The Agouti Related Protein and its Role in Human Obesity

描述（由申请人提供）：Agouti相关蛋白（AGRP）在肥胖和糖尿病小鼠中上调，并在转基因小鼠中过度表达时刺激贪食和肥胖的发生。人类同源hAGRP已被分离出来，具有相似的分子和生理特性，在弓形核和外周分别表达一个转录本。我们最近确定了基因结构，并确定了人类AGRP基因的最小启动子。转录因子的多个结合位点被确定，包括可能介导瘦素在下丘脑中的作用的STAT反激活因子的推定结合位点。5'非编码外显子仅在外周细胞系中具有显著的启动子活性，提示其在外周特异性转录物的表达中起作用。我们还发现了启动子的多态性，该多态性对启动子活性和结合转录因子的亲和力有显著影响，在下丘脑和外周来源的细胞系中进行了测试。在两个不同的队列中，具有高启动子活性的基因型与肥胖和2型糖尿病（T2DM）显著相关。此外，该基因编码区域的多态性与低体重指数和低腹部脂肪显著相关，但仅在老年人中存在，这表明hAGRP在预防晚发型肥胖方面发挥作用。我们假设，下丘脑和外周hAGRP数量的增加，由其启动子决定，将导致贪食和肥胖的发展。因此，我们建议完全表征hAGRP的启动子，并确定调节该基因表达的转录因子和序列基序（Specific Aim 1）。我们还将确定激素/燃料底物对AGRP启动子活性和内源性表达的影响，并确定该基因启动子中的激素/底物反应元件（Specific Aim 2）。此外，我们将研究hAGRP多态性与肥胖和2型糖尿病的关系，而AGRP的全身水平将与启动子多态性以及激素/燃料底物的全身水平相关（Specific Aim 3）。鉴于hAGRP是一种有效的食欲效应因子，我们有必要确定其表达的调控机制，并确定其在食物摄入和人类肥胖发展中的作用。

DESCRIPTION (provided by applicant): The Agouti Related Protein (AGRP) is upregulated in obese and diabetic mice and stimulates hyperphagia and the development of obesity when overexpressed in transgenic mice. The human ortholog, hAGRP, has been isolated and has similar molecular and physiological properties, expressing one transcript in the arcuate nucleus and one in the periphery. We recently determined the gene structure and identified the minimal promoter of the human AGRP gene. Multiple binding sites for transcription factors were identified including putative binding sites for the STAT transactivators that may potentially mediate leptin's action in the hypothalamus. The 5' non-coding exon had significant promoter activity in a periphery cell line only, suggesting its role in the expression of the periphery-specific transcript. We also identified a polymorphism in the promoter that had significant impact on promoter activity and affinity to bind transcription factors, as tested in hypothalamus- and periphery-derived cell lines. The genotype with the high promoter activity was significantly associated with obesity and Type 2 Diabetes Mellitus (T2DM) in two different cohorts. Moreover, a polymorphism in the coding region of the gene was significantly associated with low body mass index and low abdominal adiposity but in older people only, suggesting a role for hAGRP in preventing late on set obesity. We hypothesize that increased amounts of hAGRP in the hypothalamus and the periphery, as determined by its promoter, will result in hyperphagia and the development of obesity. We therefore propose to completely characterize the promoter of hAGRP and identify the transcription factors and sequence motifs that regulate expression of the gene (Specific Aim 1). We will also determine the impact of hormones/fuel substrates on promoter activity and endogenous expression of AGRP, and identify the hormone/substrate response elements in the promoter of the gene (Specific Aim 2). Furthermore, we will investigate the association of hAGRP polymorphisms with obesity and T2DM, while the systemic levels of AGRP will be correlated with promoter polymorphisms as well as with the systemic levels of hormones/fuel substrates (Specific Aim 3). Given that hAGRP is a potent appetite effector, it is important that we identify the regulatory mechanisms for its expression and determine its role in food intake and the development of human obesity.