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Molecular Regulation of Apoprotein B Degradation

载脂蛋白 B 降解的分子调控

基本信息

批准号：
8764600
负责人：
Edward A Fisher
金额：
$ 32.63万
依托单位：
NEW YORK UNIVERSITY SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-12-01 至 2015-09-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8764600
关键词：
Address Animals Apoprotein (B)Atherosclerosis Autophagocytosis Biochemical Biogenesis Biological Assay Blood Cell Culture Techniques Cell-Free System Cells Characteristics Chemicals Cholesterol Complex Coronary Arteriosclerosis Cultured Cells Data Decision Making Degradation Pathway Development Diet Dietary Fatty Acid Down-Regulation Dyslipidemias Endoplasmic Reticulum Fatty Acids Fish Oils Fractionation Genetic Goals Golgi Apparatus Hepatic Hepatocyte Human Hypobetalipoproteinemia In Vitro Incubated Insulin Lead Ligands Link Lipid Peroxidation Lipids Lipoproteins Liver Low-Density Lipoproteins Measures Mediating Mediator of activation protein Metabolic Metabolism Methods Modification Molecular Molecular Chaperones Mus Mutation Omega-3 Fatty Acids Oxidants Pathway interactions Physiologic pulse Plasma Play Polyunsaturated Fatty Acids Process Production Proteins Proteolysis Quality Control Reactive Oxygen Species Regulation Risk Factors Role Source Stream Superoxides Surveys System Techniques Testing Transport Process Ubiquitin Very low density lipoprotein Yeast Model System Yeasts apolipoprotein B-48 basal insulin base clinically relevant cohort density heart disease risk in vivo inhibitor/antagonist interest method development microsomal triglyceride transfer protein mouse model multicatalytic endopeptidase complex mutant novel novel therapeutic intervention oxidant stress particle protein p58 secretory protein success

项目摘要

The plasma level of apoprotein B (apoB) is among the strongest risk factors for coronary artery disease; thus, understanding the regulation of apoB-lipoprotein production is fundamentally interesting, clinically relevant, and may ultimately suggest new therapeutic approaches to dyslipidemias. ApoB100 is the form of apoB made by human liver and is the predominant protein component of atherogenic very low (VLDL) and low density (LDL) lipoproteins. Secretion of apoB100 from hepatic cells is controlled primarily by pre-secretory degradation. We have been at the forefront of defining apoB100 degradative pathways, and in this proposal focus on 1) endoplasmic reticulum-associated degradation (ERAD), which is mediated by the proteasome; and, 2) post- ER, presecretory proteolysis (PERPP), which is stimulated by dietary fatty acids used clinically to reduce VLDL levels, and which we believe is mediated by autophagy. In aim 1, we propose to characterize the process that targets nascent apoB for ERAD. We have previously shown that this process requires a distinct cohort of chaperones and chaperone-like proteins. We will continue to identify the factors that control apoB biogenesis in the ER using in vitro and in vivo assays, as well as a new apoB yeast expression system. In aim 2, we propose to determine the role of autophagy in post-ER apoB100 turnover under basal and perturbed metabolic states. Based on our recent data, we hypothesize that autophagy regulates the degradation of apoB100 under basal conditions and when hepatic cells are incubated with fish oil fatty acids (such as DHA) or other polyunsaturated fatty acids (PUFA) that lower VLDL levels. To address this hypothesis, we will study apoB100 turnover in hepatic cells and in mice in which the activities of specific autophagic factors have been manipulated. Further, based on the characteristics of insulin-mediated apoB degradation, we will also test the hypothesis that autophagy and the insulin-responsive pathways intersect to modulate apoB100 metabolism. In aim 3, we propose to determine the oxidant(s) responsible for apoB100 degradation and the effects of reactive oxygen species (ROS) on the VLDL assembly process. When hepatic cells are incubated with DHA and other PUFA, ROS and lipid peroxidation increase and apoB100 becomes damaged, aggregated, and targeted for autophagy. Based on our preliminary data, we hypothesize that superoxide (SO) plays a central role in apoB100 aggregation/degradation. We will test this hypothesis in hepatic cells and mice with genetic alterations in oxidant pathways. Other preliminary data suggest that PUFA decrease the amount of fully matured VLDL particles in the Golgi. By combining pulse-chase studies with sub-cellular fractionation and lipid analytical techniques, we will determine whether this results from aborted VLDL assembly or from the rapid targeting of VLDL to autophagy after assembly. In summary, these studies will lead to a more detailed molecular understanding of the intracellular metabolism of apoB100 and will contribute to the development of methods to control the production of atherogenic lipoproteins in normal and pathological states.

