Satiety signaling in Caenorhabditis elegans

秀丽隐杆线虫的饱腹感信号传导

• 批准号: 8064024
• 负责人: Leon Avery
• 金额: $ 7.74万
• 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 2011-08-15
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8064024
• 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; 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中食欲控制的分子机制。就像在哺乳动物中一样，C秀丽隐杆菌的食欲受到饥饿信号的促进，并受到饱腹感信号的抑制。对食物可用性的两种反应在行为上都是相似的，并且在分子机制中具有类似的哺乳动物行为的重叠。基于这种保护，我们旨在在秀丽隐杆线虫中建立一个遗传模型系统，以研究食欲控制。我们预计分子机制将在根本上与哺乳动物的机制相似，但更简单，因此更容易解开。此外，我们希望蠕虫中可用的强大遗传工具将允许快速阐明新途径，随后可以测试与哺乳动物行为相关的新途径。 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。我们将检验以下假设：1。ASI感觉神经元活动信号营养健康，并引起饱腹感。 2。ASI的作用是由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。饱腹激素基因的表达会因营养健康而增加。 4。饱腹激素对下游神经元作用，以改变行为。 5。环状GMP（CGMP）激活ASI和下游神经元中的CGMP依赖性蛋白激酶（PKG）以产生饱腹感。目标1。确定饱腹感信号传导机制（假设1、4、5）。确定单个蠕虫中饱腹感的时间顺序。通过从神经元特异性启动子与激光显微外科手术结合的转基因表达胰岛素，TGF-2和CGMP途径中信号和受体基因在胰岛素，TGF-2和CGMP途径中的作用位点和时间（假设2、4和5）。通过上毒研究确定信号作用的顺序，以测量功能丧失突变体背景中的过表达和功能增益突变的影响。 AIM 2。找到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，可以找到转录受饥饿和重新喂养调节的肽基因，或者由CGMP和PKG调节。突变这些基因或击倒它们的表达并确定对饱腹部行为的影响。筛选突变体在饱腹感引起的静脉下有缺陷，并通过全基因组测序鉴定突变基因。IAM3。测量突触活动和从ASI中释放的肽（假设1，2）。开发方法通过在光漂白突触囊泡pH传感器突触氟luorin的光漂白后通过荧光恢复来测量突触活动。开发方法来测量神经元释放荧光肽的方法。使用这些方法来确定ASI活性和肽释放如何应对诱发饱腹感的疾病。 公共卫生相关性：肥胖是一个严重且越来越多的健康问题，它的饮食超出了满足人的代谢需求所需的更多。更好地了解调节食欲的信号可能会导致更好地控制食物摄入的方法，从而更好地控制肥胖症。