title: ABCA1 mediates unfolding of apolipoprotein AI N terminus on the cell surface before lipidation and release of nascent high-density lipoprotein.
abstract: OBJECTIVE
  To gain insight into the mechanism by which ABCA1 generates nascent high-density lipoprotein.
  
  
  RESULTS
  HEK293 cells were stably transfected with ABCA1 vectors, encoding wild type, and the W590S and C1477R Tangier disease mutation isoforms, along with the K939M ATP-binding domain mutant. Apolipoprotein AI (ApoAI) binding, plasma membrane remodeling, cholesterol efflux, apoAI cell surface unfolding, and apoAI cell surface lipidation were determined, the latter 2 measured using novel fluorescent apoAI indicators. The W590S isoform had decreased plasma membrane remodeling and lipid efflux activities, and the C1477R isoform had decreased apoAI binding, and lipid efflux activities, whereas the K939M isoform did not bind apoAI, remodel the membrane, or efflux cholesterol. However, all ABCA1 isoforms led to apoAI unfolding at the cell surface, which was higher for the isoforms that increased apoAI binding. ApoAI lipidation was not detected on ABCA1-expressing cells, only in the conditioned medium, consistent with rapid release of nascent high-density lipoprotein from ABCA1-expressing cells.
  
  
  CONCLUSIONS
  We identified a third activity of ABCA1, the ability to unfold the N terminus of apoAI on the cell surface. Our results support a model in which unfolded apoAI on the cell surface is an intermediate in its lipidation and that, once apoAI is lipidated, it forms an unstable structure that is rapidly released from the cells to generate high-density lipoprotein.
authors.affiliation: Department of Cellular and Molecular Medicine, Cleveland Clinic, Cleveland, OH 44195, USA.
authors.forename: Shuhui
authors.initials: S
authors.lastname: Wang
authors.forename: Kailash
authors.initials: K
authors.lastname: Gulshan
authors.forename: Gregory
authors.initials: G
authors.lastname: Brubaker
authors.forename: Stanley L
authors.initials: SL
authors.lastname: Hazen
authors.forename: Jonathan D
authors.initials: JD
authors.lastname: Smith
mesh_terms: D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism*; D000818:Animals; D016632:Apolipoprotein A-I / Q000378:metabolism*; D001692:Biological Transport / Q000502:physiology; D002478:Cells, Cultured; D057809:HEK293 Cells / Q000378:metabolism*; D006801:Humans; D050356:Lipid Metabolism / Q000502:physiology*; D008075:Lipoproteins, HDL / Q000378:metabolism*; D051379:Mice; D011485:Protein Binding / Q000502:physiology; D012680:Sensitivity and Specificity; D013631:Tangier Disease / Q000378:metabolism / Q000503:physiopathology; D014162:Transfection
chemical_list: C577836:ABCA1 protein, human; D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D016632:Apolipoprotein A-I; D008075:Lipoproteins, HDL
keywords: ABCA1; apolipoprotein AI unfolding; nascent high-density lipoprotein; reverse cholesterol transport
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