© The Authors Journal compilation © 2011 Biochemical Society
Essays Biochem. (2011) 50, 265–290; doi:10.1042/BSE0500265
14
Lipid transport by
mammalian ABC proteins
Faraz Quazi and Robert S. Molday1
Department of Biochemistry and Molecular Biology, University of
British Columbia, Vancouver, BC, Canada, V6T 1Z3
Abstract
ABC (ATP‑binding cassette) proteins actively transport a wide variety of
substrates, including peptides, amino acids, sugars, metals, drugs, vitamins
and lipids, across extracellular and intracellular membranes. Of the 49 human
ABC proteins, a significant number are known to mediate the extrusion
of lipids from membranes or the flipping of membrane lipids across the
bilayer to generate and maintain membrane lipid asymmetry. Typical
lipid substrates include phospholipids, sterols, sphingolipids, bile acids
and related lipid conjugates. Members of the ABCA subfamily of ABC
transporters and other ABC proteins such as ABCB4, ABCG1 and
ABCG5/8 implicated in lipid transport play important roles in diverse
biological processes such as cell signalling, membrane lipid asymmetry,
removal of potentially toxic compounds and metabolites, and apoptosis.
The importance of these ABC lipid transporters in cell physiology is
evident from the finding that mutations in the genes encoding many of
these proteins are responsible for severe inherited diseases. For example,
mutations in ABCA1 cause Tangier disease associated with defective efflux
of cholesterol and phosphatidylcholine from the plasma membrane to the
lipid acceptor protein apoA1 (apolipoprotein AI), mutations in ABCA3
cause neonatal surfactant deficiency associated with a loss in secretion of the
1To
whom correspondence should be addressed (email molday@interchange.
ubc.ca).
265
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Essays in Biochemistry volume 50 2011
lipid pulmonary surfactants from lungs of newborns, mutations in ABCA4
cause Stargardt macular degeneration, a retinal degenerative disease linked
to the reduced clearance of retinoid compounds from photoreceptor cells,
mutations in ABCA12 cause harlequin and lamellar ichthyosis, skin diseases
associated with defective lipid trafficking in keratinocytes, and mutations in
ABCB4 and ABCG5/ABCG8 are responsible for progressive intrafamilial
hepatic disease and sitosterolaemia associated with defective phospholipid
and sterol transport respectively. This chapter highlights the involvement
of various mammalian ABC transporters in lipid transport in the context of
their role in cell signalling, cellular homoeostasis, apoptosis and inherited
disorders.
Introduction
The hydrophobic lipid bilayer separates eukaryotic cells from their
extracellular environment and further compartmentalizes these cells into
distinct organelles. Assembly and maintenance of the various cellular
membranes requires translocation of lipids from one leaflet of the bilayer
to the opposing leaflet. The plasma membrane, endosomes and lysosomes
depend completely on lipid transport and synthesis from other organelles,
in particular from the ER (endoplasmic reticulum). The lipid composition
of separate leaflets of the membrane bilayer generally show a clear
asymmetric arrangement, with the majority of the aminophospholipids
PS (phosphatidylserine) and PE (phosphatidylethanolamine), typically
present in the inner, cytoplasmic, leaflet, and PC (phosphatidylcholine),
SM (sphingomyelin) and glycolipids predominantly, if not exclusively
localized in the outer, exoplasmic, leaflet. The plasma membranes, Golgi and
endosomal membranes, unlike the membranes of the ER, display high lipid
asymmetry [1]. This bilayer asymmetry is less governed by the size, charge
and polarity of the headgroup, but more so by ATP‑dependent protein
translocators. This type of distribution influences important physiological
functions such as cell viability, membrane fusion, cell–cell recognition, and
protein function and regulation.
Maintenance of lipid asymmetry is accomplished by integral membrane
transporters that specifically flip (out‑to‑in translocation), flop (in‑to‑out
translocation) or scramble lipids across the bilayer (Figure 1). Lipid flippases
and floppases are ATP‑dependent membrane proteins that maintain trans‑
bilayer distribution of phospholipids by translocating specific phospholipid
species from the cytoplasmic to the exoplasmic leaflet of the bilayer and vice
versa respectively. P4 ATPases are a subclass of P‑type ATPases that func‑
tion as phospholipid flippases, whereas ABC (ATP‑binding cassette) trans‑
porters have been generally shown to act as lipid floppases [2,3]. In contrast,
asymmetric lipid distribution can be undone by the Ca2+‑dependent bidirec‑
tional activities of scramblases that tend to act with a low lipid headgroup
specificity.
© The Authors Journal compilation © 2011 Biochemical Society
F. Quazi and R.S. Molday
267
Figure 1. Types of protein involved in the generation and maintenance of membrane
lipid asymmetry
Lipids are translocated across the lipid bilayer by flippases (active translocation to the cytoplas‑
mic leaflet), floppases (active translocation to the exoplasmic leaflet), and scramblases (bidirec‑
tional energy‑independent translocation through a Ca2+‑dependent mechanism).
To date, 49 mammalian ABC transporters have been identified in the
human genome. These have been organized into seven subfamilies: ABCA–
ABCG. In most cases, these ABC proteins are either full‑transporters consist‑
ing of a single polypeptide, as exemplified by ABCA1, or half‑transporters
consisting of homo‑ or hetero‑dimers such as ABCG1 (see Figure 3) [4]. In
either case, the ABC transporters contain at least two TMDs (transmembrane
domains) with multiple membrane‑spanning segments and two NBDs (nucle‑
otide‑binding domains). A significant number of mammalian ABC proteins
are involved in the transport and regulation of steroids/steroid conjugates and
phospho‑/glyco‑/sphingo‑lipids and, as such, they participate in lipid traffick‑
ing, lipid asymmetry and lipid homoeostasis across cellular membranes [5,6]
(Figure 2 and Table 1).
Genetic defects in six ABC transporters that function in lipid trafficking
in cells have been linked to a variety of severe human diseases [7] (Table 1).
For instance, inherited defects in ABCA1 which mediate the efflux of cho‑
lesterol and phospholipids from cells cause Tangier disease [8], defects in
ABCA3, a transporter of lipid molecules in the lung, is associated with neona‑
tal surfactant deficiency in newborns [9], mutations in ABCA4, a retinal-PE
transporter, are linked to a variety of retinal degenerative diseases including
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Figure 2. Overview of ABC transporters involved in lipid efflux
Schematic representation of ABC transporters, known acceptors and the direction of transport.
Black arrows represent transport direction at the membrane bilayer. Cer, ceramide; Chol, chol‑
esterol; LPA, lysophosphatidic acid.
Stargardt macular degeneration [10], defects in ABCG5/ABCG8, transport‑
ers of nutritional plant sterols, are linked to sitosterolaemia [11], and defects
in ABCB4 are involved in the transport of PC on to bile micelles cause PFIC
(progressive familial intrahepatic cholestasis) [12].
