项目概要/摘要许多发育障碍和病理状况是由保守细胞间的缺陷引起的通讯途径，例如由 Hedgehog 蛋白家族介导的途径。最近的研究有记录了蛋白质输出机制在塑造 Hedgehog 和其他细胞活性方面的重要作用发育和疾病中的信号分子。刺猬可以从顶部或底部释放，大多聚体或小单体形式。它可以在长的细胞延伸中穿过许多细胞直径，例如细胞线和轴突，在释放之前。 Hedgehog 输出的机制传播是复杂且尚未解决的，但涉及许多细胞机制，这些机制会出现问题病理后果。该提案解决了刺猬生物学这一研究较少的领域。我们将解决刺猬的输出机制，特别关注轴突运输的新机制它将刺猬携带到被称为脂滴的细胞器上的轴突末端。因此我们将确定新的控制发育、再生和疾病中这一途径的目标。果蝇视觉系统为揭示刺猬输出的复杂性提供了绝佳的机会和传输。在这个系统中，顶端 Hedgehog 分泌物传播小眼的时间波产生复眼的发育；光感受器的基础靶向、运输和释放轴突末端触发大脑中突触后神经元的分化。对 Hedgehog 进行分区感光神经元对极的释放对于这些神经元的协调发展至关重要神经元，组装成精确的神经回路。 Hedgehog 由 N 端和 C 端结构域组成，通过自催化蛋白水解作用解离分裂。 N 末端产物 HhNp 具有所有已知的信号传导活性。我们最近发现极化的 HhNp 输出涉及一种不寻常的输出路径选择，该路径由 C 端自我引导裂解产物HhC。当 HhNp 与 HhC 结合时，HhNp 会并入脂滴中。这些水滴是基础靶向，绕过高尔基体，并通过快速轴突运输到达大脑。一些新生的 Hedgehog 多肽在其 C 末端附近被切割，从而消除了轴突靶向信号。与缩短的 HhC 相关的 HhNp 被分泌到顶部并保留在视网膜中。这种二元选择输出途径的数量平衡了正在发育的眼睛和大脑之间的 Hedgehog 活动。该提案旨在解决这种新型输出机制的分子细节。我们将定义顺式参与输出途径决定的作用刺猬多肽序列。既然什么都不知道关于脂滴作为轴突运输载体的作用，我们将进行一般且公正的遗传研究筛选底层分子机器的成分，并使用生化和细胞生物学方法来研究它们的功能。最后，我们将采用一种新方法对轴突进行实时成像分化感光神经元中的运输以分析 Hedgehog 和脂滴轴突的动力学运输和表征新确定的出口机械部件。研究这条新途径将揭示一套新的干预发育异常和病理的目标，涉及刺猬通路、蛋白质运输和轴突运输。