Molecular Basis of Cholesterol Metabolism

描述(由申请人提供)： 该计划项目赠款始于30年前，当时我们定义了用于控制胆固醇代谢的低密度脂蛋白受体(LDLR)途径，并表明LDLR的缺陷会导致家族性高胆固醇血症(FH)及其伴随的动脉粥样硬化。30年后，我们的目标扩大了，参与者增加了，但重点仍然是相同的：了解调节脂类代谢的遗传和分子基础，并利用这些知识预防和治疗与脂类相关的疾病，包括动脉粥样硬化、糖尿病和神经系统疾病。在上一次资助期间，我们发表了183篇论文。这些论文报道了以下主要进展：1)发现INSIG蛋白是胆固醇和脂肪酸合成反馈控制的中心；2)开发新的方法来证明胆固醇直接与类固醇敏感蛋白SCAP结合；3)证明胆固醇24-羟基酶介导成人大脑中的胆固醇转换；4)发现LDLR家族成员在大脑发育和功能(VLDL和APOE2受体)以及血管完整性和动脉粥样硬化(LRP1)中的新作用；5)证明富含胆固醇的小窝调节生长因子受体酪氨酸激酶；6)鉴定4个新基因及其在3种人类遗传性疾病中的致病作用：谷甾醇血症(AB5和ABCG8突变)；这些研究包括：1)发现了常染色体隐性遗传的FH(接头蛋白ARH)，以及选择性25-羟基维生素D缺乏(维生素D 25-羟基酶)；7)利用基因敲除小鼠证明PCSK9是肝脏低密度脂蛋白受体蛋白水平的主要调节基因；8)开发了非同义密码子方法，以寻找数量性状中的致病基因(例如，在2%的非裔美国人中，PCSK9的功能丧失突变使低密度脂蛋白降低35%)。 我们现在申请为期5年的续展(31-35年)，以通过综合和多学科的方法进一步研究这些现象和相关现象。我们建议更多地了解已知的分子，并发现调节3个过程的新分子：1)胆固醇和脂肪酸代谢(SREBPs、SCAP、Insigs、MOBP、HMG CoA还原酶、PCSK9、LDLR、NPC1L1、ABCG5/G8，可能是新的胰岛素敏感因子)；2)氧固醇和胆汁酸代谢(胆固醇24-羟基酶，新的类固醇羟基酶)；以及3)腔膜功能、脂蛋白受体和细胞信号(小窝蛋白-1、LRP1、LRP1B、PDGRF(EGFR))。我们将继续在所有可能的水平上研究这些过程--分子(即基因、信使核糖核酸、蛋白质)、完整的细胞、整个动物和人类患者。在进行这些研究时，我们将使用多种技术--生物化学、分子生物学、遗传学、细胞生物学、基因操纵的小鼠、动物生理学、临床遗传学和人类基因组学。这种跨学科的方法只有通过本计划项目赠款的支持才有可能实现。

DESCRIPTION (provided by applicant): This Program Project Grant began 30 years ago when we defined the LDL receptor (LDLR) pathway for control of cholesterol metabolism and showed that defects in LDLR produce Familial Hypercholesterolemia (FH) and its attendant atherosclerosis. After 30 years, our goals have broadened and the participants have increased, but the focus remains the same: to understand the genetic and molecular basis for regulation of lipid metabolism and to use this knowledge to prevent and treat lipid-related diseases, including atherosclerosis, diabetes, and neurologic diseases. During the last grant period, we published 183 papers. These papers report the following major advances: 1) discovery of Insig proteins as central to feedback control of cholesterol and fatty acid synthesis; 2) development of new methods to demonstrate direct cholesterol binding to the sterol-sensing protein Scap; 3) demonstration that cholesterol 24-hydroxylase mediates cholesterol turnover in adult brain; 4) discovery of new roles for LDLR family members in brain development and function (VLDL and apoE2 receptors) and in vascular integrity and atherogenesis (LRP1); 5) demonstration that cholesterol-rich caveolae regulate growth factor receptor tyrosine kinases; 6) identification of 4 new genes and their causative roles in 3 human genetic diseases: sitosterolemia (mutations in ABCG5 and ABCG8); autosomal recessive FH (adaptor protein ARH); and selective 25-hydroxyvitamin D deficiency (vitamin D 25-hydroxylase); 7) use of knockout mice to show that PCSK9 is a major regulator of LDLR protein levels in liver; and 8) development of the nonsynonymous codon approach to find causative genes in quantitative traits (e.g., loss-of-function mutations in PCSK9 lower LDL by 35% in 2% of African-Americans). We now apply for a 5-year renewal (Years 31-35) to further study these and related phenomena through an integrated and multidisciplinary approach. We propose to learn more about already-known molecules and discover new ones that regulate 3 processes: 1) cholesterol and fatty acid metabolism (SREBPs, Scap, Insigs, MOBP, HMG CoA reductase, PCSK9, LDLR, NPC1L1, ABCG5/G8, putative new insulin-sensitizing factor); 2) oxysterol and bile acid metabolism (cholesterol 24-hydroxylase, new sterol hydroxylases); and 3) caveolae membrane function, lipoprotein receptors, and cell signaling (caveolin-1, LRP1, LRP1B, PDGRF(, EGFR). We will continue to study these processes at all possible levels - the molecular (i.e., gene, mRNA, protein), the intact cell, the whole animal, and the human patient. In conducting these studies, we will use multiple techniques - biochemistry, molecular biology, genetics, cell biology, gene-manipulated mice, animal physiology, clinical genetics, and human genomics. This interdisciplinary approach is possible only through support of this Program Project Grant.