This chapter presents a snapshot of the current position of the field by
focusing on a varied selection of mammalian ABC transporters, classes of lip‑
ids which are transported, and molecular mechanisms underlying lipid trans‑
port (Figure 2). The role of lipids after membrane redistribution in response to
cellular events and their underlying disorders are also reviewed.
Mechanism of lipid efflux
Two general models have been proposed to explain the transport of lipid
substrates across membranes: ‘hydrophobic vacuum cleaner’ and ‘flippase’
models [13]. In the hydrophobic vacuum cleaner model, lipid substrates
diffuse into the membrane bilayer and are subsequently extruded from
a central channel of the transporter into the extracellular space in an
ATP‑dependent process [14]. This model typically explains the drug‑efflux
action observed for multidrug‑resistance transporters. The flippase model, on
the other hand, is used to explain the translocation of lipids, most commonly
© The Authors Journal compilation © 2011 Biochemical Society
Chol, SM, PC
Lung, pancreas, brain,
heart
Retinal photoreceptors N‑retinylidene‑PE
ABCA3
ABCA4
ABCG5/
ABCG8
ABCG4
ABCG1
ABCG2
ABCC1
ABCA12
ABCB1
(MDR1)
ABCB4
Universal
Placenta, breast, liver,
gastrointestinal tract
Macrophage, brain, eye, Chol
spleen, liver
Liver, gastrointestinal
Plant sterols, Chol
tract
LTC4, GlcCer, SM, PC, S1P, GSH, UGT, steroid
conjugates
Chol, PC, SM, 7β‑OH‑chol, 7‑oxo‑chol
PC, PS
GlcCer
SM, GlcCer, sphingoid bases, PC, PS, PE, Chol, PAF,
corticosteroids, androgens, oestrogens, progestins
PC
Chol, PS, PC, Cer
Chol, SM, PS, PE, Cer
Brain
ABCA2
Brain, skin,
myelolymphatic system
Keratinocytes
Placenta, brain, liver,
kidneys
Liver, bile, canilicular
membrane, placenta
Placenta
Chol, PS, SM, PC, S1P, 25‑OH‑chol
Universal
ABCA1
ABCA7
Lipids transported
Tissue expression
Gene
Genetic disorders
HDL bile salts
HDL
ApoAI
(Cytosolic)
[9,35,53]
[34,52]
[30,83,90]
Reference(s)
Sitosterolaemia
[12]
PFIC
[11,44]
[29]
[40,42,50,96]
[88]
[29,49,78,95]
[80]
[45–48,68]
Harlequin ichthyosis
Stargardt macular degeneration, [54,55,60]
cone–rod dystrophy, retinitis
pigmentosa, age‑related
macular degeneration
[36,81]
Fetal/ neonatal surfactant lung
deficiency
Apolipoproteins AI, AII, Tangier disease, familial
E, CI, CII, CIII and AIV hypoalphalipoproteinaemia
Acceptors
lysophosphatidic acid; LTC4, leukotriene C4; PAF, platelet‑activating factor; Sph, sphingosine; UGT, glucuronide.
Cer, ceramide; Chol, cholesterol; 7β‑OH‑chol, 7β‑hydroxycholesterol; 25‑OH‑chol, 25‑hydroxycholesterol; 7‑oxo‑chol, 7‑oxocholesterol; GSH, glutathione; LPA,
Table 1. Mammalian ABC transporters, lipid substrates and associated human genetic disorders
F. Quazi and R.S. Molday
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phospholipids, from the inner to outer leaflet of biological membranes [15].
Additionally, for many lipophilic substrates, the docking of an acceptor
protein creates energetically favourably conditions required to promote
dissociation of the lipid from the donor membrane, thereby facilitating
its efflux from the cell. In this composite model, the transporter relocates
the substrate to a region of intermediate hydrophobicity adjacent to the
transporter in the extracellular leaflet of the plasma membrane [16]. This
places the substrate in an environment which energetically favours its
binding and removal by an acceptor molecule/docking protein such as apoA1
(apolipoprotein AI).
Role of ABC transporters in cholesterol homoeostasis and
other sterol export
ABCA1 as a cholesterol transporter
Cells in peripheral tissues produce all of the cholesterol needed for cellular
homoeostasis. However, the liver is the only organ that is capable of degrading
cholesterol. Therefore cholesterol must be transported through the blood to
the liver for processing, degradation and secretion into bile, a pathway termed
RCT (reverse cholesterol transport). Because cholesterol is a highly insoluble
molecule, it must be packaged and transported by special particles in the
plasma called lipoproteins. HDLs (high‑density lipoproteins) are an essential
determinant responsible for movement of most cholesterol from peripheral
tissues through the blood back to the liver.
ABCA1, a member of the ABCA subfamily of ABC transporters, medi‑
ates the transport of excess cholesterol from cells to HDL apolipoproteins
such as apoA1 [17]. Loss of function results in Tangier disease, a rare lipid
disorder characterized by very low levels of HDLs and apoA1 [18]. Tangier
patients accumulate high concentrations of cholesteryl esters in macrophages
and various tissues, including liver, intestine, tonsils, spleen, lymph nodes and
neuronal Schwann cells. Accumulation of excess cholesterol causes athero‑
sclerotic cardiovascular disease, and may contribute to the early onset of AD
(Alzheimer’s disease) [19] and renal dysfunction [20].
Abca1‑knockout mouse models exhibit HDL deficiency and reduced
cellular cholesterol efflux activity [21]. On the other hand, overexpression of
ABCA1 increases plasma HDL levels and protects against atherosclerosis in
animal models [22]. Hepatic deletion of ABCA1 dramatically reduces apoA1
and HDL levels. However, selective knockout of ABCA1 in macrophages
has little influence on the plasma concentration of HDL, but leads to excess‑
ive accumulation of cholesterol esters. Thus, although ABCA1 in macro‑
phages is not a major determinant of plasma HDL levels, it is a crucial factor
in the prevention of excessive cholesterol accumulation in macrophages of
the arterial walls.
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F. Quazi and R.S. Molday
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Interaction of ABCA1 and apolipoproteins
ABCA1 localizes to the plasma membrane and intracellular compartments of
cells, where it can facilitate the transport of lipids to either cell‑surface‑bound
or internalized apolipoproteins (Figure 3). Overexpression of ABCA1
increases cholesterol efflux and apoA1 binding to the cell surface.
Direct interactions between apoA1 and ABCA1 have been confirmed in
numerous studies using cross‑linking, immunoprecipitation, radiolabelling
and biotinylation techniques [23–25]. Although cholesterol transport to other
lipoproteins such as apoE (apoplipoprotein E) may also occur, apoA1 is the
most physiologically relevant lipid acceptor for ABCA1 because of its high
abundance and its role in forming nascent HDL particles. It contains eight
22‑amino‑acid and two 11‑amino‑acid tandem amphipathic α‑helical domains.
