Post-translational Control of Triglyceride and Cholesterol Metabolism by ANGPTL3 & ANGPTL8 in ApoBCL Clearance

ANGPTL3 对甘油三酯和胆固醇代谢的翻译后控制

• 批准号: 10543874
• 负责人: Helen Haskell Hobbs
• 金额: $ 57.4万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2026-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10543874
• 关键词: ANGPTL3 gene; Acceleration; Affect; Angiopoietins; Antibody Specificity; Apolipoprotein E; Apolipoproteins B; Biochemical; Biological; Cardiometabolic Disease; Cells; Cholesterol; Cholesterol Homeostasis; Clinical Trials; Complement; Complex; Coronary heart disease; Cultured Cells; DNA Sequence; Defect; Development; Disease; Enzymes; Family member; Fatty Acids; Female; Functional disorder; Generations; Genetic; Genetic Screening; Genotype; Goals; Grant; Hepatic; Hepatocyte; Human; Hydrolysis; Hypertriglyceridemia; Knockout Mice; LIPG gene; Laboratories; Ligands; Link; Lipase; Lipids; Lipoprotein (a); Lipoproteins; Liver; Location; Low-Density Lipoproteins; Mediating; Metabolic; Metabolic Pathway; Metabolism; Molecular; Mus; Mutation; Mycoplasma; New Agents; Pathway interactions; Peripheral; Pharmaceutical Preparations; Phenotype; Plasma; Plasmids; Post-Translational Protein Processing; Prevention strategy; Process; Program Research Project Grants; Property; Proteins; Published Comment; Research Project Grants; Residual state; Resolution; Role; Route; Sample Size; Structure; Testing; Tissues; Triglyceride Metabolism; Triglycerides; Variant; Very low density lipoprotein; Work; antagonist; candidate identification; cardiovascular risk factor; experimental study; genetic approach; genetic manipulation; in vitro activity; in vivo; insight; lipid metabolism; lipoprotein lipase; male; novel strategies; protein function; receptor; reverse genetics; routine screening; scavenger receptor; sex; stoichiometry; success; syndecan; therapeutic development; uptake

PROJECT 3: Post-translational Control of Triglyceride and Cholesterol Metabolism by ANGPTL3 & ANGPTL8 in ApoBCL Clearance PROJECT SUMMARY The goal of this grant is to elucidate the molecular basis of the lipid-lowering effects of inactivating ANGPTL3 (A3) and ANGPTL8 (A8). Previously we discovered that inactivating mutations in A3 in humans are associated with reduced circulating levels of triglycerides (TGs). We showed that A3 forms a complex with a related protein, A8, to inactivate lipoprotein lipase (LPL), an intravascular enzyme in peripheral tissues that hydrolyzes circulating TG. Inactivation of either A3 or A8 reduced plasma TG levels by increasing LPL activity. Despite these advances in our understanding, fundamental questions remain about how A3 and A8 interact to inhibit LPL in vivo. Moreover, A3 has activities that are independent of A8. Inactivation of A3 dramatically lowers plasma cholesterol, as well as TG levels. This activity does not require A8 or any of the known hepatic lipoprotein clearance pathways. Our group, and that of Dan Rader’s, has shown that endothelial lipase (EL) is needed for the cholesterol-lowering effects of A3 inactivation. The scientific premise of this application is that elucidating the molecular mechanisms underlying the lipid-lowering effects of A3 and A8 inhibition will lead to new strategies for the prevention and treatment of cardiometabolic disorders. In AIM 1 we will determine the forms of A3/A8 and A3 that are operative in vivo in inhibiting LPL and EL, respectively. Both proteins are subject to post-translational modifications, including proteolytic cleavage and oligomerization. We will define the basic properties of the active forms of the native proteins with respect to cleavage (Aim 1A) and stoichiometry (Aim 1A), before purifying the active complexes and determining their high resolution structures (Aim 1C). In AIM 2 we will characterize molecularly the noncanonical pathway for hepatic clearance of ApoB-containing lipoproteins (ApoBL) in A3 deficiency and test 3 hypotheses regarding the molecular basis for this pathway: First, that EL-stimulated ApoBL uptake is a receptor-mediated endocytic process (Aim 2A); Second, that EL alters the composition of nascent VLDL, exposing a new ligand for ApoBL uptake by the liver (Aim 2B). Finally, we will establish an unbiased genetic screen in cultured hepatocytes to identify the receptor that mediates ApoBL uptake by liver and validate receptor activity in vivo (Aim 2C). Elucidating the molecular mechanisms by which A3 and A8 alter lipid metabolism will provide new insights into key metabolic pathways. They will also provide a mechanistic basis for a new generation of lipid-lowering agents. These goals complement the aims of Research Project 1 and Project 2 to understand what regulates the formation, location, and metabolic fate of lipids so as to develop new agents for the treatment of cardiometabolic disorders.

项目3：Angptl3&对甘油三酯和胆固醇代谢的翻译后调控 ApoBCL清除中的ANGPTL8 项目总结 这项资助的目的是阐明失活Angptl3降脂作用的分子基础。 (A3)和ANGPTL8(A8)。此前，我们发现人类A3基因的失活突变与 循环中甘油三酯(TGS)水平降低。我们发现A3与一种相关蛋白质形成了一个复合体， A8，用来灭活脂蛋白脂酶(LPL)，这是一种外周组织中的血管内酶，可以分解循环 Tg.A3或A8的失活可通过增加LPL活性来降低血浆甘油三酯水平。尽管有这些进步 在我们的理解中，基本的问题仍然是关于A3和A8如何在体内相互作用来抑制LPL。 此外，A3有独立于A8的活动。A3的失活显著降低了血浆胆固醇， 以及甘油三酯水平。这种活性不需要A8或任何已知的肝脂蛋白清除。 小路。我们和Dan Rader的团队已经表明，内皮脂肪酶(EL)是 A3失活的降胆固醇作用。这一应用的科学前提是阐明 A3和A8抑制降脂作用的分子机制将导致新的治疗策略 心脏代谢性疾病的防治。 在目标1中，我们将确定在体内抑制LPL和EL的A3/A8和A3的形式， 分别进行了分析。这两种蛋白质都受到翻译后修饰的影响，包括蛋白水解性切割和 齐聚作用。我们将定义天然蛋白质活性形式的基本性质 裂解(目标1A)和化学计量学(目标1A)，然后提纯活性络合物并测定它们的高 分辨率结构(目标1C)。在AIM 2中，我们将从分子上描述肝脏的非规范途径 A3缺乏症中含ApoB脂蛋白(ApoBL)的清除及关于A3缺乏症的3个假设检验 这一途径的分子基础：首先，EL刺激的载脂蛋白BL摄取是一种受体介导的内吞 过程(目标2A)；第二，EL改变新生极低密度脂蛋白的组成，暴露出ApoBL的新配体 肝脏摄取(目标2B)。最后，我们将在培养的肝细胞中建立一个无偏见的遗传筛选，以 确定介导肝脏摄取载脂蛋白的受体，并在体内验证受体活性(目标2C)。 阐明A3和A8改变脂代谢的分子机制将为 关键的代谢途径。它们还将为新一代降脂剂提供机理基础。 这些目标补充了研究项目1和项目2的目标，以了解是什么规范了 脂类的形成、定位和代谢去向，以开发治疗心脏代谢性疾病的新药物 精神错乱。