Genetic Analysis of Feeding Behavior and Fat Deposition

控制摄食和脂肪沉积的重要机制在 果蝇、黑腹果蝇和人类。这验证了果蝇作为遗传模型系统用于识别 以及表征调节新陈代谢过程的新基因。在初步研究中，我们有 证明5-羟色胺(5-羟色胺，5-羟色胺)是苍蝇取食行为的有效抑制因子 通过激活一个或多个5-羟色胺受体。这些发现提供了与哺乳动物的直接相似之处， 现已证实，5HT2C受体是摄食行为的重要介体，并且是一种 公认的肥胖症治疗靶点。本应用程序的目标1建议进一步探索 5-羟色胺能控制果蝇摄食的分子机制 现有的突变体和RNA干扰果蝇，目前正在我们实验室产生。这些 菌株将提供工具来研究不同的5羟色胺受体亚型以及它们的 组织特异性表达(如CMS和肠道)是5-羟色胺能控制摄食的基础。高度的 我们已经建立的用于评估5-羟色胺介导的食物摄入量的敏感食物摄入量测试也已经被 用于开始前向基因筛查(目标2)。到目前为止，1315个突变果蝇品系，每个都携带 已对单个精确定位的插入物进行了摄食异常筛查。最多的12个 已选定有前途的候选人进行详细分析(目标2 A)。为了探索， 候选基因(及其相应的哺乳动物同源基因)在调节摄食和/或脂肪沉积方面，a 将结合使用各种方法，包括生化方法(如脂肪酸分析)、组织学方法(如： (例如，脂肪组织的扫描电子显微镜)和遗传(例如，转基因拯救)方法。基于 我们初步成功地确定了候选基因，我们建议继续进行正向基因筛查 (目标2B)。这一后续工作的目标是确定(I)在基本条件下食物摄入量的修饰物 (2)5-羟色胺存在时的食物摄入量(以确定5-羟色胺和交叉性摄食的修饰因素 途径)和(Iii)脂肪沉积。为了提高发现表型和识别基因的可能性 与人类的生理相关性，我们将利用生物信息学工具预先选择插入菌株 放映。选择标准将包括(I)转录单位的中断，和(Ii)存在 与目标苍蝇基因相对应的哺乳动物同源基因。到目前为止，-2,500个插入轴承菌株 符合这些标准的已被确定。将对这些菌株中的每一种进行摄食变化的评估。 行为和脂肪沉积。异常值的特征将如上所述。我们预计， 拟议的研究将确定与人类肥胖的病理生理学相关的新基因。

Important mechanisms underlying the control of feeding and fat deposition have been conserved between Drosophila melanogaster and human. This validates the fruit fly as a genetic model system for identifying and characterizing novel genes that mediate metabolic processes. In Preliminary Studies, we have demonstrated that serotonin (5-hydroxytryptamine, 5HT) acts as a potent inhibitor of fly feeding behavior through activation of one or more 5HT receptors. These findings provide a direct parallel with mammals, for which it has been established that the 5HT2C receptor is an important mediator of feeding behavior and a well recognized target for the treatment of obesity. Aim 1 of this application proposes to further explore the molecular mechanisms underlying the serotonergic control of feeding in Drosophila, utilizing a combination of existing mutants and RNA interference flies which are currently being generated in our laboratory. These strains will provide tools to investigate the extent to which different 5HT receptor subtypes as well as their tissue specific expression (e.g. CMS vs. intestine) underlie the serotonergic control of feeding. The highly sensitive food intake assay that we have established for assessing 5HT mediated food intake has also been utilized to begin a forward genetic screen (Aim 2). To date, 1315 strains of mutant Drosophila, each carrying a single precisely mapped insertion, have been screened for abnormalities in feeding. Twelve of the most promising candidates have been selected for detailed analysis (Aim 2A). To explore the role of the , candidate gene (and its corresponding mammalian homolog) in modulating feeding and/or fat deposition, a combination of approaches will be used, including biochemical (e.g. fatty acid profiling), histologic (e.g. scanning electron microscopy of fat tissue), and genetic (e.g. transgenic rescue) methodologies. Based on our initial success in identifying candidate genes, we propose to continue the forward genetic screen (Aim 2B). The goal of this subsequent effort is to define modifiers of (i) food intake under basal conditions (ii) food intake in the presence of 5HT (to identify modifiers of feeding within the 5HT and intersecting pathways) and (iii) fat deposition. To enhance the likelihood of finding a phenotype and of identifying a gene with physiologic relevance in humans, we will utilize bioinformatic tools to pre-select insertion strains for screening. The selection criteria will include (i) disruption of the transcription unit, and (ii) existence of a mammalian homolog corresponding to the targeted fly gene. To date, -2,500 insertion bearing strains which meet these criteria have been identified. Each of these strains will be assessed for alterations in feeding behavior and fat deposition. Outliers will be characterized as described above. We anticipate that the proposed studies will identify novel genes relevant to the pathophysiology of human obesity.