Further examination of apoA1 domain mutants shows that the C‑terminal
region is important for cross‑linking with the large exocytoplasmic domains
of ABCA1. This binding is not required for the intrinsic lipid transport activ‑
ity of ABCA1, as forced overexpression of ABCA1 increases lipid domains
on the cell surface in the absence of lipoproteins [26]. Instead, binding appears
to facilitate removal of these lipids, perhaps by targeting apolipoproteins to
ABCA1‑generated lipid domains. Taken together, a two‑step model is pro‑
posed: (i) apoA1 binds to ABCA1 by different combinations of apoA1 helices
with the exocytoplasmic domain of ABCA1; and (ii) apoA1 is lipidated and
nascent HDL dissociates [27].
Figure 3. Principal cholesterol‑efflux pathway in macrophages
ABCA1 and ABCG1 mediate the transport of cholesterol to the extracellular acceptor particle,
apoA1. ApoE and other lipoproteins may also serve as acceptors for ABCA1‑mediated chol‑
esterol efflux. Phospholipids and cholesterol are loaded on to surface‑bound apoA1 by ABCA1
or HDL by ABCG1. HDL maturation is mediated by lecithin–cholesterol acyltransferase (LCAT,
not shown). Oxidized cholesterol derivatives or oxysterols such as 27‑hydroxycholesterol or
7‑oxocholesterol can serve as ABCA1 or ABCG1 substrates or endogenous ligands of the LXR
regulating ABCA1 and ABCG1 transcription.
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However, other studies indicate that only 10% of the cell‑associated
apoA1 can be cross‑linked to ABCA1 [28]. In addition, ABCA1 significantly
decreases plasma membrane rigidity independent of its interactions with apoli‑
poproteins. ABCA1 can also transport SM and PC, and function co‑ordinately
in removing these lipids together with cholesterol from peripheral cells [29].
Taken together, a second model is suggested whereby ABCA1 enriches the
outer leaflet with cholesterol and phospholipids, generates high‑affinity bind‑
ing sites for apoA1 and facilitates apoA1 docking and cholesterol uptake with‑
out direct contact [30]. This may promote the formation of apoA1‑containing
exovesicles and solubilization of apoA1‑bound phospholipids and cholesterol
for incorporation into nascent HDL.
Evidence of apoA1 internalization via receptor‑mediated endocytosis also
suggests a third model, whereby cholesterol and phospholipids are accumu‑
lated internally before secretion [31]. However, recent evidence suggests that
this retroendocytotic pathway contributes less than 2% of lipidated HDL,
reinforcing the previous models of plasma membrane being the primary site of
apoA1 lipidation [32].
Although there is strong evidence that ABCA1 plays an essential role in
the efflux of cholesterol from cells, the mechanism by which cholesterol is
transported across the lipid bilayer remains to be determined. To date, there
is no evidence for the direct binding of cholesterol to ABCA1 or the activation
of the ATPase activity of ABCA1 by cholesterol. Furthermore, the transport of
cholesterol by purified ABCA1 reconstituted into a non‑cellular system such
as liposomes has not been reported. The molecular mechanism by which
ABCA1 mediates cholesterol transport remains an area of considerable impor‑
tance in understanding cholesterol transport and metabolism.
ABCA1 and cell signalling
The interaction of apolipoproteins with ABCA1‑expressing cells
activates signalling molecules that modulate ABCA1 expression and lipid
transport activity. These include JAK2 (Janus kinase 2), protein kinase C,
Cdc42 and other protein kinases [33]. Although it remains to be shown
whether these activation steps involve direct binding of apolipoproteins to
ABCA1, these findings suggest that apolipoprotein behaves like a ligand
capable of interacting with ABCA1 as a receptor and activating intracellular
signalling pathways.
Activation of JAK2, a tyrosine kinase, is the initiating step in downstream
signalling pathways. Inhibiting JAK2 activity has no effect on the intrinsic
lipid transport function of ABCA1, but it does reduce the apolipoprotein
binding to ABCA1 required for removal of translocated lipids [33]. The inter‑
action of apoA1 with ABCA1‑expressing cells stimulates autophosphorylation
of JAK2, thus generating the active form of JAK2 that in turn phosphorylates
its target proteins [33]. This suggests that apolipoproteins influence their
own interactions with ABCA1 by activating JAK2, which in turn increases
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F. Quazi and R.S. Molday
273
apolipoprotein binding to ABCA1 required for lipid removal. This type of
JAK2‑mediated feedback mechanism has not been described for any other
transporter to date.
Other ABCA subfamily members
Overexpression of ABCA2 in CHO (Chinese‑hamster ovary) cells reduces
the trafficking of LDL (low‑density lipoprotein)‑derived cholesterol to the
ER [34]. ABCA3, which is predominantly expressed in lungs, has also been
shown to remove cholesterol in an alveolar cell line [35]. However, direct
translocation of cholesterol to apolipoproteins has not been demonstrated for
these ABC proteins, so the mechanistic explanation for these observations is
not clear.
ABCA7 shows the highest homology with ABCA1 and in fact mim‑
ics ABCA1 in mediating the production of HDL from cellular lipid
in vitro. In induced sterol uptake conditions, a notable increase in mRNA
levels of the ABCA7 gene occurs, whereas cholesterol‑depleting condi‑
tions result in down‑regulation of its expression [36]. ABCA7 generates
mostly small cholesterol‑poor HDL particles, unlike ABCA1, which forms
predominantly cholesterol‑rich HDL particles. Deletion of Abca7 in mouse
macrophages does not seem to affect phospholipid or cholesterol efflux
to apoA1, whereas inactivation of ABCA1 results in complete absence of
efflux [37]. To date, no disease‑causing genetic defect has been reported for
ABCA7.
It is suggested that ABCA7 plays an important role in lipid transport
within defined microenvironments. ABCA7 expression has been detected in
murine renal tubules in the apical brush border membrane [37]. The kidney
plays a major role in apoA1 catabolism, and expression of other ABC proteins
has been demonstrated in this organ. In female Abca7‑knockout mice, a sig‑
nificant reduction in visceral fat and serum cholesterol concentrations has been
reported [38]. Abca7 expression is also detected during late keratinocyte differ‑
entiation and significantly influences ceramide levels. These studies point to a
subtle role of ABCA7 in cholesterol and other lipid‑transport ­processes.
