ApoA1 lipidation by ABCA1 in HDL biogenesis

流行病学研究表明，高密度脂蛋白-胆固醇（HDL-C）水平高与心血管疾病（CVD）风险降低相关。然而，最近的基因和药物研究表明HDL-C本身可能不是降低心血管疾病风险的原因。相反，一种共识正在形成，即HDL的功能可以预防心血管疾病，并且治疗HDL- c的生物标志物可能并不总是与HDL功能的增加相一致。HDL可能发挥其保护作用的功能之一是其在胆固醇逆向转运（RCT）途径中的作用，在该途径中，胆固醇从外周组织移除并转移到肝脏排泄。HDL及其主要蛋白成分载脂蛋白ai （apoA1）是这一过程的关键组成部分。在RCT途径的第一步，脂质贫乏的apoA1通过细胞膜蛋白ABCA1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的HDL。ABCA1有几个很好的特征活性，磷脂酰丝氨酸的向外移位，以及它的配体apoA1的细胞表面结合。在本研究的最后一个周期中，我们描述了ABCA1的两种新活性，磷脂酰肌醇（4,5）二磷酸（PIP2）的向外易位，负责apoA1与ABCA1表达细胞的结合，以及溶酶体空泡atp酶（v - atp酶）向质膜的募集，导致apoA1的局部酸化，促进其展开和HDL的生物发生。在这些发现的基础上，我们建议开发一种无细胞的ABCA1重组系统，以便更好地了解ABCA1组装新生HDL的机制。在Aim 2中，我们探索ABCA1向质膜招募V- atp酶的机制。这些研究的成功完成将增加我们对新生HDL生成机制的认识，这可能会为增加HDL生物发生和HDL功能的新策略提供见解，并有助于预防心血管疾病。

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.