NUTRITIONAL AND HORMONAL REGULATION OF HEPATIC GENES

肝脏是人体处理摄入的碳水化合物的主要器官。 哺乳动物，因此，t负责将过量的饮食转化为 碳水化合物转化为甘油三酯。碳水化合物摄入量增加会导致肥胖 增加肝脏脂肪生成，这一过程在一定程度上由 参与形成的酶的产生的变化 甘油三酯。目前人们对这种机制知之甚少。 哪些营养因素控制哺乳动物系统中的基因表达 而这项研究的长期目标就是探索这条途径。 为此，我们重点研究两个肝脏基因的表达。 -丙酮酸激酶和S14基因。两个基因的转录速度 这些基因中的一部分对碳水化合物的摄取反应迅速而显著。 这些变化可以在培养的原代肝细胞中模拟 使我们能够识别对基因调控至关重要的DNA序列 通过碳水化合物来表达。对于S14和丙酮酸激酶，DNA 来自5‘-侧翼区的序列被证明能够 将葡萄糖激活授予一个相连的报告基因。我们假设 一种常见的碳水化合物反应转录因子结合到 位于这些基因的5‘侧翼区。或目标是确定 和表征顺式作用的dna序列和反式作用因子(S) 负责控制这一过程。这将通过以下方式实现 (1)利用诱变技术，获得了 丙酮酸激酶和S14基因的碳水化合物反应将是 在转基因肝细胞和转基因小鼠中定义；(2)鉴定 与之相互作用的肝糖反应因子(S) DNA序列；以及(3)碳水化合物的纯化和克隆- 来自肝脏的反应因子。这一克隆将提供一个关键工具 用于进一步研究，以探索信号通路，通过它 肝细胞可以“感觉”糖酵解的增加并通过改变 特定的基因表达。在现代人中，有现成的 饮食中的单糖，这一途径及其控制可能是 导致甘油三酯过度转化的因素和问题 与肥胖有关。 S14基因的表达也受甲状腺的调节 荷尔蒙。两种刺激物协同调控S14基因表达 举止。最近来自实验室的证据表明， 在T3受体和碳水化合物途径的组成部分之间。 使用先前资助中获得的T3受体的CDNA克隆 期间，我们将测试受体相互作用的各种模型 以及碳水化合物反应因子。这些研究应该提供 洞察两种不同刺激作用的机制 协调控制基因表达。

The liver is the primary organ dealing with ingested carbohydrates in mammals and, as such, t is responsible for converting excess dietary carbohydrates to triglycerides. Increased intake of carbohydrate leads t increased hepatic lipogenesis and this process is regulated in part by changes in the production of enzymes involved in the formation of triglycerides. Little is currently known regarding the mechanisms by which nutritional factors control gene expression in mammalian systems and the long range goal of this research is to explore this pathway. For this purpose, we are focusing on the expression of two hepatic genes - pyruvate kinase and the S14 gene. The rates of transcription of both of these genes respond rapidly and markedly to carbohydrate feeding. These changes can be mimicked in cultured primary hepatocytes which allows us to identify DNA sequences critical for the regulation of gene expression by carbohydrate. For both S14 and pyruvate kinase, DNA sequences from the 5'-flanking region were shown to be capable of conferring glucose- activation to a linked reporter gene. We hypothesize that a common carbohydrate-responsive transcription factor is binding to sites in the 5'-flanking region of these genes. Or goal is to identify and characterize the cis-acting DNA sequences and trans-acting factor(s) responsible for controlling this process. This will be accomplished by (1) using mutagenesis techniques, essential DNA sequences for the carbohydrate response of the pyruvate kinase and S14 genes will be defined in transfected hepatocytes and transgenic mice; (2) identifying the hepatic carbohydrate-responsive factor(s) which interacts with these DNA sequences; and (3) purifying and cloning of the carbohydrate- responsive factor from liver. This clone will provide a critical tool for further studies to explore the signalling pathway by which the hepatocyte can 'sense' increased glycolysis and respond by changing specific gene expression. In modern man with the ready availability of simple sugars in the diet, this pathway and its control could be a factor leading to excess conversion to triglycerides and problems associated with obesity. The expression of the S14 gene is also regulated in response to thyroid hormone. The two stimuli control S14 gene expression in a synergistic manner. Recent evidence from the laboratory suggests an interaction between the T3 receptor and components of the carbohydrate pathway. Using CDNA clones to the T3 receptor obtained in the previous funding period, we will test various models for the interaction of the receptor and the carbohydrate-responsive factor. These studies should provide insight into the mechanism by which two distinct stimuli can act to coordinately control gene expression.