ABCG subfamily
The human ABCG subfamily contains five half‑transporters, i.e. ABCG1,
ABCG2, ABCG4, ABCG5 and ABCG8. These are unique transporters
in which the NBD is localized on the N‑terminal side of the TMD
(NBD‑TMD). All members of the ABCG subfamily, with the exception of
ABCG2/BCRP (breast cancer‑resistance protein), function as cholesterol
transporters. Four of the five mammalian ABCG subfamily members, namely
the homodimers ABCG1–ABCG1 and ABCG4–ABCG4 and the heterodimer
ABCG5–ABCG8, have been shown to have a role in transporting sterols
across membranes. ABCG1 and ABCG4 have been implicated in lipid efflux,
predominantly cholesterol and SM, from peripheral cells to HDL [29,39]. The
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ABCG5–ABCG8 heterodimer mediates the translocation of plant sterols as
well as cholesterol.
ABCG1 displays the most widespread expression pattern, and it is cur‑
rently thought to be one of the major regulators of cellular cholesterol content
in mammals [30]. ABCG1 displays many similarities to ABCA1, such as tis‑
sue expression patterns and regulatory pathways [30] (Figure 3). However, to
date, unlike ABCA1, no mutations in ABCG1 have been linked to any human
disease. Initial studies demonstrated that transient overexpression of ABCG1
increased the efflux of cellular cholesterol to specific extracellular lipid accep‑
tors that included HDL, LDL, PC vesicles and apoE complexes, but not
lipid‑free apoA1, as is the case with ABCA1 [40]. This leads to a proposal that
ABCA1 and ABCG1 may function to redistribute cholesterol pools to either
apoA1 or HDL.
ABCG4 shares 82% amino acid identity with ABCG1, so it is not surpris‑
ing that they exhibit functional similarities: overexpression of either protein
in cultured cells facilitates the efflux of cellular cholesterol to HDL, but not
lipid‑poor apoA1 [39]. Unlike ABCG1, ABCG4 expression is highly restricted
to the brain and the neural layer of the retina [41]. Indeed, a concerted model
for cholesterol efflux has been proposed for ABCA1, ABCG1 and ABCG4,
in which initial lipidation of apoA1 to nascent HDL is performed by ABCA1
followed by HDL maturation via ABCG1/ABCG4 [42].
The ABCG5 and ABCG8 obligate heterodimers are expressed in the intes‑
tines and the liver. They function to limit the absorption of plant sterols found
in our diet by transporting these sterols back into the intestinal lumen and by
facilitating efficient secretion of plant sterols and cholesterol from hepato‑
cytes into the bile. Mutations in these genes are linked to β‑sitosterolaemia, a
rare disorder caused by accumulation of cholesterol and plant sterols, even‑
tually leading to premature coronary atherosclerosis [43]. Wang et al. [44]
showed that cholesterol and sitosterol are the direct substrates of ABCG5
and ABCG8. Interestingly, acceptors for cholesterol are bile salts and not
apolipoproteins or HDL.
Sterol transport by other ABC proteins
Active redistribution of cholesterol across the cell membrane by ABCB1
[Pgp (P‑glycoprotein)] has been described. Considering the above‑mentioned
transporters, Pgp is unlikely to be a primary cholesterol transporter.
Additionally, ATP‑dependent cholesterol trafficking from the plasma
membrane to the ER has been reported for Pgp [45]. Some studies suggest that
this may be a consequence of its sphingolipid‑efflux function, since cholesterol
is known to interact with GlcCer (glucosylceramide) [46]. Recently,
a variety of other steroids have been shown to be transported by Pgp.
Overexpression of Pgp in human colon carcinoma cells reduces intracellular
levels of glucococorticoids (cortisol, dexamethasone), mineralocorticoids
(aldosterone, corticosterone) and, to a lesser extent, androgens (testosterone,
© The Authors Journal compilation © 2011 Biochemical Society
F. Quazi and R.S. Molday
275
dihydrotestosterone), oestradiol and progestins [47]. The steroid role of Pgp in
endocrine function awaits further clarification.
Role of ABC proteins in phospholipid transport
A number of ABC proteins have been shown to transport or flip
phospholipids such as PC, PE and PS. Using lipid substrates labelled with
the fluorophore 7‑nitrobenz‑2‑oxa‑1,3‑diazole in conjunction with cellular
or reconstituted systems, Pgp (ABCB1), responsible for pleiotropic drug
resistance in tumour cells, was found to transport various phospholipid
analogues, including PC and PE, as well as the sphingolipid GlcCer from
the cytoplasmic to the exoplasmic leaflet of the plasma membrane [47].
Expressed mainly in the liver, ABCB4 is primarily a floppase for PC [48].
Mutations in ABCB4 have been associated with PFIC, a disorder involving
liver inflammation and fibrosis [12]. Cholestasis is thought to result from the
toxicity of bile in which detergent bile salts are not effectively neutralized
by phospholipids, leading to bile canaliculi and biliary epithelium injuries.
PC and PS also act as substrates to for the other xenobiotic transporter
BCRP (ABCG2) and MRP1 (multidrug‑resistance protein 1) (ABCC1) [49].
Taken together, these studies support a key role of these ABC proteins in
regulating membrane composition, asymmetry, and stability via translocation
of key membrane phospholipids across cellular membranes.
ABCA1 and ABCG1 also regulate cellular phospholipid levels. Early evi‑
dence suggests that ABCA1 acts as a translocator of PC, PS and PE, whereas
ABCG1 transports only PC [25,29,50]. As discussed above, generation of new
HDL requires ABCA1‑mediated phospholipid efflux to apoA1. More recent‑
ly, missense mutations of ABCA1 within conserved residues of the Walker A
motif and selected disease mutations (in exocytoplasmic domains) demonstrate
impaired phospholipid export function, apoA1‑binding activity and JAK2
activation [51].
ABCA2, a lysosome‑associated protein, is predominantly localized in the
brain in the cell bodies of oligodendrocytes. Recently, a correlation between
ABCA2 and AD was reported from co‑localization studies with amyloid
β‑peptides and other AD markers. As AD is associated with lipid‑transport
disorders and abnormal myelination, an indirect link has been proposed for
ABCA2 in controlling lipid trafficking from the neuronal cell body to the
membrane [52].
ABCA3 is localized in the lamellar bodies of lung alveolar type II cells.
Lamellar bodies are densely packed lysosome‑like structures which store pul‑
monary surfactant, a mixture of phospholipids and proteins. Upon secretion
from lamellar bodies, pulmonary surfactants coat the airways of the lung
with a lipid‑rich monolayer thereby reducing the surface tension at the air/
liquid interface. Secretion of pulmonary surfactant represents a critical feature
in the switch of the lung from an aqueous environment at birth. Abca3‑null
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mice display low PC content in lamellar bodies of alveolar type II cells [53].
Mutations in Abca3 are associated with fatal surfactant deficiency in newborns
[9]. On the basis of these studies, ABCA3 has been implicated in the transport
of lipids and pulmonary surfactant metabolism.
