ATP-binding cassette transporter A7 (ABCA7) binds apolipoprotein A-I and mediates cellular phospholipid but not cholesterol efflux

ATP-binding cassette transporter 1 (ABCA1), the defective transporter in Tangier disease, binds and promotes cellular cholesterol and phospholipid efflux to apolipoprotein I (apoA-I). Based on a high degree of sequence homology between ABCA1 and ABCA7, a transporter of unknown function, we investigated the possibility that ABCA7 might be involved in apolipoprotein binding and lipid efflux. Similarly to cells expressing ABCA1, HEK293 cells overexpressing ABCA7 showed specific binding and cross-linking of lipid-poor apoA-I. ABCA7 expression increased cellular phosphatidylcholine and sphingomyelin efflux to apoA-I in a manner similar to ABCA1 but had no effect on cholesterol efflux. Western analysis showed a high protein level of ABCA7 in mouse spleen, lung, adrenal, and brain but low expression in liver. In contrast to ABCA1, ABCA7 showed moderate basal mRNA and protein levels in macrophages and lymphocytes but no induction by liver X receptor activation. These studies show that ABCA7 has the ability to bind apolipoproteins and promote efflux of cellular phospholipids without cholesterol, and they suggest a possible role of ABCA7 in cellular phospholipid metabolism in peripheral tissues.     

Molecular structure of a novel cholesterol-responsive A subclass ABC transporter,ABCA9

We recently identified a novel ABC A subclass transporter, ABCA6, in human macrophages. Here, we report the molecular cloning of an additional ABC A subfamily transporter from macrophages denoted ABCA9. The identified coding sequence is 4.9 kb in size and codes for a 1624 amino acid protein product. In accordance with the proposed nomenclature, the novel transporter was designated ABCA9. The putative full-length ABC transporter polypeptide consists of two transmembrane domains and two nucleotide binding folds and thus conforms to the group of full-size ABC transporters. We identified alternative ABCA9 mRNA variants in human macrophages that predict the existence of three truncated forms of the novel transporter. Among the human ABC A subfamily transporters, ABCA9 exhibits the highest amino acid sequence homology with ABCA8 (72%) and ABCA6 (60%), respectively. The striking amino acid sequence similarity between these transporter molecules supports the notion that they represent an evolutionary more recently emerged subgroup within the family of ABC A transporters, which we refer to as "ABCA6-like transporters." ABCA9 mRNA is ubiquitously expressed with the highest mRNA levels in heart, brain, and fetal tissues. Analysis of the genomic structure revealed that the ABCA9 gene consists of 39 exons that are located within a genomic region of approximately 85 kb size on chromosome 17q24.2. In human macrophages, ABCA9 mRNA is induced during monocyte differentiation into macrophages and suppressed by cholesterol import indicating that ABCA9, like other known ABC A subfamily transporters, is a cholesterol-responsive gene. Based on this information, ABCA9 is likely involved in monocyte differentiation and macrophage lipid homeostasis.     

ABCA17 mediates sterol efflux from mouse spermatozoa plasma membranes

Mammalian spermatozoa lose plasma membrane cholesterol during maturation in the epididymis and during capacitation in the female reproductive tract. While cholesterol acceptors such as high-density lipoproteins (HDL) and apolipoproteins A-I (apoA-I) and J (Apo J) have been found in male and female reproductive tracts, transporters that mediate cholesterol efflux from plasma membranes of spermatozoa to acceptors are not well defined. Candidates include members of the ATP-binding cassette (ABC) transporter superfamily including ABCA1, ABCA7, ABCA17, and ABCG1. In this study, we utilize immunocytochemistry on sections of adult mouse testis and epididymis and RT-PCR on isolated germ cells. The data reveal that ABCA17 is expressed by steps 12-16 elongated spermatids in the mouse in testis and by spermatozoa in the lumen of the epididymis where ABCA17 localizes to the sperm head and tail midpiece. It also localizes on these areas of mouse sperm isolated from the epididymis. Moreover, ABCA17 antibody interferes with cholesterol efflux from spermatozoa to lipid acceptors apoA-I. Taken together, these results suggest that ABCA17 plays an important role in the process of sterol efflux which renders spermatozoa capable of fertilizing an oocyte.     

