Neuronal events and molecular pathways in active dendrite degeneration

描述（由申请人提供）：树突和轴突从神经元细胞体延伸很远，因此可以独立于神经元细胞体受到损伤。已知轴突具有自己的主动退化程序，该程序是单独控制的，并且在分子上与细胞体不同。目前尚不清楚树突是否具有类似的程序。树突与轴突一样对正常的神经元功能至关重要，在癫痫发作和中风时树突表现出形态变化。因此，我们将在一个简单的模型系统中研究树突退化的过程。果蝇树突状树突神经元是幼虫角质层下的感觉神经元，在复杂程度上与中枢神经元相似。我们可以在每一种动物身上识别出同一个细胞，并且可以用紫外激光从这个细胞中分离出一个单独的树突。我们有初步证据表明，这种损伤引发了一种类似于横断后轴突变性（Wallerian degeneration）的活性变性程序：远端树突在24小时内完全解体，这种解体被Wld(s)或Wallerian degeneration slow蛋白的表达所阻断。在本提案中，我们概述了一系列实验，以确定发生在树突变性过程中的主要事件和执行变性程序的分子途径。我们将利用我们在实时成像和神经元基本细胞生物学方面的专业知识来确定哪个亚细胞系统首先被拆除：肌动蛋白、微管或膜运输。我们将使用现有的遗传工具来阻断已知的自毁机制，包括半胱天冬酶和泛素蛋白酶体系统，以确定树突是否使用凋亡或轴突变性程序。我们还将研究候选细胞器，线粒体和溶酶体，是否对拆除树突很重要。从我们的实验中，我们知道过表达Wld(s)蛋白是可能阻止树突变性程序的。因此，我们将筛选候选基因和转基因果蝇的过表达文库，以确定改变树突变性过程的其他方法。使用一个简单的模型系统来识别树突退化过程中的主要细胞和分子事件，以及改变它们的方法，将迅速推进我们对神经元如何应对包括癫痫和中风在内的环境压力的理解。公共卫生相关性：树突在包括癫痫和中风在内的压力下表现出损伤和退化的形态学迹象。我们将使用一个简单的模型系统来研究树突退化过程中发生的细胞和分子事件，并将确定可以减缓退化程序的蛋白质。

DESCRIPTION (provided by applicant): Dendrites and axons extend long distances from the neuronal cell body and so can be damaged independently from it. It is known that axons possess their own active degeneration program that is controlled separately and is molecularly distinct from that of the cell body. It is not known whether dendrites possess a similar program. Dendrites are as important for normal neuronal function as axons, and exhibit morphological changes during seizures and stroke. We will therefore investigate the process of dendrite degeneration in a simple model system. Drosophila dendritic arborization neurons are sensory neurons under the larval cuticle that elaborate large dendritic trees similar in complexity to central neurons. We can identify the same individual cell in every animal, and can sever a single dendrite from this cell with a UV laser. We have preliminary evidence that this injury elicits an active degeneration program similar to axon degeneration after transection (Wallerian degeneration): distal dendrites are completely disassembled within 24 hours, and this disassembly is blocked by expression of the Wld(s), or Wallerian degeneration slow, protein. In this proposal, we outline a series of experiments to identify the major events that occur during dendrite degeneration and the molecular pathways that execute the degeneration program. We will use our expertise in live imaging and the basic cell biology of neurons to determine which subcellular systems are dismantled first: actin, microtubules or membrane trafficking. We will use available genetic tools to block known self-destruct machinery including caspases and the ubiquitin proteasome system to determine whether dendrites use the apoptotic or axonal degeneration program. We will also investigate whether candidate organelles, mitochondria and lysosomes, are important for dismantling dendrites. From our experiments overexpressing the Wld(s) protein, we know that it is possible to block the program of dendrite degeneration. We will therefore screen candidate genes and an overexpression library of transgenic Drosophila to identify additional ways to alter the course of dendrite degeneration. Using a simple model system to identify major cellular and molecular events during dendrite degeneration, and ways to change them, will rapidly advance our understanding of how neurons respond to environmental stresses including seizures and stroke. PUBLIC HEALTH RELEVANCE: Dendrites exhibit morphological signs of damage and degeneration under stresses including seizures and stroke. We will use a simple model system to investigate the cellular and molecular events that take place during dendrite degeneration, and will also identify proteins that can slow the degeneration program.