ABCA4 is expressed in rod and cone outer segment disks of vertebrate
retinal photoreceptors, where it plays an important role in the removal
of retinal derivatives following the photobleaching of the photopigment
proteins rhodopsin and cone opsin [54]. Photon absorption induces the
isomerization of 11‑cis‑retinal to all‑trans‑retinal chromophore within the
opsin protein (Figure 4A). This is followed by the release of the chromo‑
phore into the disc membrane. All‑trans‑retinal can be directly reduced to
all‑trans‑retinol by retinol dehydrogenase as a key step in the regeneration
of 11‑cis‑retinal as part of the visual cycle. However, a substantial fraction
of all‑trans­‑retinal reacts with PE in disc membranes to form the Schiff base
adduct N‑retinylidene‑PE, which can be trapped on the luminal side of the
disc membrane. ABCA4 is generally thought to function in the transport of
N‑retinylidene‑PE from the lumen to the cytoplasmic leaflet of the disc mem‑
brane [55]. Dissociation of N‑retinylidene‑PE into all‑trans‑retinal and PE
enables retinol dehydrogenase to reduce all‑trans‑retinal to all‑trans‑retinol
for entry into the visual cycle. This ensures that all of the retinoid obtained
from photoexcitation is removed from disc membranes. In the absence of
ABCA4, all‑trans‑retinal and N‑retinylidene‑PE in disc membranes react to
form fluorescent di‑retinoid compounds including A2PE, which, after phago‑
cytosis of photoreceptor outer segments, is hydrolysed to A2E in RPE (retinal
pigment epithelial) cells and progressively accumulate as lipofuscin deposits
[56] (Figure 4B). A2E and related di‑retinoids are toxic, resulting in RPE and
photoreceptor cell death and a loss in vision. The accumulation of A2E in RPE
cells has been reported for individuals with Stargardt macular degeneration
linked to mutations in ABCA4 [57].
Several lines of evidence support the role of ABCA4 as a N‑retinylidene‑PE
transporter. The ATPase activity of detergent‑solubilized and reconstituted
ABCA4 is activated by retinal in the presence of PE [58,59]. Furthermore, ABCA4
immobilized on an immunoaffinity matrix binds stoichiometric amounts of
N‑retinylidene‑PE and its reduced analogue N‑retinyl‑PE with an apparent
Kd of 4–5 μM [55]. Addition of ATP (or GTP) to ABCA4 containing bound
N‑retinylidene‑PE causes the dissociation of the substrate from the protein.
Finally, Abca4‑knockout mice display light‑dependent elevated levels of PE,
N‑retinylidene‑PE and all‑trans‑retinal in the outer segments of photorecep‑
tors and A2E in RPE cells, consistent with the role of ABCA4 in the removal of
N‑retinylidene‑PE from disc membranes following photoexcitation [57,60].
ABCA12 is predominantly expressed on the skin and in the stomach and
in particular keratinocytes [61]. Mutations in the ABCA12 gene are responsible
for harlequin ichthyosis, a rare hereditary disease characterized by skin desq‑
uamations over the whole body [61]. A disorder in the keratinization process
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F. Quazi and R.S. Molday
277
Figure 4. Proposed role for ABCA4 in the transport of N‑retinylidene‑PE and the
removal of retinoids from disc membranes of photoreceptors
(A) All‑trans‑retinal released from rhodopsin after photoexcitation can be directly reduced to
all‑trans‑retinol by retinol dehydrogenase or react with PE to produce a Schiff base conjugate
N‑retinylidene‑PE (N‑ret PE). ABCA4 can bind and transport N‑retinylidene‑PE from the lumen
to the cytoplasmic side of the disc membrane. All‑trans‑retinal derived from the dissociation
of N‑retinylidene‑PE can be reduced to all‑trans‑retinol for removal from discs by the visual
cycle (not shown). (B) Chemical reactions involved in the formation of N‑retinylidene‑PE and
the production of the di‑retinoid A2PE in disc membranes in the absence of ABCA4 transport
activity. A2PE is subsequently hydrolysed to A2E upon phagocytosis of photoreceptor outer
segments by retinal pigment epithelial cells. A2E progressively accumulates as fluorescent lipo‑
fuscin deposits in retinal pigment epithelial cells, a characteristic feature of Stargardt macular
degeneration.
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results in a thick ‘armour’‑like scale layer covering the whole body. ABCA12
localizes in the lamellar granules of keratinocytes, where it may play a major
role in lipid trafficking.
Eukaryotes primarily express outward directed ABC transporters.
However, in the yeast Candida albicans, Cdr3p, a subfamily member of ABC
proteins, has been found to exhibit an inward‑directed phospholipid activity
translocating NBD‑PE, NBD‑PC and NBD‑PS [62]. This is in contrast with
its related homologues, Cdr1p and Cdr2p. Two other Saccharomyces cerevisiae
transporters, Aus1p and Pdr11p, facilitate exogenous sterol uptake when sterol
biosynthesis is compromised [63]. In humans, ABCA4 is implicated to be an
inwardly directed flippase translocating N‑retinylidene‑PE from the luminal
to the cytosolic leaflet of the outer segment disc membranes in the retina [54].
Finally, the presence of CFTR (cystic fibrosis transmembrane conductance
regulator) (ABCC7), which is not a true transporter, but a channel, is corre‑
lated with an increased uptake of signalling lipids sphingosine 1‑phosphate and
lysophosphatidic acid [64].
Members of the eicosanoid family of lipid mediators are also substrates for
ABC proteins. Leukotriene C4 is effluxed by ABCC1 (MRP1) and ABCC2
(MRP2) [65], whereas ABCC4 (MRP4) is a transporter of PG (prostaglandin)
E1 and E2 [66]. ABCC4 may also transport other related eicosonoids such
PGA2, PGF2α and thromboxane A2 [66]. The physiological importance of these
transporters with respect to eicosanoid secretion remains to be determined.
Phospholipid‑ and sterol‑transport assays
Transport of lipids by Pgp has been studied most extensively. A variety of
approaches and techniques have been used, including the efflux of lipids from
confluent monolayers of transfected kidney cells, fluorescence microscopy,
FACS analysis of NBD fluorescence, and fluorescence quenching of
reconstituted proteoliposomes. Pgp has been shown to transport short‑chain
analogues of membrane lipids across the plasma membrane, including
C6‑NBD and C8‑short chains of PC and PE, and the sphingolipids GlcCer
and SM.
