ApoA1 lipidation by ABCA1 in HDL biogenesis

在流行病学研究中，高密度脂蛋白胆固醇（HDL-C）水平与心血管疾病（CVD）风险降低相关。然而，最近的遗传和药物研究表明，HDL-C本身可能不是降低CVD风险的原因。相反，一个共识是HDL功能可以防止CVD，并且治疗HDL-C的生物标志物可能并不总是与增加HDL功能相一致。HDL在其保护作用中可能发挥作用的功能之一是其在胆固醇逆向转运（RCT）途径中的作用，其中胆固醇从外周组织中去除并转移到肝脏进行排泄。HDL及其主要蛋白质成分载脂蛋白AI（apoA 1）是这一过程的关键组成部分。在RCT途径的第一步中，贫脂apoA 1通过细胞膜蛋白ABCA 1作为细胞胆固醇和磷脂的受体，通过分子水平上尚不清楚的机制产生新生HDL。ABCA 1具有几种充分表征的活性，磷脂酰丝氨酸的向外易位，以及其配体apoA 1的细胞表面结合。在该资助的最后一个周期中，我们表征了ABCA 1的两种新活性，即负责apoA 1与ABCA 1表达细胞结合的磷脂酰肌醇（4，5）二磷酸（PIP 2）的向外易位，以及溶酶体空泡ATP酶（V-ATP酶）向质膜的募集，导致apoA 1的局部酸化，从而促进其展开和HDL生物合成。基于这些发现，我们建议开发一种无细胞ABCA 1重建系统，从而更好地了解ABCA 1组装新生HDL的机制。目的2：探讨ABCA 1募集V-ATP酶至质膜的机制.成功完成拟议的研究将增加我们对从头HDL产生机制的了解，这可能会产生对增加HDL生物合成和HDL功能的新策略的见解，并有助于预防CVD。

High levels of high density lipoprotein-cholesterol (HDL-C) are associated with lowered risk for cardiovascular disease (CVD) in epidemiological studies. However, recent genetic and drug studies have shown that HDL-C itself is probably not causal in reducing CVD risk. Instead, a consensus is building that HDL functions may protect against CVD, and that treating for the biomarker of HDL-C may not always coincide with increased HDL function. One of the functions of HDL that may play a role in its protective effect is its role in the reverse cholesterol transport (RCT) pathway, in which cholesterol is removed from peripheral tissues and transferred to the liver for excretion. HDL and its major protein constituent, apolipoprotein-AI (apoA1), are critical components of this process. In the first step of the RCT pathway, lipid-poor apoA1 acts as an acceptor for cell cholesterol and phospholipids via the cell membrane protein ABCA1, generating nascent HDL through a mechanism which is not understood at the molecular level. ABCA1 has several well characterized activities, the outward translocation of phosphatidylserine, and the cell surface binding of its ligand apoA1. In the last cycle of this grant we characterized two novel activities of ABCA1, the outward translocation of phosphatidylinositol (4,5) bis-phosphate (PIP2) that is responsible for apoA1 binding to ABCA1 expressing cells, and the recruitment of the lysosomal vacuolar ATPase (V-ATPase) to the plasma membrane leading to local acidification of apoA1 that promotes its unfolding and HDL biogenesis. Building on these discoveries we propose to develop a cell-free ABCA1 reconstituted system, allowing a better understanding of the mechanism by which ABCA1 assembles nascent HDL. In Aim 2, we explore the mechanism by which ABCA1 recruits V- ATPase to the plasma membrane. Successful completion of the proposed studies will increase our knowledge of the mechanism of de novo HDL production, which may yield insights into new strategies to increase HDL biogenesis and HDL function, and aid in the prevention of CVD.