血浆载脂蛋白 B (apoB) 水平是冠状动脉疾病的最强危险因素之一；因此，了解 apoB-脂蛋白产生的调节非常有趣，具有临床相关性，并且可能最终提出血脂异常的新治疗方法。 ApoB100 是 apoB 的形式，由人类肝脏，是致动脉粥样硬化性极低 (VLDL) 和低密度 (LDL) 的主要蛋白质成分脂蛋白。肝细胞中 apoB100 的分泌主要受分泌前降解的控制。我们一直处于定义 apoB100 降解途径的最前沿，在本提案中重点关注 1) 内质网相关降解（ERAD），由蛋白酶体介导；以及，2) 后 ER，分泌前蛋白水解 (PERPP)，由临床上用于降低 VLDL 的膳食脂肪酸刺激水平，我们认为这是由自噬介导的。在目标 1 中，我们建议描述以下过程： ERAD 的目标是新生 apoB。我们之前已经表明，这个过程需要一群不同的人分子伴侣和分子伴侣样蛋白。我们将继续确定控制 apoB 生物发生的因素使用体外和体内测定以及新的 apoB 酵母表达系统的 ER。在目标 2 中，我们提议确定自噬在基础代谢和扰动代谢下 ER 后 apoB100 周转中的作用州。根据我们最近的数据，我们假设自噬在条件下调节 apoB100 的降解基础条件以及当肝细胞与鱼油脂肪酸（例如 DHA）或其他多不饱和脂肪酸（PUFA）可降低极低密度脂蛋白（VLDL）水平。为了解决这个假设，我们将研究 apoB100 肝细胞和小鼠体内特定自噬因子活性的更新被操纵。进一步，根据胰岛素介导的apoB降解的特点，我们还将测试假设自噬和胰岛素反应途径交叉调节 apoB100 代谢。在目标 3，我们建议确定导致 apoB100 降解的氧化剂以及反应性的影响氧自由基 (ROS) 对 VLDL 组装过程的影响。当肝细胞与 DHA 和其他物质一起孵育时 PUFA、ROS 和脂质过氧化增加，apoB100 受损、聚集并靶向自噬。根据我们的初步数据，我们假设超氧化物 (SO) 在 apoB100 聚集/降解。我们将在肝细胞和具有遗传基因的小鼠中测试这一假设氧化途径的改变。其他初步数据表明，PUFA 完全减少了高尔基体中成熟的 VLDL 颗粒。通过将脉冲追踪研究与亚细胞分离和脂质相结合分析技术，我们将确定这是否是由于 VLDL 装配中止或快速组装后将 VLDL 靶向自噬。总而言之，这些研究将带来更详细的对 apoB100 细胞内代谢的分子理解，将有助于开发在正常和病理状态下控制致动脉粥样硬化脂蛋白产生的方法。

项目成果

期刊论文数量（36）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Docosahexaenoic acid impairs the maturation of very low density lipoproteins in rat hepatic cells.

二十二碳六烯酸会损害大鼠肝细胞中极低密度脂蛋白的成熟。

DOI：
10.1194/jlr.m043026
发表时间：
2014
期刊：
Journal of lipid research
影响因子：
6.5
作者：
Maitin,Vatsala;Andreo,Ursula;Guo,Liang;Fisher,EdwardA
通讯作者：
Fisher,EdwardA

Inhibition of miR-33a/b in non-human primates raises plasma HDL and lowers VLDL triglycerides.

DOI：
10.1038/nature10486
发表时间：
2011-10-19
期刊：
NATURE
影响因子：
64.8
作者：
Rayner, Katey J.;Esau, Christine C.;Hussain, Farah N.;McDaniel, Allison L.;Marshall, Stephanie M.;van Gils, Janine M.;Ray, Tathagat D.;Sheedy, Frederick J.;Goedeke, Leigh;Liu, Xueqing;Khatsenko, Oleg G.;Kaimal, Vivek;Lees, Cynthia J.;Fernandez-Hernando, Carlos;Fisher, Edward A.;Temel, Ryan E.;Moore, Kathryn J.
通讯作者：
Moore, Kathryn J.

Insulin-stimulated degradation of apolipoprotein B100: roles of class II phosphatidylinositol-3-kinase and autophagy.