MOLECULAR GENETICS OF THERMOGENESIS

描述：（改编自申请人摘要）：研究者寻求 了解兼性和专性产热在 调节体重，并找到机制，将减少 通过增加产热而导致肥胖。 非颤抖 产热，棕色脂肪的主要生理功能，是基于 线粒体解偶联蛋白的棕色脂肪特异性表达 （UCP1），线粒体内膜中的质子载体， 氧化磷酸化以产生热量。 小鼠中的遗传操作 在转基因小鼠中增加UCP1可以减少肥胖。 在小鼠中， Ucp1基因已被基因打靶灭活，对 冷;然而，矛盾的是，他们不会发展肥胖。 结果 表明存在替代产热机制， UCP1的丢失 研究人员假设，一种新发现的 Ucp1，称为Ucp2，是一种强制性产热机制， 恢复Ucp1缺陷小鼠的能量平衡。 实验 提出了描述Ucp2在野生型小鼠中的表达， 对于Ucp1敲除，就其在 发育，组织之间，并作为喂养高脂肪/高 蔗糖日粮 为了确定UCP2分子是否产热， 将使用双替代技术构建转基因小鼠 在ES细胞中，将Ucp2 cDNA插入Ucp1基因。 这将 将Ucp2基因置于交感神经系统的控制之下， 将使我们能够确定UCP2分子是否可以保护小鼠 UCP1缺乏的人 第三个具体目标是 用Cre-loxP噬菌体系统克隆Ucp2基因， 重组 由于Ucp2是一种普遍表达的基因， 使一个基因失活将使我们能够在这些组织中抑制Ucp2， 所述Cre重组酶被选择性表达。 有了这个遗传系统 对UPC 2在组织能量平衡中的作用的明确评估可以 实现。 第四个具体目标是为世界各国提供一种新的模式。 用白喉毒素的细胞特异性表达消融棕色脂肪。 这种新的模型将通过基因替换技术产生， 白喉毒素基因插入Ucp1基因， 其表达的特异性和测试明确的假设， 褐色脂肪的消融导致食欲过盛。

DESCRIPTION: (Adapted from applicant's abstract): The investigators seek to understand the role of facultative and obligatory thermogenesis in the regulation of body weight and to find mechanisms that will reduce the development of obesity by increasing thermogenesis. Nonshivering thermogenesis, the major physiological function of brown fat, is based on the brown fat specific expression of the mitochondrial uncoupling protein (UCP1), a proton carrier in the inner mitochondrial membrane, that uncouples oxidative phosphorylation to produce heat. Genetic manipulations in mice which increase UCP1 in transgenic mice reduce obesity. Mice in which the Ucp1 gene has been inactivated by gene targeting are extremely sensitive to cold; however, paradoxically, they do not develop obesity. The result suggests that alternative thermogenic mechanisms exist that compensate for the loss of UCP1. The investigators have hypothesized that a newly discovered homologue of Ucp1, called Ucp2, is a mechanism for obligatory thermogenesis that can restore energy balance in mice with a defective Ucp1. Experiments are proposed to characterize the expression of Ucp2 in wild-type mice in homozygous for the Ucp1 knockout with respect to its changes during development, among tissues and as a consequence of feeding high fat/high sucrose diets. To establish whether the UCP2 molecule is thermogenic, a transgenic mouse will be constructed using the double replacement technique in ES cells that will insert the Ucp2 cDNA into the Ucp1 gene. This will bring the Ucp2 gene under the control of the sympathetic nervous system and will enable us to determine whether the UCP2 molecular can protect mice deficient in UCP1 from the cold exposure. A third specific aim will inactivate the Ucp2 gene with the Cre-loxP bacteriophage system of recombination. Since Ucp2 is a ubiquitously expressed gene, this system for inactivating a gene will enable us to inactivate Ucp2 in those tissues for which Cre recombinase is selectively expressed. With this genetic system the unambiguous evaluation of the role of UPC2 in tissue energy balance can be achieved. The fourth specific aim seeks to produce a new model for the ablation of brown fat with the cell specific expression of diphtheria toxin. This new model will be generated by gene replacement techniques to insert the diphtheria toxin gene into the Ucp1 gene in order to maximize the cell specificity of its expression and test unequivocally the hypothesis that the ablation of brown fat leads to hyperphagia.