Biogenesis and speciation of nascent apoA-I-containing particles in various cell lines

Krimbou, Larbi; Hassan, Houssein Hajj; Blain, Sacha; Rashid, Shirya; Denis, Maxime; Marcil, Michel; Genest, Jacques

doi:10.1194/jlr.m500038-jlr200

Cited by 66 publications

(80 citation statements)

References 44 publications

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“…6D) as we have previously documented (17,20). Noninduced BHK cells were unable to form such particles (data not shown).…”

Section: Development Of a Quantitative Biotinylation Assay-tosupporting

confidence: 51%

“…Freeman and co-workers (10) demonstrated that in ABCA1-expressing 293 cells, the apoA-I-ABCA1 complex has a half-life of less than 30 min and that 50% of the cell-associated apoA-I is lipidated within 40 min. Similarly, we demonstrated previously (17) that in HepG2 cells and fibroblasts, formation of nascent particles occurs within 1 h at 37°C and presently (Table 1) that the kinetics of dissociation from the ICCs is rapid, with a halflife of 20 min. As such, a large number of lipidation cycles will have occurred within 2 h of incubation and, in doing so, will have reduced the lipid burden of the cell.…”

Section: Discussionmentioning

confidence: 81%

“…rHDL particles were further concentrated by ultrafiltration (spiral ultrafiltration cartridge, molecular weight cutoff 50,000, Amicon) to discard any lipid-free apoA-I or proteolytic peptides. ApoA-I-lipid complex formation was verified by analysis with two-dimensional PAGGE as we have described previously (17).…”

Section: Methodsmentioning

confidence: 99%

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Quantitative Analysis of ABCA1-dependent Compartmentalization and Trafficking of Apolipoprotein A-I

Hassan¹,

Bailey²,

Lee

et al. 2008

Journal of Biological Chemistry

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The molecular mechanisms underlying the apoA-I/ABCA1 endocytic trafficking pathway in relation to high density lipoprotein (HDL) formation remain poorly understood. We have developed a quantitative cell surface biotinylation assay to determine the compartmentalization and trafficking of apoA-I between the plasma membrane (PM) and intracellular compartments (ICCs). Here we report that 125 I-apoA-I exhibited saturable association with the PM and ICCs in baby hamster kidney cells stably overexpressing ABCA1 and in fibroblasts. The PM was found to have a 2-fold higher capacity to accommodate apoA-I as compared with ICCs. Overexpressing various levels of ABCA1 in baby hamster kidney cells promoted the association of apoA-I with PM and ICCs compartments. The C-terminal deletion of apoA-I ⌬(187-243) and reconstituted HDL particles exhibited reduced association of apoA-I with both the PM and ICCs. Interestingly, cell surface biotinylation with a cleavable biotin revealed that apoA-I induces ABCA1 endocytosis. Such endocytosis was impaired by naturally occurring mutations of ABCA1 (Q597R and C1477R). To better understand the role of the endocytotic pathway in the dynamics of the lipidation of apoA-I, a pulse-chase experiment was performed, and the dissociation (re-secretion) of 125 I-apoA-I from both PM and ICCs was monitored over a 6-h period. Unexpectedly, we found that the time required for 50% dissociation of 125 I-apoA-I from the PM was 4-fold slower than that from ICCs at 37°C. Finally, treatment of the cells with phosphatidylcholine-specific phospholipase C, increased the dissociation of apoA-I from the PM. This study provides evidence that the lipidation of apoA-I occurs in two kinetically distinguishable compartments. The finding that apoA-I specifically mediates the continuous endocytic recycling of ABCA1, together with the kinetic data showing that apoA-I associated with ICCs is rapidly re-secreted, suggests that the endocytotic pathway plays a central role in the genesis of nascent HDL.

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“…6D) as we have previously documented (17,20). Noninduced BHK cells were unable to form such particles (data not shown).…”

Section: Development Of a Quantitative Biotinylation Assay-tosupporting

confidence: 51%

Section: Discussionmentioning

confidence: 81%

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Quantitative Analysis of ABCA1-dependent Compartmentalization and Trafficking of Apolipoprotein A-I

Hassan¹,

Bailey²,

Lee

et al. 2008

Journal of Biological Chemistry

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“…The samples (300 l) were loaded on a 1.5-mm-thick 0.75% agarose gel prepared in Tris-glycine buffer, pH 8.3, using 10% polyacrylamide to seal the bottom of the gel as described previously (18). The second dimension, a 4 -22.5% polyacrylamide nondenaturing concave gradient gel, was run at 4°C for 24 h at 10 mA/gel, as previously described (19). After transfer (30 mA, 24 h, 4°C) onto nitrocellulose membranes, molecular weight markers were revealed by Ponceau S. The membranes were either directly revealed by autoradiography or processed for immunoblotting.…”

Section: Methodsmentioning

confidence: 99%

ATP-binding Cassette A1-mediated Lipidation of Apolipoprotein A-I Occurs at the Plasma Membrane and Not in the Endocytic Compartments

