Multi-disciplinary Approaches to HDL Structure, Assembly, and Functional Heterogeneity

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SUMMARY (Entire PPG) The development of new cardioprotective drugs and accurate metrics for cardiovascular risk is being hindered, we believe, by the failure to correctly identify the cardioprotective forms of high-density lipoprotein (HDL) and inadequate knowledge about the mechanisms that remove cholesterol from the artery wall. This renewal application will address these issues by continuing dynamic interactions among seven world-class scientists who study HDL from diverse, but complementary, viewpoints. HDL, an important mediator of cholesterol transport, is created when its most abundant scaffold protein, apolipoprotein A-I (APOA1), interacts with ATP-binding cassette transporter A1 (ABCA1). This process is critical for removing cholesterol from macrophages. Indeed, the capacity for human plasma HDL to promote cholesterol efflux (CEC) is more diagnostic for cardiovascular disease (CVD) risk than the traditional measure of HDL’s cholesterol content. The central hypothesis of this multidisciplinary Program Project Grant is that ABCA1 engages in highly specific interactions with APOA1 particles that may or may not contain lipid to produce cardioprotective HDL. Our objective is to derive a molecular understanding of this pathway and the roles played by HDL subspecies in the setting of diabetes. We will use three general overlapping approaches: 1) calibrated ion mobility spectrometry to relate the size and number of specific HDL particles to CEC and CVD risk in diabetic patients, 2) detailed in vitro mechanistic approaches to unravel the factors that allow specific lipidated HDL particles to interact with ABCA1, and 3) cryo-EM and computational studies of the structure and mechanism of ABCA1’s action. Our plan focuses on three Projects at four sites: Project 1: Cardioprotection by extra-small HDL particles – Jay Heinecke, Project Leader; Karin Bornfeldt, Co-I (University of Washington); Project 2: Mechanism of ABCA1-mediated cholesterol efflux to lipidated HDL – W. Sean Davidson, Project Leader and PPG Principal Investigator (University of Cincinnati); Project 3: Mechanisms of phospholipid/cholesterol translocation by ABCA1 – Jere Segrest, Project Leader (Vanderbilt University); Steve Aller, Co-I (University of Alabama at Birmingham). Additionally, four core facilities will drive scientific synergy and cost-effective use of NIH resources: Core A: Administration – W. Sean Davidson, Core Leader, will provide administrative support for the Program. Core B: Computational Biology – Jere Segrest, Core Leader, will perform molecular modeling, homology modeling, and double/single state normal mode analyses for structural studies. Core C: Lipoprotein Quantitation and Function – Tomas Vaisar, Core Leader; Chongren Tang, Co-I, will quantify i) HDL particle number (the sizes and concentrations of HDL particles) and ii) the cholesterol efflux capacity of various HDLs and ABCA1 mutants. Core D: Apo/Lipoprotein Production – W. Sean Davidson, Core Leader, will supply the Program with high-quality apolipoproteins, customized mutants, and lipoprotein samples, both recombinantly derived and isolated from human plasma.