Van Helvoort et al. [48] showed the translocation of lipids with stable
transfectants of pig kidney epithelial (LLC‑PK1) cells containing Pgp. To
generate C6‑NBD‑PC and C6‑NBD‑PE in cytosol, they incubated confluent
monolayers of cells and Pgp transfectants with a precursor, C6‑NBD‑PA (phos‑
phatidic acid), complexed to BSA for 3 h at 15°C (Figure 5A). C6‑NBD‑PA
partitions into the plasma membrane and is dephosphorylated to
C6‑NBD‑diacylglycerol and further converted by cells into C6‑NBD‑PC and
C6‑NBD‑PE. Vectorial transport of lipids to the cell surface is discriminated by
lowering the temperature to 15°C, at which point lipid transport by vesicular
processes is blocked. After 3 h, only a small fraction of each lipid was found
in apical BSA medium of control cells, whereas 20–25% of both C6‑NBD‑PC
and C6‑NBD‑PE was found in the apical medium of Pgp‑transfected cells,
© The Authors Journal compilation © 2011 Biochemical Society
F. Quazi and R.S. Molday
279
Figure 5. The application of NBD‑labelled lipids for lipid flippase measurements
The structural formula in the Figure illustrates the position and length of the chains. B, base
(choline in PC or ethanolamine in PE); NBD, fluorescent group; P, phosphate; asterisk, 3H
radiolabel; ‘=O’ and ‘‑OH” have been omitted for clarity. (A) Cell monolayers are incubated
with C6NBD‑PA on both sides for 3 h at 15°C. Fluorescent lipid products, C6‑NBD‑PC and
C6‑NBD‑PE, appearing at the cell surface, were depleted into the BSA‑containing medium. (B)
Dithionite reaction with reconstituted proteoliposomes containing Pgp and fluorescently labelled
NBD phospholipid. The reaction mixture is incubated at 37°C with ATP and terminated by vana‑
date addition. Fluorescence emission is monitored and dithionite is added. The sample is incu‑
bated further until a stable baseline is reached and Triton X‑100 detergent is added to make the
vesicles permeable to dithionite. The fluorescent traces involved with ATP and inhibitors enable
flippase activity determination.
which suggests that Pgp can translocate both C6‑NBD‑PC and C6‑NBD‑PE
across the apical membrane. This was confirmed by NBD fluorescence
microscopy. Translocation was also confirmed by [3H]choline radioactivity.
C6‑NBD‑PC in apical medium was 3H‑labelled, so it was derived from intra‑
cellular C6‑NBD‑PC and confirms the selective translocation of C6‑NBD‑PC
by Pgp transfectants. The continuous presence of precursor ensured synthe‑
sis of C6‑NBD‑PC and C6‑NBD‑PE in Pgp cells and was linear over 5 h at
15°C. Translocation of these fluorescence lipids was efficiently blocked by Pgp
inhibitors verapamil, PSC833 and by energy depletion, demonstrating a direct
involvement for the functional involvement of Pgp. Surprisingly, Pgp also dem‑
onstrated broad specificity by translocating C6‑NBD‑SM and C6‑NBD‑GlcCer
(derived from the precursor C6‑NBD‑ceramide) and most natural versions of
these lipids lacking the NBD moiety.
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Essays in Biochemistry volume 50 2011
Short‑chain lipid analogues have also been studied with other tech‑
niques. Bosch et al. [67] demonstrated by fluorescence microscopy decreased
accumulation of NBD‑PC and NBD‑PE in multidrug resistant cell line
(CEM/VBL300) compared with the parental cell line (CEM, human
T‑lymphoblastic cell). Exogenous NBD‑PC, NBD‑PE and NBD‑PS were
added to CEM/VBL 300 and insect cells expressing recombinant Pgp. These
lipid compounds are internalized and eventually translocated to the plasma
membrane. Dithionite treatment was used to quench NBD fluorescence on
the outer leaflet of the plasma membrane. Quantification of internal cel‑
lular fluorescence was measured by FACS analysis. Drug‑resistant CEM/
VBL 300 cells accumulated approximately 10% of the amount of NBD‑PE
and NBD‑PC compared with drug‑sensitive CEM cells. No internal accu‑
mulation of NBD‑PS was found between drug‑resistant and drug‑sensitive
cell lines. Pgp reversal agents (verapamil, cyclosporin A and SDZ PSC 833)
both increase accumulation and inhibit efflux of these phospholipids in
CEM/VBL 300 cells, but not in CEM cells. The increased accumulation was
dose‑dependent, and the relative potency of the reversal agents substanti‑
ates the evidence that PC and PE, but not PS, behave as substrates for Pgp.
However, the possibility of NBD‑PS accumulating in mitochondrial mem‑
branes after internalization may explain its inaccessibility for Pgp efflux as
opposed to physicochemical differences.
The use of dithionite quenching of fluorescently labelled lipids for Pgp
in in vitro systems was first employed by Romsicki and Sharom [68]. The
distribution of NBD‑labelled lipids can be monitored between the inner and
outer leaflets of reconstituted proteoliposomes before and after incubation
with ATP. Addition of dithionite quenches the NBD lipids partitioned in
the outer leaflet and produces a stable baseline. Owing to the inaccessibility
of the fluorescently labelled lipids in the inner leaflet of vesicles to dithionite,
quantification of lipid distribution can be monitored in the presence of ATP,
drugs and inhibitors (Figure 5B). Purified Pgp reconstituted into proteolipo‑
somes was found to flip both short‑ and long‑chain NBD analogues of PC, PS
and PE in an ATP‑dependent and vanadate‑sensitive manner [68]. Similarly,
NBD‑GlcCer, NBD‑galactosylceramide and NBD‑SM flipping was also
observed by Pgp proteoliposomes and was inhibited by various drugs and
modulators in a concentration‑dependent manner [69].
The ATPase and transport activities of reconstituted protein are modulated
by the lipid environment in which it is embedded. The possibility that cholester‑
ol might serve as a substrate for Pgp was also studied. When NBD‑cholesterol
was used in the fluorescence‑based flippase assay, a transbilayer gradient of
the lipid was not observed after dithionite treatment in both PC liposomes
and Pgp proteoliposomes [70]. This behaviour arose from the rapid flip–flop
of NBD‑cholesterol between the inner and outer leaflet portions. A modest
decrease in NBD‑PC flippase activity was observed at 20–30% cholesterol, but
cholesterol itself was not a substrate for Pgp. Cholesterol content, however,
© The Authors Journal compilation © 2011 Biochemical Society
F. Quazi and R.S. Molday
281
has modest effects on both basal and drug‑­stimulated ATPase activity of Pgp.
In the presence of verapamil, Pgp reconstituted into PC proteoliposomes
exhibits a 2.2‑fold increase in ATPase stimulation. In the presence of 30%
(w/w) cholesterol, the stimulation is a modest decrease to 1.7‑fold. In addition,
it has no effect on Pgp conformation and neither does Pgp cross‑link with
[3,5,6‑3H]7‑azi‑5‑α‑cholestan‑3β‑ol, a photoactive cholesterol analogue, show‑
ing that it is not a direct substrate for Pgp. However, evidence from whole cell
assays support cholesterol translocation by Pgp. Wang et al. [71] reported that
in an NIH‑G‑185 cell line overexpressing Pgp, cholesterol caused a dramatic
inhibition of daunorobucin transport, yet had no effect in the parent cell line,
suggesting that cholesterol directly interacts with cholesterol transported by
Pgp [71]. Gayet et al. [72] found in human VEM acute lymphoblastic leukaemia
cells expressing Pgp, the amount of cholesterol increased linearly with the level
of resistance to vinblastine, whereas the amounts of total and free cholesterol
increased in a non‑linear manner.
