Minimal Lipidation of Pre-β HDL by ABCA1 Results in Reduced Ability to Interact with ABCA1

Abstract

Objectives-The aim of this study was to determine the role of ATP binding cassette transporter A1 (ABCA1) on generation of different-sized nascent HDLs. Methods and Results-HEK293 cells stably-transfected with ABCA1 (HEK293-ABCA1) or non-transfected (control) cells were incubated with lipid free 125 I-apoA-I for 24 hours. Incubation of apoA-I with HEK293-ABCA1 cells, but not control cells, led to the formation of heterogeneous-sized, pre-␤ migrating nascent HDL subpopulations (pre-␤1 to -4) that varied in size (7.1 to 15.7 nm), lipid, and apoA-I content. Kinetic studies suggested that all subpopulations were formed simultaneously, with no evidence for a precursor-product relationship between smaller and larger-sized particles. When isolated nascent pre-␤ HDLs (pre-␤1 to -4) were added back to HEK293-ABCA1 cells, their ability to bind to ABCA1 and efflux lipid was severely compromised. Heat-denaturation of pre-␤1 HDL resulted in partial recovery of ABCA1 binding, suggesting that initial interaction of apoA-I with ABCA1 results in a constrained conformation of apoA-I that decreases subsequent binding. Key Words: ATP binding cassette transporter A1 Ⅲ apolipoprotein AI Ⅲ high density lipoprotein T he inverse relationship between plasma high-density lipoprotein (HDL) cholesterol concentration and the risk of premature atherosclerotic vascular diseases has generated interest in understanding the steps involved in HDL assembly and catabolism. HDLs are proposed to be antiatherogenic because of their ability to accept excess cellular cholesterol in peripheral tissues and transport it to the liver in a process denoted as reverse cholesterol transport (RCT). 1 HDLs are classified into two subpopulations based on electrophoretic mobility on agarose gels: ␣-HDL (90% to 95% total HDL in plasma) and pre-␤ HDL (5% to 10%). 2 Heterogeneity is found among pre-␤ and ␣-HDLs. Using 2-dimensional gel electrophoresis (2D-gel), several pre-␤ and ␣-HDL subpopulations have been identified as a function of increasing size. 3 Although heterogeneity among pre-␤ and ␣-HDLs is well-documented, little is known about the mechanism of formation and metabolism of these pre-␤ and ␣-HDL subpopulations. Conclusions-InteractionABCA1 is critical for maintaining normal plasma HDL levels and nascent HDL biogenesis. A critical role for ABCA1 in HDL metabolism was demonstrated in patients with Tangier disease, a genetic disorder in which the ABCA1 gene is mutated. 4 -6 These patients have plasma HDL-cholesterol and apoA-I concentrations Ͻ5% of normal, accumulation of CE in macrophages, and increased risk of atherosclerosis. 7,8 Fibroblasts from these patients also have a significant reduction in lipid efflux to apoA-I compared with controls, suggesting ABCA1 was essential for mediating transport of intracellular lipid to apoA-I. 9 Additionally, ABCA1 knockout mice recapitulate the Tangier disease phenotype, verifying the role of ABCA1 in cellular lipid homeostasis and plasma HDL maintenance. 10 Plasma pre-␤ HDLs exist as several subpopulations...

Effects of Dietary Flavonoids on Reverse Cholesterol Transport, HDL Metabolism, and HDL function

“…Accordingly, deficiency of these ATP-binding cassette transporters in macrophages has been shown to reduce RCT in mice (26). HDL formation, or biogenesis, involves the efflux of phospholipids and free cholesterol by ABCA1 to lipid-free apoA-I in the extracellular space, generating nascent HDL particles (27). After ABCA1 facilitates HDL biogenesis, these nascent HDLs become efficient substrates for further lipidation by other cellular pathways (e.g., ABCG1) and esterification of free cholesterol to cholesteryl esters (CEs) by lecithin-cholesterol acyltransferase (LCAT) (28,29).…”

Structure of Apolipoprotein A-I N Terminus on Nascent High Density Lipoproteins

“…First, lipid free/lipid-poor apoA-I acquires phospholipid and cholesterol from the ATP-binding cassette transporter A1 (ABCA1) (6,7) and generates nascent HDL. Second, cholesterol on nascent HDL is esterified by lecithin: cholesterol acyltransferase to yield cholesteryl ester (8,9) and mature spherical HDL.…”

Characterization and properties of preβ-HDL particles formed by ABCA1-mediated cellular lipid efflux to apoA-I

“…The mechanism involves membrane phospholipid (PL) translocation via ABCA1 that induces bending of the membrane to create high-curvature sites to which apolipoproteins, such as apolipoprotein A-I (apoA-I), can bind and solubilize membrane PL and free (unesterified) cholesterol (FC) to create nascent HDL particles (7). In the case of apoA-I, the principal protein of HDL, the major nascent HDL products [lipoprotein A-I (LpA-I)] comprise discoidal particles containing two, three, or four apoA-I molecules per particle (8)(9)(10)(11). These discoidal particles are the progenitors of spherical HDL 2 and HDL 3 particles that form the major fraction of circulating HDL in human plasma.…”