Denis¹,

Landry²,

Zha

2008

Journal of Biological Chemistry

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ATP-binding cassette transporter (ABC) A1 is required for the lipidation of apolipoprotein A-I to generate high density lipoprotein (HDL). This process is proposed to occur through a retro-endocytosis pathway in which apoA-I internalizes with ABCA1 and generates HDL from the endosomal compartments before resecretion. The aim of this study was to determine the route of apoA-I endocytosis and whether endocytosis contributes to HDL biogenesis. Using confocal microscopy, we found that internalized apoA-I only transiently colocalized with transferrin, a retro-endocytosis marker. Instead, apoA-I perfectly colocalized with a bulk phase uptake marker (fluorescein isothiocyanate-dextran) and, at later time points, with LysoTracker in several cell models including macrophages, fibroblasts, and baby hamster kidney cells. ABCA1 colocalized poorly with internalized apoA-I. To determine the contribution of internalized apoA-I to HDL biogenesis, we specifically removed apoA-I from the cell surface and analyzed the fate of internalized apoA-I. We found that 23% of cell-associated apoA-I was internalized at steady state. Of internalized apoA-I, only 20% was converted to HDL, and the rest was degraded, consistent with a lysosomal destination. We also found that apoA-I was released approximately five times faster from the plasma membrane than from the intracellular compartments. From these kinetic parameters, we estimated that ϳ5.6% of apoA-I that interacts with cells is degraded and that internalized apoA-I contributes to ϳ1.4% of total HDL production. We also found that blocking endocytosis with sucrose or cytochalasin D did not decrease cholesterol efflux or HDL biogenesis. We therefore conclude that the plasma membrane is the main platform where ABCA1-mediated lipidation of apoA-I occurs.

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“…However, larger sized rHDL containing 3 and 4 molecules of apoA-I can be formed in the presence of cholesterol both by DMPC solubilization [28][29][30] and by cholate dialysis methods [31,32]. Incubating apoA-I with cells expressing ABCA1 also generates numerous discrete particles that vary in size (diameters from ~ 90-200 Å) and number of apoA-I molecules (from 2 to 8 molecules) [25,26,33,34]. Thus, ABCA1 formed nascent HDL and cholesterol-rich rHDL demonstrate size speciation and apoA-I structural organization that are different form the well studied rHDL containing 2 molecules of apoA-I.…”

Section: Introductionmentioning

confidence: 99%

Cholesterol is a determinant of the structures of discoidal high density lipoproteins formed by the solubilization of phospholipid membranes by apolipoprotein A-I

Massey

Pownall

2008

Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biolog

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Formation of discoidal high density lipoproteins (rHDL) by apolipoprotein A-I (apoA-I) mediated solubilization of dimyristoyl phosphatidylcholine (DMPC) multilamellar vesicles (MLV) was dramatically affected by bilayer cholesterol concentration. At a low ratio of DMPC/apoA-I (2 mg DMPC/mg apoA-I, 84/1 mol/mol), sterols (cholesterol, lathosterol, and β-sitosterol) that form ordered lipid phases increase the rate of solubilization similarly, yielding rHDL with similar structures. By changing the temperature and sterol concentration, the rates of solubilization varied almost 3 orders of magnitude; however, the sizes of the rHDL were independent of the rate of their formation and dependent upon the bilayer sterol concentration. At a high ratio of DMPC/apoA-I (10/1 mg DMPC/mg apoA-I, 420/1 mol/mol), changing the temperature and cholesterol concentration yielded rHDL that varied greatly in size, phospholipid/protein ratio, mol% cholesterol, and number of apoA-I molecules per particle. rHDL were isolated that had 2, 4, 6, and 8 molecules of apoA-I per particle, mean diameters of 117, 200, 303, and 396 Å, and a mol% cholesterol that was similar to the original MLV. Kinetic studies demonstrated the different sized rHDL are formed independently and concurrently. The rate of formation, lipid composition, and three-dimensional structures of cholesterol-rich rHDL are dictated primarily by the original membrane phase properties and cholesterol content. The size speciation of rHDL and probably nascent HDL formed via the activity of the ABCA1 lipid transporter are mechanistically linked to the cholesterol content of the membranes from which they were formed.

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Biogenesis and speciation of nascent apoA-I-containing particles in various cell lines

Cited by 66 publications

References 44 publications

Quantitative Analysis of ABCA1-dependent Compartmentalization and Trafficking of Apolipoprotein A-I

Quantitative Analysis of ABCA1-dependent Compartmentalization and Trafficking of Apolipoprotein A-I

ATP-binding Cassette A1-mediated Lipidation of Apolipoprotein A-I Occurs at the Plasma Membrane and Not in the Endocytic Compartments

Cholesterol is a determinant of the structures of discoidal high density lipoproteins formed by the solubilization of phospholipid membranes by apolipoprotein A-I

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