Satiety signaling in Caenorhabditis elegans

秀丽隐杆线虫的饱腹感信号传导

• 批准号: 7778044
• 负责人: Leon Avery
• 金额: $ 30.12万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7778044
• 关键词: Afferent Neurons; Animals; Anorexia; Behavior; Biological Models; Biology; Caenorhabditis elegans; Complex; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Desire for food; Disease; Eating; Environment; Epidemic; Fluorescence Recovery After Photobleaching; Food; Genes; Genetic; Genetic Epistasis; Genetic Models; Genetic Transcription; Genome; Goals; Health; Hormones; Human; Hunger; Insulin; Lasers; Lead; MAP Kinase Signaling Pathways; Mammals; Measures; Mediating; Metabolic; Methods; Microsurgery; Molecular; Motion; Muscarinic Acetylcholine Receptor; Mutate; Nematoda; Neurons; Nutritional; Obesity; Organism; Pathway interactions; Peptides; Personal Satisfaction; Persons; Proteins; Receptor Gene; Reverse Transcriptase Polymerase Chain Reaction; Satiation; Satiety Response; Signal Transduction; Site; Sleep; Source; Starvation; Synapses; Synaptic Vesicles; Testing; Time; base; behavior change; feeding; gain of function mutation; genome sequencing; knock-down; loss of function; mutant; novel; overexpression; promoter; public health relevance; response; sensor; tool; transgene expression

DESCRIPTION (provided by applicant): We propose to investigate the molecular mechanisms of appetite control in the nematode Caenorhabditis elegans. Just as in mammals, C elegans appetite is promoted by hunger signals and suppressed by satiety signals. Both responses to food availability are similar behaviorally and overlap in molecular mechanism with analogous mammalian behaviors. Based on this conservation, we aim to establish in C elegans a genetic model system to study appetite control. We anticipate that molecular mechanisms will be fundamentally similar to those of mammals, but simpler and therefore easier to unravel. Furthermore, the powerful genetic tools available in the worm will, we hope, allow rapid elucidation of new pathways, whose relevance to mammalian behavior can subsequently be tested. Our broad long-term objective is to understand at a molecular level how an animal regulates its food intake. Our goal in the next five years is to investigate the cellular and molecular mechanisms that control satiety. We will test the following hypotheses: 1. ASI sensory neuron activity signals nutritional well-being and provokes a satiety response. 2. ASI's effects are mediated by the release of peptide and protein hormones, including insulin-like peptides and the TGF-2-like peptide DAF-7. 3. Expression of satiety hormone genes increases in response to nutritional well-being. 4. Satiety hormones act on downstream neurons to change behavior. 5. Cyclic GMP (cGMP) activates cGMP-dependent protein kinase (PKG) in ASI and in downstream neurons to produce satiety. Aim 1. Identify satiety signaling mechanisms (hypotheses 1, 4, 5). Determine the time-course of satiety behavior in single worms. Determine the sites and times of action of signal and receptor genes in the insulin, TGF-2, and cGMP pathways by expression of transgenes from neuron-specific promoters in combination with laser microsurgery (hypotheses 2, 4, and 5). Determine the order of signal action by epistasis studies measuring the effects of overexpression and gain-of-function mutations in loss-of-function mutant backgrounds. Aim 2. Find peptide and protein hormone genes whose transcription correlates with satiety (hypotheses 2, 3). Using microarrays and quantitative RT-PCR, find peptide genes whose transcription is regulated by starvation and refeeding, or by cGMP and PKG. Mutate these genes or knock down their expression and determine the effect on satiety behavior. Screen for mutants defective in satiety- induced quiescence and identify the mutated genes by whole-genome sequencing.Aim 3. Measure synaptic activity and release of peptides from ASI (hypotheses 1, 2). Develop methods to measure synaptic activity by recovery of fluorescence after photobleaching of the synaptic vesicle pH sensor synaptopHluorin. Develop methods to measure release of fluorescent peptides from neurons. Use these methods to determine how ASI activity and peptide release respond to conditions that induce satiety. PUBLIC HEALTH RELEVANCE: Obesity is a serious and growing health problem, brought on by eating more than is necessary to fulfill a person's metabolic needs. A better understanding of the signals that regulate appetite may lead to better methods for controlling food intake, and therefore to better control of obesity.

描述（由申请人提供）：我们建议研究线虫秀丽隐杆线虫食欲控制的分子机制。就像哺乳动物一样，线虫的食欲受到饥饿信号的促进，并受到饱足信号的抑制。对食物供应的两种反应在行为上相似，并且在分子机制上与类似的哺乳动物行为重叠。基于这种保守性，我们的目标是在秀丽隐杆线虫中建立一个遗传模型系统来研究食欲控制。我们预计分子机制将与哺乳动物的分子机制基本相似，但更简单，因此更容易解开。此外，我们希望，线虫中可用的强大遗传工具将能够快速阐明新的途径，随后可以测试其与哺乳动物行为的相关性。 我们广泛的长期目标是在分子水平上了解动物如何调节其食物摄入量。我们未来五年的目标是研究控制饱腹感的细胞和分子机制。我们将测试以下假设： 1. ASI 感觉神经元活动发出营养健康信号并引发饱腹感反应。 2. ASI 的作用是通过肽和蛋白质激素的释放介导的，包括胰岛素样肽和 TGF-2 样肽 DAF-7。 3. 饱腹感激素基因的表达随着营养状况的改善而增加。 4. 饱腹感激素作用于下游神经元以改变行为。 5. 环鸟苷酸 (cGMP) 激活 ASI 和下游神经元中的环鸟苷酸 (cGMP) 依赖性蛋白激酶 (PKG) 以产生饱腹感。目标 1. 确定饱腹感信号机制（假设 1、4、5）。确定单个蠕虫饱足行为的时间过程。通过神经元特异性启动子的转基因表达结合激光显微手术，确定胰岛素、TGF-2 和 cGMP 途径中信号和受体基因的作用位点和时间（假设 2、4 和 5）。通过测量功能丧失突变体背景中过度表达和功能获得突变的影响的上位研究来确定信号作用的顺序。目标 2. 找到其转录与饱腹感相关的肽和蛋白质激素基因（假设 2、3）。使用微阵列和定量 RT-PCR，找到其转录受饥饿和再喂食或 cGMP 和 PKG 调节的肽基因。突变这些基因或敲低它们的表达，并确定对饱腹感行为的影响。筛选在饱腹感诱导的静止中存在缺陷的突变体，并通过全基因组测序鉴定突变基因。目标 3. 测量突触活性和 ASI 肽的释放（假设 1、2）。开发通过突触囊泡 pH 传感器 synaptopHluorin 光漂白后恢复荧光来测量突触活性的方法。开发测量神经元荧光肽释放的方法。使用这些方法来确定 ASI 活性和肽释放如何响应引起饱腹感的条件。 公众健康相关性：肥胖是一个严重且日益严重的健康问题，是由于吃得超过满足一个人的新陈代谢需求所必需的量而引起的。更好地了解调节食欲的信号可能会带来更好的控制食物摄入的方法，从而更好地控制肥胖。