Retroendocytosis pathway of ABCA1/apoA‐I contributes to HDL formation

Azuma, Yuya; Takada, Mie; Shin, Hye‐Won; Kioka, Noriyuki; Nakayama, Kazuhisa; Ueda, Kazumitsu

doi:10.1111/j.1365-2443.2008.01261.x

Cited by 67 publications

(71 citation statements)

References 40 publications

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“…Because the expression levels of ABCA1 were kept very low (<0.3 copies/μm 2 ) for singlemolecule imaging in the present study, we observed very limited numbers of oligomer-like bright spots before apoA-I application. The interconversions of ABCA1 between dimers and monomers observed here occurred on the PM, likely to be independent of the endocytic pathway (30,31). The observation that an ATPasedeficient ABCA1 mutant, ABCA1-MM, which is unable to generate HDL, did not form dimers or show immobilization on the PM implies an obvious medical impact that dimerization of ABCA1 is physiologically critical.…”

Section: Discussionmentioning

confidence: 73%

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ABCA1 dimer–monomer interconversion during HDL generation revealed by single-molecule imaging

Nagata

Nakada

Kasai

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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The generation of high-density lipoprotein (HDL), one of the most critical events for preventing atherosclerosis, is mediated by ATPbinding cassette protein A1 (ABCA1). ABCA1 is known to transfer cellular cholesterol and phospholipids to apolipoprotein A-I (apoA-I) for generating discoidal HDL (dHDL) particles, composed of 100-200 lipid molecules surrounded by two apoA-I molecules; however, the regulatory mechanisms are still poorly understood. Here we observed ABCA1-GFP and apoA-I at the level of single molecules on the plasma membrane via a total internal reflection fluorescence microscope. We found that about 70% of total ABCA1-GFP spots are immobilized on the plasma membrane and estimated that about 89% of immobile ABCA1 molecules are in dimers. Furthermore, an ATPase-deficient ABCA1 mutant failed to be immobilized or form a dimer. We found that the lipid acceptor apoA-I interacts with the ABCA1 dimer to generate dHDL and is followed by ABCA1 dimermonomer interconversion. This indicates that the formation of the ABCA1 dimer is the key for apoA-I binding and nascent HDL generation. Our findings suggest the physiological significance of conversion of the ABCA1 monomer to a dimer: The dimer serves as a receptor for two apoA-I molecules for dHDL particle generation.membrane protein | transporter P lasma high-density lipoprotein (HDL) is critical for preventing coronary artery disease (1). A member of the ATPdependent transporter family of ABC proteins, ATP-binding cassette protein A1 (ABCA1) initiates the generation of discoidal HDL (dHDL), a bilayer fragment consisting of 100-200 lipids wrapped by two molecules of apolipoprotein A-I (apoA-I) (2-4), by exporting cholesterol and phospholipids to lipid-free apoA-I in serum (5). More than 70 mutations have been identified in the ABCA1 gene. Indeed, mutations in ABCA1 lead to Tangier disease, which is characterized by plasma HDL deficiency (6-10). ABCA1 has two large extracellular domains (ECDs), and the two intramolecular disulfide bonds between them are necessary for apoA-I binding and HDL formation (11-13) (Fig. 1A). Two pieces of evidence suggest that apoA-I interacts with a specific conformation of the ECDs in an ATP-dependent manner: Chemical cross-linkers can cross-link apoA-I with ABCA1, and ATPasedeficient ABCA1 mutants fail to mediate apoA-I binding and crosslinking. However, the importance of direct binding of ABCA1-apoA-I in HDL formation is still controversial and, furthermore, how a dHDL particle containing two molecules of apoA-I is formed from lipid-free apoA-I monomers and membrane lipids is unknown.To address these issues, we performed single-molecule fluorescence imaging (14, 15) of ABCA1 and apoA-I on the plasma membrane (PM) via a total internal reflection fluorescence (TIRF) microscope in living cells. We examined the dynamic behaviors of ABCA1 as well as the interaction of ABCA1 with apoA-I, and found that ABCA1 forms an immobile dimer on the PM; the ABCA1 dimer then dissociates into diffusing monomers upon interaction with apoA-I, finally gen...

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Section: Discussionmentioning

confidence: 73%

“…The dye:protein ratio was calculated to be between 2.5 and 5.0 by measuring the absorbance at 280 and 601 nm. Alexa546-apoA-I was prepared as previously described (30).…”

Section: Methodsmentioning

confidence: 99%

ABCA1 dimer–monomer interconversion during HDL generation revealed by single-molecule imaging

Nagata

Nakada

Kasai

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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“…Substitution of the C-terminal valine in the PDZ binding motif (13) Co-localization of LXR␤ and ABCA1 on the Plasma Membrane-To better characterize the interaction of LXR␤ with WT and mutant ABCA1, we examined the localization of ABCA1(207HA), a construct bearing an HA tag between residues 207 and 208 within the first extracellular domain, by confocal microscopy. Insertion of this HA tag does not affect apoA-I binding (22,23), cholesterol efflux (22,23), internalization (22,23), or the interaction with LXR␤ (data not shown). HEK293 cells stably expressing FLAG-LXR␤ were transiently transfected with ABCA1(207HA) (Fig.…”

