Carbohydrate Regulation of Hepatic Gene Expression

描述（由申请人提供）：进化的压力有利于机制，允许身体在食物丰富时有效地将营养素储存为脂肪，以防止偶尔的饥荒。随着现代生活方式的巨大变化，包括高碳水化合物和高脂肪食物的消费，这些机制现在可能导致美国肥胖症的主要流行，其中大多数人口超重。葡萄糖不仅是所有哺乳动物组织的主要燃料，也是脂肪和蛋白质合成的碳源。肝脏是负责将过量的膳食碳水化合物转化为甘油三酯的主要器官。摄入高碳水化合物饮食诱导超过15个基因的转录，这些基因参与葡萄糖向储存脂肪的转化。本实验室首次鉴定了一种从大鼠肝脏中纯化的转录因子，该转录因子对脂肪生成所需的多个基因的启动子中发现的碳水化合物响应元件（ChRE）具有特异性，并显示适当的饮食响应调节，并被称为“碳水化合物响应元件结合蛋白，ChREBP”。ChREBP响应于过量碳水化合物而被激活以诱导脂肪生成酶基因的转录，然后被关闭的完整过程尚未完全理解。我们已经表明，葡萄糖和cAMP在脂肪生成的调节中具有相反的活性，部分通过ChREBP上多个位点的去磷酸化/磷酸化。葡萄糖通过激活Xu 5 P刺激的蛋白磷酸酶Xu 5 P-PP 2A来刺激这些位点中的至少一些的去磷酸化。我们最近发现，ChREBP与14-3-3的相互作用是调节ChREBP核定位的最重要步骤之一。ChREBP的磷酸化激活其与14-3-3的结合，并且是饥饿条件下ChREBP输出到细胞核外所必需的。此外，我们最近发现，肝脏中的特定代谢物促进ChREBP与14-3-3的相互作用。为了进一步表征代谢物，我们建议（1）分离纯形式的代谢物并确定结构，（2）确定其激活ChREBP和14-3-3之间相互作用的机制，（3）确定代谢物如何合成和降解，哪些酶负责这些反应以及它们如何被调节，（4）研究其代谢产物是否可能是营养和饥饿条件下调节ChREBP亚细胞定位的信号化合物;（5）利用X射线晶体学方法阐明Xu 5 P如何激活ChREBP和双功能酶Fru 6 P，2 kinase/Fru 2，6 bisphosphate。 公共卫生相关性：肥胖及其相关疾病，糖尿病，高血压，心脏病和一些癌症是美国目前面临的最严重的健康问题。将多余的碳水化合物有效地转化为脂肪以供长期储存，直到现代，这似乎是适应不确定的食物供应的进化优势，现在似乎对肥胖的发展起着重要作用。碳水化合物反应元件结合蛋白（ChREBP）是一种转录因子，当摄入碳水化合物时变得活跃，并增加脂肪酸合成所需基因的表达。本申请中提出的工作结果将阐明参与激活和终止ChREBP依赖性增加产生脂肪酸所需的酶的机制。这些知识将为制定预防和治疗肥胖症的策略提供有用的信息。

DESCRIPTION (provided by applicant): Evolutionary pressure has favored mechanisms that allow the body to efficiently store nutrients as fat when food is abundant as a safeguard against occasional famine. With the dramatic changes in modern lifestyle including consumption of high carbohydrate and high fat foods, these mechanisms may now be contributing to a major epidemic of obesity in the US where the majority of the population is overweight. Glucose is not only a major fuel of all mammalian tissues but also a source of carbon for fat and protein synthesis. The liver is the principal organ responsible for the conversion of excess dietary carbohydrate into triglycerides. Ingestion of a high carbohydrate diet induces transcription of more than 15 genes involve in the conversion of glucose to storage fat. A transcription factor, purified from rat liver, with specificity for carbohydrate responsive elements (ChRE) found in the promoters of multiple genes required for lipogenesis and which displayed appropriate dietary responsive regulation was first identified in this laboratory and termed "carbohydrate response element binding protein, ChREBP". The complete process by which ChREBP is activated in response to excess carbohydrate in order to induce the transcription of lipogenesis enzyme genes, and then is turned off, is not yet fully understood. We have shown that glucose and cAMP have opposing activities in the regulation of lipogenesis, in part through dephosphorylation/phosphorylation of multiple sites on ChREBP. Glucose stimulates dephosphorylation of at least some of these sites by activating a Xu5P-stimulated protein phosphatase, Xu5P-PP2A. We recently found that the interaction of ChREBP with 14-3-3 is one of the most important steps regulating the nuclear localization of ChREBP. Phosphorylation of ChREBP activates its binding to 14-3-3 and is essential for ChREBP export out of the nucleus under starvation conditions. In addition, we found more recently that a specific metabolite in liver promotes the interaction of ChREBP with 14-3-3. To characterize the metabolite further we propose to (1) isolate the metabolite in pure form and determine the structure, (2) determine the mechanism by which it activates the interaction between ChREBP and 14-3-3, (3) determine how the metabolite is synthesized and degraded, what enzymes are responsible for these reactions and how they are regulated, (4) investigate whether the metabolite might be a signaling compound for regulation of the subcellular localization of ChREBP in response to nutrients and starvation, and (5) elucidate how Xu5P activates ChREBP and the bifunctional enzyme, Fru6P, 2kinase/Fru2,6 bisPase, using X-ray crystallography. PUBLIC HEALTH RELEVANCE: Obesity and its associated diseases, diabetes, hypertension, heart disease and some cancers are among the most serious health problems now facing the US. The efficient conversion of excess carbohydrates to fat for long-term storage, which until modern times it would appear was an evolutionary advantage in adapting to uncertain food supplies, now appears to contribute significantly to development of obesity. Carbohydrate response element binding protein (ChREBP) is a transcription factor that becomes active when carbohydrates are eaten and increases the expression of genes required for synthesis of fatty acids. The results of the work proposed in this application will elucidate the mechanisms involved both in activation and termination of ChREBP dependent increases in enzymes needed to make fatty acids. This knowledge will provide information that may be useful in developing strategies for the prevention and treatment of obesity.