HDL GENE DISCOVERY--GENOME WIDE EXPRESSION SCREENS

基于我们对高密度脂蛋白（HDL）代谢的不完全理解，本提案的重点是通过全基因组表达筛选鉴定参与这种脂蛋白代谢的新基因。将使用含有>5000个小鼠基因的小鼠cDNA阵列来鉴定在基于HDL代谢的特征性异常选择的几种转基因和敲除系小鼠的肝脏和肾上腺中表达改变的基因。这最初将包括载脂蛋白A-I（apo A-I）、清道夫受体b1类（sr-bi）、肝脂肪酶（HL）和脂蛋白胆固醇酰基转移酶（LCAT）的转基因和敲除小鼠。这些研究中的一个基本假设是，已知参与HDL代谢的基因表达的改变将影响也参与该脂蛋白代谢的其他基因的表达。从这些研究中鉴定的新基因将优先用于基于多种参数的进一步生物学表征，所述参数包括：表达变化水平、不同HDL突变基因型小鼠之间的表达模式聚类、以及与已知参与脂蛋白代谢的其他基因的序列或表达模式相似性。每年将通过转基因小鼠中的过表达以及对转基因过表达对脂蛋白代谢的后果的仔细分析来评估有限数量的新型“HDL候选”基因（约10个）的功能。在这些研究中，我们将利用新技术和先前开发的实验底物的组合来解决哪些基因直接或间接参与体内HDL代谢的基本问题。

Based on the incompleteness of our understanding of High Density Lipoprotein (HDL) metabolism the focus of this proposal is the identification of new genes involved in the metabolism of this lipoprotein through genome-wide expression screens. Mouse cDNA arrays containing >5000 mouse genes will be used to identify genes whose expression is altered in the liver and adrenals of several transgenic and knockout lines of mice chosen based on their characterized abnormalities in HDL metabolism. This will initially include transgenic and knockout mice for apolipoprotein A-I (apo A-I), Scavenger Receptor class b1 (sr- bi), Hepatic lipase (HL), and Lecithin Cholesterol Acyl Transferase (LCAT). A basic assumption in these studies is that alterations in the expression of genes known to be involved in HDL metabolism will affect the expression of other genes also participating in the metabolism of this lipoprotein. The novel genes identified from these studies will be prioritized for further biological characterization based on a variety of parameters including: level of expression change, clustering of expression patterns between mice of different HDL mutant genotypes, and sequence or expression pattern similarities to other genes known to participate in lipoprotein metabolism. The function of a limited number of novel "HDL candidate" genes (approximately 10) will be assessed each year through their over-expression in transgenic mice coupled with careful analysis of the consequence of transgene over-expression over-expression on lipoprotein metabolism. In these studies we will be utilizing a combination of new technologies and previously developed experimental substrates to address the fundamental question of what genes are directly or indirectly involved in the metabolism of HDL in vivo.