Although fluorescently labelled lipids have been widely employed to
measure Pgp‑mediated lipid transport across membranes, such techniques
have not been successfully used to study lipid transport for ABCA proteins.
For the most part, transport of lipids by this class of ABC transporters has
been inferred from cell‑based assays or through the phenotypic analysis of
knockout mice.
It is unclear whether one can correlate ATPase stimulation as seen with
drug transport to identify lipid substrates. Reconstituted ABCG5/ABCG8
and ABCG2 display sterol uptake in reconstituted proteoliposomes in an
ATP‑dependent and vanadate‑sensitive manner. However, the ATPase activ‑
ity is slightly decreased, rather than stimulated by sterols [44,73]. Although
ABCA1 is a well‑accepted phospholipid transporter, its ability to transport
cholesterol directly is still unclear. First, cholesterol efflux from cells with
low ABCA1 expression was enhanced by cell culture media which were con‑
ditioned by pre‑incubation with ABCA1‑expressing cells [74]. Secondly, in
experiments with photoactivated lipids, ABCA1 was found to bind phospholi‑
pids, but not cholesterol, although overexpression of ABCA1 increased efflux
of photoactive cholesterol [74]. Thirdly, it has been reported that the ATPase
activity of ABCA1 shows robust ATPase activity (400–900 nmol/min per mg
of protein) when reconstituted into liposomes made of choline headgroups,
PC and SM [75]. Addition of cholesterol, β‑sitosterol and campesterol mildly
suppresses ABCA1 ATPase activity. Glibenclamide, an effective inhibitor
of apoA1‑dependent cellular cholesterol efflux suppresses the ATPase activ‑
ity of ABCA1 in a dose‑dependent manner [75]. These studies suggest that
sterols may affect membrane fluidity, which suppresses the ATPase activity
of ABCA1. It is not known whether sterol transfer and its lack of stimulation
in ATPase activity is an artefact of the systems used or an intrinsic property
of the sterol transporters. However, further studies are necessary to reveal a
direct biochemical interaction between cholesterol and sterol transporters.
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Export of sphingolipid metabolites and S1P
(sphingolipid 1‑phosphate)
ABCB1 [MDR1 (multidrug resistance 1)]
Sphingolipids comprise a vital fraction of membrane lipids in eukaryotic
cells. Individual sphingolipids are known to be signalling molecules and are
important mediators of survival, stress response and apoptosis. Sphingolipids
form a relatively non‑fluid membrane which can associate with cholesterol
to form microdomains commonly known as lipid rafts. The trafficking of
sphingolipids between organelles is also an important regulated process.
SM, sphingosine, ceramide and GlcCer are mainly localized in the plasma
membranes, but a significant fraction can be found in the Golgi and ER.
Among the ABCB subfamily, Pgp is rather promiscuous in its specificity:
besides analogues of PC, PE and PS, Pgp translocates short‑chain fluorescent
sphingolipid substrates across membranes [76]. It has been shown that the
ability of Pgp to transport drugs and act as a sphingolipid floppase occurs
via the same mechanism [47]. There is also evidence that trafficking of the
two sphingolipid species between the Golgi and plasma membrane may be
MDR1‑dependent [77]. Finally, Pgp also regulates the translocation of GlcCer
synthase activity and synthesis of complex glycosphingolipids [69].
ABCC1 (MRP1)
Like Pgp, ABCC1 has been reported to translocate short‑chain fluorescent
cholesterol, SM and GlcCer across membranes [49]. Both Pgp and ABCC1
transport hydrophobic cytotoxic drugs; Pgp transports compounds in an
unmodified state. In contrast, ABCC1 mainly transports substrates conjugated
to glutathione, glucuronide and sulfate [78].
ABCA and ABCG members
Abca2‑null mice exhibit low SM levels in oligodendrocytes, suggesting a
role for ABCA2 in the transport of SM and possibly other sphingolipids
[79]. SM is also a substrate of ABCG1, which translocates this lipid to HDL
particles [29]. Indeed, other ABCA proteins, including ABCA2, ABCA7 and
ABCA12 mediate transport of ceramide [79,80]. In keratinocytes, ABCA7
expression increases significantly during differentiation, coinciding with the
accumulation of ceramide, suggesting a link between ABCA7 and ceramide
levels [81]. Cultures of keratinocytes obtained from harlequin ichthyosis
patients (mutations in the ABCA12 gene) show abnormality in GlcCer
distribution, which is restored by genetic correction of the ABCA12 gene
[80]. Hence, ABCA12 may function as a ceramide transporter, but a direct
interaction between sphingolipids and ABCA7 or ABCA12 has not been
reported.
S1P exerts a wide range of actions in the nervous system. S1P receptors
are highly expressed in cerebellum; Anelli et al. [82] and Sato et al. [83] inves‑
tigated S1P release from astrocytes. On the basis of glyburide inhibition and
© The Authors Journal compilation © 2011 Biochemical Society
F. Quazi and R.S. Molday
283
siRNA (short interfering RNA) down‑regulation, translocation of S1P appears
to be highly dependent on ABCA1 expression [83]. Moreover, S1P release
from astrocytes is coupled with HDL formation [83]. Inhibitors of ABCB1
(Pgp) and ABCC1 (MRP1) (cyclosporin A and MK571 respectively) dimin‑
ish S1P release, suggesting that ABCA1, rather than ABCB1 or ABCC1, is
involved in the process [84].
Transport of pro‑apoptotic lipids by ABC transporters
Pgp
Modulation of lipid distribution in cells may also prevent cell death.
Cells induced to express Pgp either by drug stimulation or retroviral gene
transduction with Pgp cDNA are resistant to cell death induced by a range
of death stimuli such as FasL (Fas ligand), TNFα (tumour necrosis factor
α), UV irradiation and other factors that activate the caspase apoptotic
cascade. Overexpression of Pgp transporters in drug‑resistant cells is
accompanied by altered membrane content of cholesterol, SM, GlcCer and
other glycosphingolipids [85]. Interestingly, ceramide, a precursor of SM,
acts as an intracellular signalling molecule during apoptosis and is liberated
by apoptotic cascade factors. Pgp promotes SM externalization, resulting in
lower levels of SM available for ceramide synthesis on the intracellular side of
the membrane [86]. In response to treatment with PSC833, a Pgp inhibitor,
SM levels increase together with enhanced sphingomyelinase activity and
ceramide synthesis [87]. However, it is unclear whether PSC833 can act in a
Pgp‑independent manner.
