出版社：American Society for Biochemistry and Molecular Biology
摘要：The nascent HDL created by ABCA1-mediated efflux of cellularphospholipid (PL) and free (unesterified) cholesterol (FC) toapolipoprotein A-I (apoA-I) has not been defined. To addressthis issue, we characterized the lipid particles released whenJ774 mouse macrophages and human skin fibroblasts in which ABCA1is activated are incubated with human apoA-I. In both cases,three types of nascent HDL containing two, three, or four moleculesof apoA-I per particle are formed. With J774 cells, the predominantspecies have hydrodynamic diameters of 9 and 12 nm. These discoidalHDL particles have different FC contents and PL compositions,and the presence of acidic PL causes them to exhibit -electrophoreticmobility. These results are consistent with ABCA1 located inmore than one membrane microenvironment being responsible forthe production of the heterogeneous HDL. Activation of ABCA1also leads to the release of apoA-I-free plasma membrane vesicles(microparticles). These larger, spherical particles releasedfrom J774 cells have the same PL composition as the 12 nm HDLand contain CD14 and ganglioside, consistent with their originbeing plasma membrane raft domains. The various HDL particlesand microparticles are created concurrently, and there is noprecursor-product relationship between them. Importantly, alarge fraction of the cellular FC effluxed from these cellsby ABCA1 is located in microparticles. Collectively, these resultsshow that the products of the apoA-I/ABCA1 interaction includediscoidal HDL particles containing different numbers of apoA-Imolecules. The cellular PLs and cholesterol incorporated intothese nascent HDL particles originate from different cell membranedomains.Supplementary key words phospholipids • cholesterol • lipoprotein • reverse cholesterol transport • mouse macrophages • human skin fibroblasts • cell membranes • high density lipoprotein • ATP binding cassette transporter A1 • apolipoprotein A-I
Abbreviations: apoA-I, apolipoprotein A-I; BS3, bis(sulfosuccinimidyl) suberate; FC, free (unesterified) cholesterol; FPLC, fast-protein liquid chromatography; GM1, ganglioside; PC, phosphatidylcholine; PE, phosphatidylethanolanime; PL, phospholipid; PS, phosphatidylserine; SM, sphingomyelin
9 and 12 nm. These discoidal HDL particles have different FC contents and PL compositions, and the presence of acidic PL causes them to exhibit 
-electrophoretic mobility. These results are consistent with ABCA1 located in more than one membrane microenvironment being responsible for the production of the heterogeneous HDL. Activation of ABCA1 also leads to the release of apoA-I-free plasma membrane vesicles (microparticles). These larger, spherical particles released from J774 cells have the same PL composition as the 12 nm HDL and contain CD14 and ganglioside, consistent with their origin being plasma membrane raft domains. The various HDL particles and microparticles are created concurrently, and there is no precursor-product relationship between them. Importantly, a large fraction of the cellular FC effluxed from these cells by ABCA1 is located in microparticles. Collectively, these results show that the products of the apoA-I/ABCA1 interaction include discoidal HDL particles containing different numbers of apoA-I molecules. The cellular PLs and cholesterol incorporated into these nascent HDL particles originate from different cell membrane domains.
Supplementary key words phospholipids • cholesterol • lipoprotein • reverse cholesterol transport • mouse macrophages • human skin fibroblasts • cell membranes • high density lipoprotein • ATP binding cassette transporter A1 • apolipoprotein A-I
Abbreviations: apoA-I, apolipoprotein A-I; BS3, bis(sulfosuccinimidyl) suberate; FC, free (unesterified) cholesterol; FPLC, fast-protein liquid chromatography; GM1, ganglioside; PC, phosphatidylcholine; PE, phosphatidylethanolanime; PL, phospholipid; PS, phosphatidylserine; SM, sphingomyelin
