Molecular Regulation of Apoprotein B Degradation

载脂蛋白 B 降解的分子调控

• 批准号: 7066018
• 负责人: Edward A Fisher
• 金额: $ 34.77万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-07-15 至 2008-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7066018
• 关键词: antioxidants; apolipoprotein B; blood lipoprotein biosynthesis; cell free system; endoplasmic reticulum; fatty acids; fungal genetics; heat shock proteins; hyperlipidemia; insulin sensitivity /resistance; laboratory mouse; laboratory rat; liver cells; molecular chaperones; omega 3 fatty acid; oxidative stress; proteasome; protein degradation; protein transport; tissue /cell culture; ubiquitin

DESCRIPTION (provided by applicant): This is a renewal application to continue studies on the molecular regulation of apoprotein B (apoB) degradation. ApoB is the predominant protein component of the atherogenic lipoproteins. Thus, knowledge of the regulation of the assembly and secretion of apoB-lipoproteins is not only of fundamental interest, but is also medically relevant. An important control of the net secretion of apoB from cells of hepatic origin is the level of its pre-secretory degradation, one of the many unusual features of the biosynthesis and assembly of apoB into lipoprotein particles. We have recently established powerful cell-free systems to complement studies in cultured cells and animal models to identify and investigate the molecular factors that target apoB to degradation by proteasomal and non-proteasomal mechanisms as well as those that mediate apoB exit from the ER as part of a lipoprotein particle. The application is divided into 3 proposed aims: Aim 1 is to determine the spectrum of factors regulating the proteasome-mediated ER-associated degradation (ERAD) of apoB. We have previously established that cytosolic Hsp70 and Hsp90 promote ERAD of apoB and propose that known Hsp70 and Hsp90 co-chaperones and accessory factors also play important roles. Aim 2 is to determine the role of oxidant stress in the post-ER, non-proteasomal degradation of apoB induced by n-3 fatty acids. N-3 fatty acids are known hypolipidemic agents and we have recent data that in hepatic cells they stimulate a post-ER, non-proteasomal, degradative pathway (with remarkable similarity to the one induced by insulin) that is blocked by anti-oxidants. The molecular characteristics of n-3-stimulated apoB degradation, particularly with regard to oxidant stress, and its relationship to the insulin-stimulated process, will be tested in cell culture systems and in 2 recently described mouse models of human diseases, one of familial combined hyperlipidemia (and which has increased hepatic antioxidant levels) and one of insulin resistance (the Akt2 KO mouse). Aim 3 is to determine the factors regulating the ER-exit of apoB-containing lipoproteins. Based on classical studies of protein trafficking, the large size and other properties of apoB lipoproteins make it likely that there are novel features that drive their packaging and exit from the ER. Using cell-free model systems (established, to our knowledge, for the first time for this purpose), and hepatic cells, we will identify the molecular characteristics of the ER-exit process.

描述（由申请方提供）：这是一份延续申请，旨在继续研究载脂蛋白B（apo B）降解的分子调控。ApoB是致动脉粥样硬化脂蛋白的主要蛋白组分。因此，了解apoB-脂蛋白的组装和分泌的调节不仅具有根本意义，而且与医学相关。apoB从肝源细胞的净分泌的一个重要控制是其分泌前降解的水平，这是apoB生物合成和组装成脂蛋白颗粒的许多不寻常的特征之一。我们最近建立了强大的无细胞系统，以补充在培养细胞和动物模型中的研究，以确定和研究靶向apoB通过蛋白酶体和非蛋白酶体机制降解的分子因子，以及介导apoB作为脂蛋白颗粒的一部分从ER退出的分子因子。本申请分为3个目标：目标1是确定apoB蛋白酶体介导的ER相关降解（ERAD）的调控因子谱。我们以前已经建立了胞质Hsp 70和Hsp 90促进ERAD的载脂蛋白B，并提出，已知的Hsp 70和Hsp 90的辅助分子和辅助因子也发挥重要作用。目的2是确定氧化应激在ER后n-3脂肪酸诱导的apoB非蛋白酶体降解中的作用。N-3脂肪酸是已知的降血脂剂，我们最近的数据表明，在肝细胞中，它们刺激ER后非蛋白酶体降解途径（与胰岛素诱导的途径非常相似），该途径被抗氧化剂阻断。将在细胞培养系统和2种最近描述的人类疾病小鼠模型中测试n-3刺激的apoB降解的分子特征，特别是关于氧化应激及其与胰岛素刺激过程的关系，所述小鼠模型为家族性混合型高脂血症（并且具有增加的肝脏抗氧化剂水平）和胰岛素抵抗（Akt 2 KO小鼠）。目的3是确定含apoB脂蛋白ER出的调控因素。基于蛋白质运输的经典研究，apoB脂蛋白的大尺寸和其他特性使得可能存在驱动其包装并从ER退出的新特征。使用无细胞模型系统（据我们所知，首次为此目的建立）和肝细胞，我们将确定ER退出过程的分子特征。