Determining the effects of genetic variation and early life stress on the regulation of the galanin gene in fat and alcohol selection.

英国67%的男性和57%的女性被归类为超重，过度沉迷于包括脂肪和酒精在内的高卡路里营养已经对英国人的健康产生了重大影响。由于肥胖与2型糖尿病和心血管疾病有关，我们需要找到这种卡路里过度消耗的原因，而英国人消耗的卡路里的主要来源是脂肪和酒精。除了心血管疾病和癌症，过量饮酒还与肝硬变和胰腺炎有关。由于DNA的差异，我们中的许多人都容易过度摄入脂肪和酒精，但也有证据表明，早期生活压力等环境因素也起到了作用。然而，我们仍然对影响我们脂肪和酒精摄入量的机制以及这些机制如何受到遗传变异和环境的影响知之甚少。重要的线索已经浮现。例如，大脑中控制脂肪和酒精摄入的下丘脑区域表达一种名为甘丙肽的小神经肽。去除编码甘丙素的基因可以减少脂肪和酒精的摄入量，减少体重增加。甘丙肽功能的一个关键方面是，它应该在正确的细胞中表达，以正确的数量和对正确的刺激做出反应。然而，对于控制下丘脑甘丙素产生的机制，或者控制这种表达的DNA区域的差异如何影响脂肪和酒精摄入量，我们一无所知。我们首先发现了一个DNA开关序列，它是控制下丘脑甘丙素的未知开关序列的极好候选者。然后我们发现，存在于人类群体中的这种开关内的序列变化与酗酒有关，增加了下丘脑这种开关的强度。我们还发现了开关内的表观遗传学变化，称为DNA甲基化，这些变化会因应激而改变，似乎也会改变开关的强度。我们假设，这些序列变化和GAL基因开关中的甲基化变化共同作用，改变了下丘脑中甘丙素的产生，并影响了脂肪和酒精的摄入。为了解决这一假设，我们使用了一种名为CRISPR基因组编辑的非凡新方法，快速有效地从基因组中删除了这个开关。我们将初步确定移除开关对下丘脑甘丙素产生的影响，并确定其对脂肪和酒精选择的影响。我们将再次使用CRISPR技术来复制开关内已知的人类序列变化(这一过程被称为人性化)，以模拟它们对甘丙素生产以及脂肪和酒精摄入的影响。然后，我们将使用这些人性化的模型来确定早期生活压力对开关序列甲基化的影响，以及这种甲基化如何影响其功能。使用这项革命性的CRISPR技术，我们将确定人类人口中由于遗传或环境而自然发生的变化如何影响我们的脂肪和酒精摄入量。考虑到肥胖和过量饮酒在我们社会中的巨大问题，这些研究不仅将为了解遗传和环境在过度脂肪和酒精摄入中的作用开辟道路，而且还将通过控制人类人口的过量卡路里摄入量为降低肥胖的易感性提供新的机会。

Overindulgence in high calorie nutrients, which include fat and alcohol, has had a major effect on the health of the UK population where 67% of males and 57% of females are classed as overweight. Because obesity is linked to type 2 diabetes and cardiovascular disease we need to find the causes of this calorie overconsumption and a major source of the calories consumed by the UK population come in the form of fat and alcohol. In addition to cardiovascular disease and cancer, excessive alcohol consumption is also linked to liver cirrhosis and pancreatitis. Many of us are susceptible to over consume fat and alcohol as a result of differences in our DNA but there is also evidence that environmental factors such as early life stress also play a role. However, we still know very little about the mechanisms that influence our intake of fat and alcohol and how these mechanisms are affected by genetic variation and environment. Important clues have emerged. For example, a small neuropeptide, called galanin is expressed in a region of the brain called the hypothalamus that controls fat and alcohol intake. Removal of the gene which encodes galanin reduces fat and alcohol intake and decreases weight gain. A critical aspect of the function of galanin is that it should be expressed in the correct cells, in the correct amount and in response to the correct stimuli. However, nothing was known about the mechanisms controlling the production of galanin in the hypothalamus or how differences in the regions of DNA that control this expression might affect fat and alcohol intake. We first discovered a DNA switch sequence that was an excellent candidate for the undiscovered switch sequence that controls galanin in the hypothalamus. We then found that sequence changes within this switch that exist within the human population, and were linked to alcohol abuse, increased the strength of this switch in the hypothalamus. We also found epigenetic changes within the switch, called DNA methylation, that were altered by stress and seemed to also change the strength of the switch. We hypothesise that these sequence changes and methylation changes within the GAL gene switch work together to alter the production of galanin in the hypothalamus and to influence fat and alcohol intake. To address this hypothesis we used a remarkable new method called CRISPr genome editing to quickly and efficiently delete the switch from the genome. We will initially determine the effects of removing the switch on galanin production in the hypothalamus and determine its effects on fat and alcohol selection. We will use CRISPr technology again to reproduce known human sequence changes within the switch (a process known as humanisation) to model their effects on galanin production and fat and alcohol intake. We will then use these "humanised" models to determine the effects of early life stress on the methylation of the switch sequence and how this methylation affects its function. Using this revolutionary CRISPr technology we will determine how changes that occur naturally in the human population, as a result of genetics or environment, affect our fat and alcohol intake. Considering the huge problem of obesity and excessive alcohol intake in our society these studies will open avenues for not only understanding the role of genetics and environment in excess fat and alcohol intake but will also provide novel opportunities for reducing susceptibility to obesity by controlling excess calorie intake in the human population.