HDL GENE DISCOVERY--GENOME WIDE EXPRESSION SCREENS

基于我们对高密度脂蛋白(高密度脂蛋白)代谢的了解还不完全，这项建议的重点是通过全基因组表达筛选来识别参与高密度脂蛋白代谢的新基因。含有&gt；5000小鼠基因的小鼠基因阵列将被用来识别在几个转基因和基因敲除小鼠的肝脏和肾上腺中表达发生变化的基因，这些基因是根据它们在高密度脂蛋白代谢方面的特征异常而选择的。这将首先包括载脂蛋白A-I(apo A-I)、清道夫受体b1类(sr-bi)、肝脂酶(HL)和卵磷脂胆固醇酰基转移酶(LCAT)的转基因和基因敲除小鼠。这些研究中的一个基本假设是，已知的参与高密度脂蛋白代谢的基因表达的变化将影响也参与这种脂蛋白代谢的其他基因的表达。从这些研究中确定的新基因将被优先用于基于各种参数的进一步生物学特征，包括：表达水平的变化，不同高密度脂蛋白突变类型小鼠之间的表达模式聚集，以及与其他已知的参与脂蛋白代谢的基因的序列或表达模式相似性。每年将通过在转基因小鼠中过度表达以及仔细分析转基因过度表达对脂蛋白代谢的影响来评估有限数量的新“高密度脂蛋白候选”基因(约10个)的功能。在这些研究中，我们将利用新技术和以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高密度脂蛋白代谢的根本问题。

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.