Structural and Functional Investigations on Cholesterol Signaling and Metabolism

胆固醇信号传导和代谢的结构和功能研究

• 批准号: 10623526
• 负责人: Xiaochun Li
• 金额: $ 46.74万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-22 至 2028-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10623526
• 关键词: Acetyl Coenzyme A; Adult; Anabolism; Atherosclerosis; Bile Acids; Binding; Biological; Blood; Cardiovascular Diseases; Cells; Cholesterol; Cholesterol Esters; Cholesterol Homeostasis; Coenzyme A; Complex; Cryoelectron Microscopy; Embryonic Development; Endoplasmic Reticulum; Enzymes; Erinaceidae; Growth; Homeostasis; Human; Incidence; Intestines; Investigation; LDL Cholesterol Lipoproteins; Lanosterol; Ligands; Ligase; Lipids; Lipoproteins; Liver; Maintenance; Malignant Neoplasms; Mammalian Cell; Mediating; Membrane; Membrane Proteins; Metabolism; Methods; Mevalonic Acid; Molecular; Oxidoreductase; Pathway interactions; Permeability; Pharmaceutical Preparations; Plasma; Play; Reaction; Regulation; Role; Series; Signal Transduction; Sterol O-Acyltransferase; Sterols; Structure; Tissues; Transducers; Vitamin D; Work; acyl group; cholesterol biosynthesis; hedgehog signal transduction; insight; novel; pharmacologic; prevent; small molecule; smoothened signaling pathway; steroid hormone biosynthesis; sterol O-acyltransferase 1; sterol O-acyltransferase 2; synthetic enzyme; trafficking; tumorigenesis; ubiquitin-protein ligase

Summary Cholesterol is an essential lipid that plays an important role in the maintenance of membrane rigidity and permeability and is essential for the growth and viability of mammalian cells. Cholesterol also functions as a precursor in the biosynthesis of steroid hormones, bile acids, vitamin D, and is a Hedgehog signaling transducer. In this project, we will employ cell biological and structural approaches to study the membrane proteins involved in cholesterol signaling (Hedgehog signaling pathway), biosynthesis (cholesterol synthetic enzymes), and storage (cholesterol esterification enzymes). 1) Dysregulation of Hedgehog (HH) signaling, which is required for proper embryonic development and adult tissue homeostasis, leads to tumorigenesis. Over the past five years, we have determined the structures of human PTCH1 alone, human PTCH1-HH complexes, SMO-Gi complexes in distinct states, and DISP1 alone and in complex with HH. These structures, along with our functional studies, provide molecular insights into HH signal transduction. In this project, we will continue to work on this pathway. Specifically, we will focus on the trafficking and signal regulation of the HH ligand via the HH-PTCH1 axis. 2) Eighteen enzymes convert acetyl-CoA into lanosterol, the first sterol-like intermediate in a series of reactions that synthesize cholesterol in the endoplasmic reticulum (ER). One key reaction is mediated by HMGCR (3- Hydroxy-3-methylglutaryl coenzyme A reductase), which catalyzes the conversion of acetyl-CoA to mevalonic acid. HMGCR is a target of the cholesterol-lowering drugs statins due to its role as the rate-limiting enzyme in cholesterol synthesis. UBIAD1 stabilizes HMGCR and prevents its degradation, while the binding of the E3 ligase gp78 and Insig trigger the degradation of HMGCR. Recently, we determined the cryo-EM structures of HMGCR bound to UBIAD1. The successful completion of this project will aid our investigation into how HMGCR is ubiquitinated by gp78 via Insig. In addition, we will study how the membrane-embedded cholesterol synthetases carry out the cholesterol biosynthesis. 3) In the ER, the ACAT enzymes (ACAT-1 and ACAT-2) catalyze the transfer of long-chain fatty acyl groups to cholesterol, generating cholesterol esters that are integrated into lipoproteins for secretion or storage in lipid droplets. This reaction results in a low cholesterol concentration in the ER and is crucial for maintaining intracellular cholesterol homeostasis. ACAT-1 is ubiquitously expressed in tissues, whereas ACAT-2 expression is restricted to the liver and intestine. Pharmacological inhibition of ACAT- 2 reduces blood cholesterol levels and atherosclerosis. Using structural methods, we plan to investigate mechanisms for lipid-mediated regulation of ACATs to gain insight into novel modes of inhibition and use this information to develop small molecules that specifically inhibit ACAT-2 and lower plasma LDL cholesterol.

概括 胆固醇是一种必不可少的脂质，在维持膜刚度和 渗透性对于哺乳动物细胞的生长和生存力至关重要。胆固醇也充当 类固醇激素，胆汁酸，维生素D的生物合成中的前体，是刺猬信号传导剂。 在这个项目中，我们将采用细胞生物学和结构方法来研究所涉及的膜蛋白 在胆固醇信号（刺猬信号通路）中，生物合成（胆固醇合成酶）和 储存（胆固醇酯化酶）。 1）刺猬（HH）信号的失调，这是必需的 适当的胚胎发育和成人组织稳态导致肿瘤发生。在过去的五年中， 我们已经确定了人类PTCH1，人PTCH1-HH复合物，SMO-GI复合物的结构 在不同的状态下，单独使用DESP1，并与HH复杂。这些结构以及我们的功能研究 向HH信号转导提供分子见解。在这个项目中，我们将继续在这一途径上工作。 具体而言，我们将通过HH-PTCH1轴关注HH配体的运输和信号调节。 2） 18酶将乙酰辅酶A转化为羊毛醇，这是一系列反应中的第一个类固醇样中间体 这可以合成内质网中（ER）中的胆固醇。一个关键反应是由HMGCR介导的（3- 羟基-3-甲基戊二酰辅酶A还原酶），该辅酶会催化乙酰辅酶A向甲硅酸盐的转化 酸。 HMGCR是降低胆固醇药物汀类药物的靶标，因为它是限速酶的作用 胆固醇合成。 UBIAD1稳定HMGCR并防止其降解，而E3连接酶的结合 gp78和Insig触发了HMGCR的降解。最近，我们确定了HMGCR的冷冻EM结构 绑定到ubiad1。该项目的成功完成将有助于我们调查HMGCR的方式 通过Insig泛滥成灾。此外，我们将研究膜上添加的胆固醇合成酶 进行胆固醇生物合成。 3）在ER中，ACAT酶（ACAT-1和ACAT-2）催化 将长链脂肪酰基基团转移至胆固醇，产生胆固醇酯的整合 脂蛋白用于分泌或在脂质液滴中储存的脂蛋白。该反应导致胆固醇浓度较低 ER，对于维持细胞内胆固醇稳态至关重要。 ACAT-1无处不在 组织，而ACAT-2表达仅限于肝脏和肠。药理学抑制ACAT- 2降低了血液胆固醇水平和动脉粥样硬化。使用结构方法，我们计划调查 脂质介导的ACAT调节的机制，以深入了解新型抑制模式并使用 信息以开发专门抑制ACAT-2和较低血浆LDL胆固醇的小分子。