ABCG2
ABCG2 (BCRP) has also been suggested to play an important role in cell
survival. Trophoblast cells, which typically express BCRP at high levels,
undergo loss of plasma membrane symmetry during cell fusion without
further progression to terminal phases of apoptosis. Suppression of BCRP
expression by siRNA leads to a marked increase in PS externalization fol‑
lowed by accumulation of ceramides and increased apoptosis. This can
occur as a result of cell–cell fusion during the differentiation process [88],
or in response to activation of the extrinsic apoptosis pathway by TNFα
and interferon‑γ [89]. However, whereas BCRP silencing was associated
with increased susceptibility to ceramide‑induced apoptosis and increased
intracellular ceramide levels, evidence of ceramide transport by BCRP was
not provided.
Oxidation products of cholesterol transport
In addition to cholesterol, ABCA1 and ABCG1 may prevent the accumulation
of cytotoxic cholesterol‑derived oxysterols in cells. Evidence from in vitro
studies indicate that oxidized LDLs contain oxysterols such as 25‑hydroxy‑ and
7‑oxo‑cholesterol, both abundant oxysterols in atherosclerotic plaques, can
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Essays in Biochemistry volume 50 2011
up‑regulate the expression of death mediators, including p53, Fas and FasL,
promote cytochrome c release and trigger an increase in intracellular calcium
concentrations. ABCA1 mediates high‑affinity uptake of 25‑hydroxycholesterol
in membrane vesicles (inside‑outside orientation) [90], whereas 7‑oxocholesterol
is effluxed by ABCG1 macrophages [91]. In addition, oxysterols are also
implicated in the cellular response to increased levels of cholesterol by binding
to and activating LXRs (liver X receptors). LXRs function as heterodimers with
the RXRs (retinoid X receptors) and induce transcription of genes involved in
cholesterol catabolism, including ABCA1 and ABCG1 [90] (Figure 3).
Phagocytosis
In mammals, PS externalization is associated with phagocytosis. Macrophages
deficient in ABCA1 fail to engulf apoptotic cells, whereas forced transfection
in non‑phagocytic HeLa cells promotes phagocytic behaviour [92]. At the
in vivo level, Abca1‑null mice exhibit transient accumulation of apoptotic
corpses in limb buds that can be seen during embryonic development [92].
Following phagocytosis of apoptotic cells, cholesterol efflux is enhanced in
an ABCA1‑dependent manner [93]. Indeed, cholesterol efflux may depend
on PS externalization, since PS vesicles are able to stimulate efflux. However,
annexin, a PS‑binding protein, neither competes with apoA1 for ABCA1
nor inhibits cholesterol efflux [94]. Hence the molecular control exerted by
ABCA1 in phagocytosis remains uncertain.
Conclusions
Lipid‑efflux activity of mammalian ABC transporters accomplishes a
wide variety and diverse range of activities related to homoeostasis and
cytoprotection. The structure and function of these transporters dictate
an intimate association with the constituents of the plasma membrane
lipid bilayer. Not surprisingly, some ABC transporters show preferential
localization to different organelles and, like other domain‑associated
proteins, their activity is affected by their surrounding lipid environment.
Hence they are well placed to regulate the efflux and redistribution of
lipids during periods of cellular division, stress and recovery, and, in turn,
are regulated by alterations in membrane lipid composition. A number
of ABC proteins are also able to transport ceramide and sphingolipids, as
well as phospholipids, which play important roles in cellular signalling,
differentiation and apoptosis.
There is strong indirect evidence that cholesterol serves as a substrate for sev‑
eral ABC transporters. ABCA1 mediates the transport of cholesterol and phos‑
pholipids such as PS from the inner to the outer leaflet of the membrane, but it
remains to be shown whether cholesterol and related sterols and phospholipids are
transported directly by ABCA1 and ABCG proteins. The involvement of ABCA1
in the reverse cholesterol pathway comprises distinct cellular events, including
intrinsic formation of cell‑surface lipid domains that interact with apolipoproteins,
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F. Quazi and R.S. Molday
285
solubilization of these lipid domains, direct binding of apoA1 to ABCA1, and
activation of signalling molecules. But the complexity and diversity of this process
is far from being understood. It is less likely that ABCA7 or ABCB1 play a major
role in cholesterol homoeostasis, but this does not exclude a more subtle role in
sterol trafficking. Several other species of sterols are also effluxed by ABC trans‑
porters, including toxic oxysterols, consistent with the view that ABC proteins
function to prevent build‑up of toxins and waste products.
Although the majority of ABC proteins transport substrates from the
cytoplasmic to the extracellular/lumen leaflet of membranes, several transport‑
ers, including Cdr3p, Aus1p, Pdr11p and ABCA4, have been implicated in
transport in the reverse direction. Additional biochemical transport assays are
needed to confirm the identity of physiological substrates and the direction
of transport by various ABC proteins. ABC lipid transporters expressed else‑
where in other organelles, including the Golgi and the ER, perform important
functions with respect to distribution of sphingolipids between the organelles.
Through regulation of ceramide and SM levels, and the availability of SM for
conversion into ceramide by sphingomyelinase, ABC transporters are able to
influence cellular sensitivity to apoptotic signals.
Further clarification of the mechanism of transfer of lipid substrates by
ABC transporters, and their regulation at both the transcriptional and activity
levels by lipid substrates is underway. These studies will enhance further our
understanding of the role ABC transporters in a wide variety of cellular pro‑
cesses and functions, and offer further insight into various lipid diseases.
Summary
•
•
•
•
•
Members of the ABC family of transporters are involved in multiple
aspects of lipid transport. Defective sterol and phospholipid transport
are linked to mutations in genes encoding ABCA1 (Tangier disease),
ABCA3 (fatal newborn surfactant disease), ABCA4 (Stargardt macu‑
lar degeneration), ABCA12 (harlequin ichthyosis), ABCB4 (PFIC) and
ABCG5/ABCG8 (sitosterolaemia).
A concerted model for cholesterol efflux has been proposed for ABCA1,
ABCG1 and ABCG4, in which initial lipidation of apoA1 to nascent
HDL is performed by ABCA1 followed by HDL maturation via
ABCG1/ABCG4.
Ceramide plays a major role in apoptosis. Increased turnover of cera‑
mide by modulating the plasma membrane pool of SM may allow
Pgp‑expressing cells to escape apoptosis.
Oxysterols may contribute to the regulation of their elimination by
inducing LXRs to increase expression of ABCA1 and ABCG1 genes.
Apoptotic cell engulfment and efflux of cellular lipids depend on
ABCA1‑induced perturbation of PS externalization.
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Essays in Biochemistry volume 50 2011
F.Q. is supported by a Natural Sciences and Engineering Research Council (NSERC)
graduate studentship. R.S.M. is a Canada Research Chair in Vision and Macular
Degeneration. This work was supported by the National Institutes of Health (NIH)
[grant number EY02422] and Canadian Institutes of Health Research (CIHR).
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