ALCOHOL METABOLISM GENES IN TRANSGENIC MICE

描述：肝脏中的乙醇代谢是由第I类物质调节的。 乙醇脱氢酶(ADH)和细胞色素P4502E1(CYP2E1)。第I类ADH 在小鼠体内是由ADH-1基因编码的，该基因在复杂的激素和 发育控制。转基因小鼠将被用来确定这种作用 基因内序列和5‘和3’侧翼序列的总控制性 表达表型。连接到ADH的扩展到-10KB的区域 迷你基因控制肾脏、肾上腺和其他组织的适当表达； 然而，肝脏表达还需要其他顺式作用元件。 包含所有基因内序列的Cosmid和lambda基因组克隆 现在隔离了不同的5‘和3’侧翼距离，并将使用 在转基因分析中鉴定肝脏所需的序列 表情。同样的转基因方法，使用替代启动子和 粘粒克隆将用于生产高表达ADH的转基因小鼠 和/或细胞色素P450-2。一种独特的分析方法将使RNA和蛋白质 从转基因产生的区别于生产与生产 内源基因。对于ADH-1，要使用的唯一基因变体将 能够在组织内的细胞类型水平上进行适当的表达 下定决心。过度表达的小鼠将被用来检验主要假说 关于酒精性肝损伤，包括自由基形成、脂质 过氧化和脂肪肝的产生。此外，一种基因变异 在近交系中确定的ADH-3的表达将在 分子水平，并将提供一种分化组织的机制 这种基因的表达。

DESCRIPTION: Ethanol metabolism in the liver is mediated by class I alcohol dehydrogenase (ADH) and cytochrome P4502E1 (CYP2E1). Class I ADH in mice is encoded by the Adh-1 gene which is under complex hormonal and developmental control. Transgenic mice will be used to determine the role of intragenic and 5'- and 3'-flanking sequences in control of the total expression phenotype. A region extending to -10kb attached tot an ADH minigene controls proper expression in kidney, adrenal, and other tissues; however, other cis-acting elements are required for liver expression. Cosmid and lambda genomic clones containing all intragenic sequences and different 5'- and 3'-flanking distance are now isolated and will be used in transgenic analysis to identify sequences necessary for liver expression. The same transgenic approach, using alternative promoters and cosmid clones, will be used to produce transgenic mice over expressing ADH and/or CYP2E1. A unique approach to analysis will allow RNA and protein produced from the transgene to be distinguished from production from the endogenous gene. For Adh-1, the unique genetic variant to be used will enable proper expression at the level of cell type within a tissue to be determined. Over expressing mice will be use to test major hypotheses about ethanol-induced liver injury including free radical formation, lipid peroxidation and production of fatty-liver. In addition, a genetic variant in Adh-3 expression identified among inbred strains will be defined at the molecular level and will provide a mechanism for differential tissue expression of this gene.