METABOLIC HEPATIC FLUX OF ISOPRENOIDS

描述(改编自申请者摘要)：本报告的主要目的 建议研究胆固醇中的第一种特异酶 生物合成途径，角鲨烯合成酶，以了解 其调控机制。这个目标现在是可以实现的，因为在 该项目的头三年，研究人员已经获得了 大鼠肝酶和人肝酶的克隆以及 人类基因5‘侧翼序列的基因组克隆。基于 在本授权期内完成的工作，现申请续期5年 其中调查人员提议调查潜在的机制 参与了类固醇介导的该基因的转录调控。 研究人员已经确定了一个69个碱基对的启动子序列 负责这项规定，并建议调查和澄清 本条例中涉及的顺式作用元件的结构。 进一步提出了研究细胞反式作用蛋白的建议。 参与该基因调控的因子并进行比较 其中，监管机制对其他高度监管 胆固醇生物合成途径中的调节酶。这将是 通过分离和克隆这些蛋白因子和一种 它们与角鲨烯编码基因相互作用的详细研究 合成酶。根据本授权期内取得的成果， 调查人员建议调查这种病毒存在的意义 不同大小类角鲨烯合成酶mRNA亚型的调控 在不同的胆固醇生成条件下这种肝酶的活性。 最后，调查人员建议核实人类的规则 角鲨烯合成酶基因在转基因小鼠中的表达。我们也 建议研究这种酶的过度生产的含义，在 转基因小鼠对甾醇和非甾醇整体代谢的影响 类异戊二烯。这些研究的圆满完成 资助申请应加强对分子基础的理解 肝脏胆固醇的产生。

DESCRIPTION (Adapted from applicant's Abstract): The main objective of this proposal is to study the first specific enzyme in the cholesterol biosynthetic pathway, squalene synthase, in order to understand the mechanisms of its regulation. This objective is now obtainable since in the first three years of the project the investigators have obtained the cDNA clones of both the rat hepatic and the human hepatic enzymes as well as genomic clones of the 5' flanking sequences of the human gene. Based on work completed in the current grant period we now apply for a 5-year renewal in which the investigators propose to investigate the underlying mechanisms involved in the sterol-mediated transcriptional regulation of this gene. The investigators have identified a 69 bp promoter sequence which is responsible for this regulation and propose to investigate and elucidate the structure of the cis-acting elements which are involved in this regulation. It is further proposed to investigate the cellular trans-acting protein factors that are involved in the regulation of this gene and to compare them, and the regulatory mechanism to the regulation of other highly regulated enzymes in the cholesterol biosynthetic pathway. This will be achieved by the isolation and cloning of these protein factors and a detailed study of their interaction with the gene encoding squalene synthase. Based on results obtained in the current grant period the investigators propose to investigate the significance of the existence of different size-classes isoforms of squalene synthase mRNA in the regulation of this hepatic enzyme activity under varying cholesterogenic conditions. Finally, the investigators propose to verify the regulation of the human squalene synthase gene by its expression in transgenic mice. We also propose to study the implication of over production of this enzyme, in transgenic mice, on the overall metabolism of sterol and non sterol isoprenoids. The successful completion of these studies outlined in the grant application should enhance the understanding of the molecular basis of hepatic cholesterol production.