Section: Leucine Residues Of Abca1 Mediate Its Interaction Withmentioning

confidence: 99%

Liver X Receptor β (LXRβ) Interacts Directly with ATP-binding Cassette A1 (ABCA1) to Promote High Density Lipoprotein Formation during Acute Cholesterol Accumulation

Hozoji-Inada

Munehira

Nagao

et al. 2011

Journal of Biological Chemistry

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Cells have evolved multiple mechanisms for maintaining cholesterol homeostasis, and, among these, ATP-binding cassette protein A1 (ABCA1)-mediated cholesterol efflux is highly regulated at the transcriptional level through the activity of the nuclear receptor liver X receptor (LXR). Here, we show that in addition to its well defined role in transcription, LXR␤ directly binds to the C-terminal region ( Disruption of cellular cholesterol homeostasis can lead to a variety of pathological conditions, including cardiovascular disease (1). ATP-binding cassette protein A1 (ABCA1), 2 an important regulator of cholesterol homeostasis, mediates the release of cellular excess free cholesterol and phospholipids to apolipoprotein A-I (apoA-I), an extracellular acceptor circulating in plasma, to form high density lipoprotein (HDL) (2-5). HDL formation is the only pathway through which excess cholesterol can be eliminated from nonhepatic cells. Defects in ABCA1 cause Tangier disease (6 -8), a condition in which patients have a near absence of circulating HDL, prominent cholesterol-ester accumulation in tissue macrophages, and premature atherosclerotic vascular disease (1, 9).The ABCA1-mediated release of cholesterol is highly regulated at the transcriptional level. When excess cholesterol accumulates in cells, intracellular concentrations of oxysterols increase and activate liver X receptor (LXR), which, in turn, stimulates ABCA1 gene transcription and increased expression of ABCA1 with associated elimination of excess cholesterol (10 -12). However, cholesterol is required for cell function and proliferation, and the intracellular cholesterol concentration must be maintained within a narrow range. Consequently, ABCA1-mediated cholesterol release is also regulated at the post-translational level. Several proteins, including syntrophins (13, 14), JAK2 (15), and LXR␤ (16), have been reported to interact with ABCA1 and modulate its degradation and function. However, the precise mechanism(s) by which ABCA1 activity is regulated post-translationally remains unclear.We previously reported (16) that a fraction of cytosolically localized LXR␤ may interact with ABCA1 on the plasma membrane and modulate the function of ABCA1. In WI-38 and THP-1 cells, endogenous LXR␤ interacts with ABCA1 under conditions in which LXR ligands do not accumulate, i.e. when cholesterol is not in excess. LXR␤ suppresses ABCA1-mediated cholesterol efflux. However, the mechanism by which LXR␤ suppresses ABCA1 functions was not clear. In this study, we identified two leucine residues in the C-terminal region of ABCA1 responsible for the interaction with LXR␤ and showed that LXR␤ interaction suppresses ATP binding to ABCA1 and thereby keeps ABCA1 standby on the plasma membrane for acute cholesterol accumulation. EXPERIMENTAL PROCEDURESMaterials-The LXR ligand TO901317, 22(R)-hydroxycholesterol, 25-hydroxycholesterol, and Alexa Fluor 546 succinimidyl esters were purchased from Cayman Co., Sigma, Stelaroids, and Molecular Probes, respectively. A cholesterol oxidas...

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“…This peptide insertion had no effect on the subcellular localization or the function of ABCA1. 17) The relative surface expression of each mutant was calculated by dividing HA immunofluorescence by GFP fluorescence, and was compared with that of wild-type ABCA1 ( Fig. 1(B)).…”

Section: Resultsmentioning

confidence: 99%

Position 834 in TM6 plays an important role in cholesterol and phosphatidylcholine transport by ABCA1

Itoh

Nagao

Kimura

et al. 2015

Bioscience, Biotechnology, and Biochemistry

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ATP-binding cassette protein A1 (ABCA1) plays a key role in eliminating excess cholesterol from peripheral cells by generating nascent high-density lipoprotein (HDL). However, it remains unclear whether both phospholipids and cholesterol are directly loaded onto apolipoprotein A-I (apoA-I) by ABCA1. To identify the amino acid residues of ABCA1 involved in substrate recognition and transport, we applied arginine scan mutagenesis to residues L821–E843 of human ABCA1 and predicted the environment to which each residue is exposed. The relative surface expression of each mutant suggested that residues L821–E843 pass through the plasma membrane as TM6, and the four residues (S826, F830, L834, and V837) of TM6 are exposed to the hydrophilic internal cavity of ABCA1. Furthermore, we showed that L834 is critical for the function of ABCA1.

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Retroendocytosis pathway of ABCA1/apoA‐I contributes to HDL formation

Cited by 67 publications

References 40 publications

ABCA1 dimer–monomer interconversion during HDL generation revealed by single-molecule imaging

ABCA1 dimer–monomer interconversion during HDL generation revealed by single-molecule imaging

Liver X Receptor β (LXRβ) Interacts Directly with ATP-binding Cassette A1 (ABCA1) to Promote High Density Lipoprotein Formation during Acute Cholesterol Accumulation

Position 834 in TM6 plays an important role in cholesterol and phosphatidylcholine transport by ABCA1

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