Identifying New Regulators of Leptin-Like Signaling in Drosophila Brain Neurons

鉴定果蝇脑神经元中瘦素样信号传导的新调节因子

基本信息

批准号：
8653630
负责人：
KAI G ZINN
金额：
$ 20.6万
依托单位：
CALIFORNIA INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-05-01 至 2015-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8653630
关键词：
Adipocytes Affect Anti-Obesity Agents Biological Models Body Weight Brain Candidate Disease Gene Cells Chronic Disease Cultured Cells Disease Down-Regulation Drosophila genus Drug Targeting Eating Elements Energy Metabolism Fatty acid glycerol esters Gene Expression Generic Drugs Genes Genetic Genetic Epistasis Genetic Screening Goals Hormones Human Hypothalamic structure Incidence Individual Insertion Mutation Leptin Leptin resistance Mammals Maps Metabolism Molecular Neurons Obesity Orthologous Gene Outcomes Research Pathway interactions Pattern Proteins Public Health Publishing RNA Interference Receptor Signaling Relative (related person)Reporting Research Resistance Signal Pathway Signal Transduction Site System Therapeutic Transgenic Organisms Triglycerides Work base drug development energy balance feeding fly genetic regulatory protein in vivo insight knock-down leptin receptor lipid metabolism loss of function mutation neuromechanism obesity management public health relevance receptor receptor coupling research study screening

项目摘要

DESCRIPTION (provided by applicant): Leptin is a hormone that controls fat storage and energy expenditure. Leptin is made by adipocytes and acts on hypothalamic neurons that regulate energy balance. High leptin levels are an indicator of sufficient energy stores and should cause cessation of eating. However, most obese humans are leptin resistant. Leptin interacts with a JAK/STAT pathway-coupled receptor, and leptin resistance can be caused by downregulation of the pathway downstream of the receptor. If additional regulators of leptin receptor signaling in hypothalamic neurons could be identified, manipulation of their activities might provide ways to overcome leptin resistance. This proposal describes a new way to find such regulators using Drosophila genetics. Drosophila has dedicated adipocytes, and the activities of specific sets of brain neurons control fat storage and metabolism. The preliminary results reported here show that Drosophila fat content is regulated by a leptin-like JAK/STAT signaling pathway that acts in fat-regulating 'Fru' neurons. These findings show that flies and mammals use some similar genetic and neural mechanisms for control of fat storage. Drosophila is a model system that is amenable to fast and inexpensive forward genetic screening, and our results suggest that conserved JAK/STAT regulators identified in Drosophila would have relevance for research in mammalian systems. The objective of the first specific aim is to find proteins that affect fat content by regulating JAK/STAT signaling in Fru neurons. These will be identified by screening a set of ~200 genes identified as modulators of the JAK/STAT pathway in cultured cells, about 75% of which have human orthologs or relatives. Expression of each gene will be knocked down in Fru neurons using transgenic RNAi, and those genes for which knockdown affects triglyceride levels will be selected for further study. The identified candidate genes will be prioritized based on sequence relationships with human genes, effect size and direction, and availability of viable loss-of- function mutations and inserted elements that can be used for tagging. The objective of the second specific aim is to place the highest-priority subset of the regulators identified in Specific Aim 1 into their molecular and cellular contexts. Analysis of genetic epistasis will be used to determine whether the regulators act upstream or downstream of the receptor, and if they control the firing of Fru neurons. Regulators will be fluorescently tagged in vivo to reveal their cellular and subcellular expression patterns. The same tagging strategy will be used to create 'driver' lines that will confer gene expression in neurons that normally express the regulator, allowing manipulation of their activities. The expected outcome of the research proposed in specific aims 1 and 2 is the definition of a set of conserved modulators of JAK/STAT signaling that function downstream of the receptor in fat-regulating neurons. Negative regulators identified in these experiments might be potential drug targets whose inhibition could upregulate anorexigenic leptin signaling in leptin-resistant obese individuals.

描述（由申请人提供）：瘦素是控制脂肪储存和能量消耗的激素。瘦素是由脂肪细胞制成的，并作用于调节能量平衡的下丘脑神经元。高瘦素水平是足够的能量储存的指标，应导致饮食停止。但是，大多数肥胖的人是耐瘦素的。瘦素与JAK/STAT途径偶联受体相互作用，瘦素耐药性可能是由于受体下游的途径下调引起的。如果可以确定下丘脑神经元中瘦素受体信号传导的其他调节剂，则对其活性的操纵可能会提供克服瘦素耐药性的方法。该提案描述了一种使用果蝇遗传学找到此类调节剂的新方法。果蝇具有专用的脂肪细胞，并且特定的脑神经元组的活性控制脂肪储存和代谢。此处报告的初步结果表明，果蝇脂肪含量受瘦素样JAK/Stat信号传导途径的调节，该途径作用于脂肪调节的“ FRU”神经元。这些发现表明，苍蝇和哺乳动物使用一些类似的遗传和神经机制来控制脂肪储存。果蝇是一种模型系统，可与快速且廉价的远期遗传筛查相应，我们的结果表明，果蝇中鉴定出的保守的JAK/Stat调节剂与哺乳动物系统的研究具有相关性。第一个特定目的的目的是通过调节FRU神经元中的JAK/STAT信号来找到影响脂肪含量的蛋白质。这些将通过筛选一组200个基因在培养细胞中识别为JAK/Stat途径的调节剂的一组基因，其中约75％具有人类直系同源物或亲戚。使用转基因RNAi将在FRU神经元中击倒每个基因的表达，而敲低影响甘油三酸酯水平的基因将被选择进行进一步研究。确定的候选基因将根据与人类基因，效应大小和方向的序列关系以及可行的功能丧失突变和可用于标记的插入元素的可用性进行优先级。第二个特定目的的目的是将特定目标1中确定的调节剂的最高优先级子集放入其分子和细胞环境中。分析遗传性上学将用于确定受体上游或下游的调节剂以及是否控制FRU神经元的发射。调节器将在体内荧光标记，以揭示其细胞和亚细胞表达模式。相同的标记策略将用于创建“驱动程序”线，以赋予基因表达通常表达调节剂的神经元可以操纵其活动。特定目的1和2中提出的研究的预期结果是对脂肪调节神经元下游功能的JAK/STAT信号的一组保守调节剂的定义。在这些实验中鉴定出的负调节剂可能是潜在的药物靶标，其抑制作用可以上调抗瘦素耐药肥胖个体中的厌食症瘦素信号传导。