Genetic dissection of neuropeptide cell biology at high and low activity in a defined sensory neuron.
Genetic dissection of neuropeptide cell biology at high and low activity in a defined sensory neuron.
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DOI：
10.1073/pnas.1714610115
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发表时间：
2018-07-17
影响因子：
11.1
通讯作者：
de Bono M
中科院分区：
文献类型：
--
作者：
Laurent P;Ch'ng Q;Jospin M;Chen C;Lorenzo R;de Bono M
神经肽是普遍存在的行为和生理调节剂。它们被包装在称为致密核心囊泡（DCV）的特殊分泌细胞器中，在神经刺激时释放。尽管突触小泡的局部回收已被深入研究，但关于 DCV 蛋白回收的研究却很少。我们建立了一个范式来研究我们可以控制其活动的神经元中的 DCV。我们通过证实之前对 DCV 细胞生物学的许多观察结果来验证我们的模型。我们鉴定了一组参与 DCV 蛋白质回收的基因。我们还发现证据表明 DCV 启动和胞吐作用的不同机制可能在高和低神经活动下发挥作用。神经肽是普遍存在的行为和生理调节剂。它们被包装在称为致密核心囊泡（DCV）的特殊分泌细胞器中，在神经刺激时释放。与可以在释放位点附近回收和重新填充的突触小泡不同，DCV 必须通过细胞体内的从头合成来补充。在这里，我们剖析了秀丽隐杆线虫感觉神经元体内的 DCV 细胞生物学，我们可以使用自然刺激来控制其强直活动。我们在神经元中表达荧光标记的神经肽，并定义描述其亚细胞分布的参数。我们测量了 187 个突变体在高和低神经活动时的这些参数，这些突变体在涉及膜运输、神经内分泌分泌和神经元或突触活动的蛋白质方面存在缺陷。使用无监督的层次聚类方法，我们分析这些数据并识别出 62 组具有相似突变表型的基因。我们探索这些组的子集的功能。我们概括了许多以前的发现，验证了我们的范式。我们发现了大量参与 DCV 膜蛋白回收的蛋白质，但迄今为止对此的研究还很少。我们表明，未折叠的蛋白质反应促进 DCV 的产生，这可能有助于压力的组织间通讯。我们还发现证据表明不同的启动和胞吐作用机制可能在高和低神经活动下发挥作用。我们的工作为研究不同神经活动水平的 DCV 生物学提供了一个明确的框架。
Neuropeptides are ubiquitous modulators of behavior and physiology. They are packaged in specialized secretory organelles called dense core vesicles (DCVs) that are released upon neural stimulation. Whereas local recycling of synaptic vesicles has been investigated intensively, there are few studies on recycling of DCV proteins. We set up a paradigm to study DCVs in a neuron whose activity we can control. We validate our model by confirming many previous observations on DCV cell biology. We identify a set of genes involved in recycling of DCV proteins. We also find evidence that different mechanisms of DCV priming and exocytosis may operate at high and low neural activity. Neuropeptides are ubiquitous modulators of behavior and physiology. They are packaged in specialized secretory organelles called dense core vesicles (DCVs) that are released upon neural stimulation. Unlike synaptic vesicles, which can be recycled and refilled close to release sites, DCVs must be replenished by de novo synthesis in the cell body. Here, we dissect DCV cell biology in vivo in a Caenorhabditis elegans sensory neuron whose tonic activity we can control using a natural stimulus. We express fluorescently tagged neuropeptides in the neuron and define parameters that describe their subcellular distribution. We measure these parameters at high and low neural activity in 187 mutants defective in proteins implicated in membrane traffic, neuroendocrine secretion, and neuronal or synaptic activity. Using unsupervised hierarchical clustering methods, we analyze these data and identify 62 groups of genes with similar mutant phenotypes. We explore the function of a subset of these groups. We recapitulate many previous findings, validating our paradigm. We uncover a large battery of proteins involved in recycling DCV membrane proteins, something hitherto poorly explored. We show that the unfolded protein response promotes DCV production, which may contribute to intertissue communication of stress. We also find evidence that different mechanisms of priming and exocytosis may operate at high and low neural activity. Our work provides a defined framework to study DCV biology at different neural activity levels.
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