Olfactory Adaptation in C. Elegans

描述(申请人提供)：虽然神经元必须能够对刺激做出可靠的反应，但同样重要的是，它要根据特定的经验来改变其反应。了解这种可塑性的分子和细胞基础以及如何发生刺激特有的可塑性变化非常重要，因为它既是学习和记忆等正常过程的基础，也是成瘾和抑郁等疾病状态的基础。我们的目标是利用遗传上易驯化的线虫的嗅觉反应来确定小RNA调控途径是否指导刺激特异性神经元的可塑性。重复刺激改变神经元反应性的一种方式是通过表观遗传“标记”引起的转录变化，如DNA甲基化和组蛋白修饰(回顾文献1)。在S.pombe、植物和果蝇中，染色质“标记”已被证明是由小RNA2引导的。这些表观遗传变化被认为调节着重要的发育过程。作为特定行为的结果，小RNA是否可以动态地调节神经元的表观遗传变化还没有得到检验。一个有吸引力但完全未经检验的假说是，小RNA可能为引导神经元活动发生长期变化的表观遗传事件提供指导和特异性。作为检验这一假说的第一步，我们询问了在解剖学上简单但遗传功能强大的模式生物线虫中，RNA干扰(RNAi)所需的基因是否可能是神经元可塑性所必需的。线虫天生就会被通过G蛋白偶联受体(GPCRs)感觉到的特定气味所吸引，然而，如果气味不伴随食物，它的吸引力就会减弱。我们称之为对气味嗅觉适应反应的经验依赖性抑制。开启嗅觉适应的关键“开关”是cGMP依赖的蛋白激酶(PKG)EGL-4进入气味刺激的感觉嗅觉神经元AWC的核(Lee等人提交)。在我们的初步研究中，我们发现一类特定的小RNA在气味暴露时与感觉神经元中编码染色质相关蛋白HPL-2(组蛋白H3赖氨酸9三甲基结合蛋白)的基因合作，以促进适应。这两个因子在EGL-4核进入的下游起作用，并且这两个因子在相同的适应遗传途径中起作用。因此，我们的研究提出了一种新颖而令人兴奋的可能性，即环境刺激可以通过小RNA来指导染色质的变化。我们建议通过确定小RNA和染色质是否在适应过程中起核心作用，它们在适应过程中如何发挥作用，以及它们是否可以调节候选靶标的转录来响应长期的气味暴露来检验这一假说。这项工作的意义在于，我们正在研究的那种大规模的表观遗传变化在成瘾和抑郁的模型中被发现(3，4)。在抑郁症模型中，这些变化发生在长时间的GPCR刺激的背景下。这些变化是如何发生的，目前尚不清楚。了解神经元刺激转化为染色质标记的途径的分子细节是理解这些疾病的关键。

DESCRIPTION (provided by applicant): Though a neuron must be able to respond reliably to stimulation, it is equally important that it alters its response as a function of specific experiences. The molecular and cellular basis for this plasticity and how stimulus-specific changes in plasticity occurs is important to understand as it underlies both normal processes such as learning and memory as well as the disease states of addiction and depression. Our goal is to use the olfactory response of the genetically tractable nematode C. elegans to determine whether a small RNA regulatory pathway directs stimulus-specific neuronal plasticity. One means by which repeated stimulation alters neuronal responsiveness is via the changes in transcription elicited by epigenetic "marks" such as DNA methylation and histone modification (reviewed in1). In S. pombe, plants and Drosophila, chromatin "marks" have been shown to be directed by small RNAs2. These epigenetic changes are thought to regulate important developmental processes. Whether small RNAs can dynamically regulate epigenetic changes in neurons as a consequence of specific behaviors has not been examined. An attractive but completely untested hypothesis is that small RNAs might provide the guidance and specificity for epigenetic events that direct long-lasting changes in neuronal activity. As a first step towards testing this hypothesis, we asked whether genes required for RNA-interference (RNAi) might be required for neuronal plasticity in the anatomically simple but genetically powerful model organism, C. elegans. C. elegans is inherently attracted to specific odors which it senses using G-protein coupled receptors (GPCRs), however, its attraction is dampened if the odors are not accompanied by food. We term this experience-dependent dampening of the response to odor olfactory adaptation. The key "switch" that turns on olfactory adaptation is the entry of the cGMP-dependent protein kinase (PKG), EGL-4, into the nucleus of the odor-stimulated sensory olfactory neuron AWC (Lee et al., submitted). In our preliminary studies, we found that a specific class of small RNAs work with a gene encoding a chromatin associated protein, HPL-2, (a histone H3 lysine 9 tri-methyl binding protein), within the sensory neuron at the time of odor-exposure to promote adaptation. Both factors act downstream of EGL-4 nuclear entry and both factors act in the same genetic pathway for adaptation. Thus, our studies have raised the novel and exciting possibility that environmental stimuli can act via small RNAs to direct changes in chromatin. We propose to test this hypothesis by determining whether small RNAs and chromatin are central players in the adaptation process, how they function in adaptation and whether they can regulate transcription of candidate targets in response to prolonged odor-exposure. The significance of this work is that large scale epigenetic changes of the sort we are studying are found in models for addiction and depression (3,4). In the models of depression, these changes occur in the context of prolonged GPCR stimulation. How these changes occur, is unknown. Understanding the molecular details of the pathways by which neuronal stimulation is translated into chromatin marks is key to understanding these diseases.