Identification of a novel human sterol-sensitive ATP-binding cassette transporter (ABCA7)

We report the identification of the full-length cDNA for a novel ATP-binding cassette (ABC) transporter from human macrophages. The mRNA is of 6.8 kb size and contains an open reading frame encoding a polypeptide of 2146 amino acids with a calculated molecular weight of 220 kDa. The predicted protein product is composed of two transmembrane domains and two nucleotide binding folds indicating that it pertains to the group of full-size ABC transporters. The novel transporter shows highest protein sequence homology with the recently cloned human cholesterol and phospholipid exporter ABCA1 (54%) and the human retinal transporter ABCR (49%), both members of the ABC transporter subfamily A. In accordance with the currently proposed classification, the novel transporter was designated ABCA7. ABCA7 mRNA was detected predominantly in myelo-lymphatic tissues with highest expression in peripheral leukocytes, thymus, spleen, and bone marrow. Expression of ABCA7 is induced during in vitro differentiation of human monocytes into macrophages. In macrophages, both the ABCA7 mRNA and protein expression are upregulated in the presence of modified low density lipoprotein and downregulated by HDL(3). Our results suggest a role for ABCA7 in macrophage transmembrane lipid transport.     

Distinct spatio-temporal expression of ABCA and ABCG transporters in the developing and adult mouse brain

Using in situ hybridization for the mouse brain, we analyzed developmental changes in gene expression for the ATP-binding cassette (ABC) transporter subfamilies ABCA1-4 and 7, and ABCG1, 2, 4, 5 and 8. In the embryonic brains, ABCA1 and A7 were highly expressed in the ventricular (or germinal) zone, whereas ABCA2, A3 and G4 were enriched in the mantle (or differentiating) zone. At the postnatal stages, ABCA1 was detected in both the gray and white matter and in the choroid plexus. On the other hand, ABCA2, A3 and A7 were distributed in the gray matter. In addition, marked up-regulation of ABCA2 occurred in the white matter at 14 days-of-age when various myelin protein genes are known to be up-regulated. In marked contrast, ABCA4 was selective to the choroid plexus throughout development. ABCG1 was expressed in both the gray and white matters, whereas ABCG4 was confined to the gray matter. ABCG2 was diffusely and weakly detected throughout the brain at all stages examined. Immunohistochemistry of ABCG2 showed its preferential expression on the luminal membrane of brain capillaries. Expression signals for ABCG5 and G8 were barely detected at any stages. The distinct spatio-temporal expressions of individual ABCA and G transporters may reflect their distinct cellular expressions in the developing and adult brains, presumably, to regulate and maintain lipid homeostasis in the brain.     

Potential role of ABCA7 in cellular lipid efflux to apoA-I

ABCA7 is homologous to ABCA1 and has recently been shown in cell culture to bind apolipoprotein A-I (apoA-I) and to promote the efflux of phospholipids. However, it is not known if ABCA7 promotes lipid efflux in vivo. When expressed in HEK293 cells, both human and mouse ABCA7 promoted phospholipid efflux to apoA-I but no detectable cholesterol efflux. However, genetic knockdown of ABCA7 in mouse peritoneal macrophages did not affect phospholipid or cholesterol efflux to apoA-I. Moreover, in ABCA1-knockout macrophages, there was no detectable apoA-I-stimulated phospholipid efflux, inconsistent with a residual role of ABCA7. In contrast to plasma membrane localization of ABCA7 in transfected embryonic kidney cells, immunofluorescence microscopy of endogenous ABCA7 in macrophages showed a predominantly intracellular localization of the protein. Strikingly, immunofluorescence studies of adult mouse kidney revealed an apical brush border membrane localization of ABCA7 in the proximal tubule, suggesting that ABCA7 may come in contact with apoA-I in the glomerular filtrate. Although ABCA7 does not contribute to apolipoprotein-mediated lipid efflux in resting macrophages, its cell surface location in the kidney suggests that it could serve such a role in tissue microenvironments.     

Immunopathological consequences of the loss of engulfment genes: the case of ABCA1

Programmed cell death plays a crucial role in the maintenance of cell homeostasis. An initial, effector phase leads to the generation of apoptotic corpses and is closely followed by a swift clearance by professional or amateur phagocytes. Several aspects distinguish this latter process of engulfment of dying cells from the classical forms of phagocytosis. They concern all aspects of the process from the recognition of the prey to the final outcome, i.e. immunological silence. The engulfment of dead cells is a process highly conserved through evolution and it has been studied in parallel in two systems, mammalian cells and the nematode C. elegans. ABCA1 and its ortholog CED-7 in the nematode are key players of engulfment. Their mode of action is somehow original in the panorama of engulfment receptors since they act as lipid transporters. While in the worm the loss of CED-7 has phenotypic consequences exclusively on engulfment, in the mouse the deletion of ABCA1 by homologous recombination has highlighted broad consequences on macrophage biology. Among those we will discuss here the aberrant responses of ABCA1-/- mice to Plasmodium berghei ANKA infection, concerning in particular the development of cerebral malaria (CM), a cytokine-induced immunopathology. This syndrome involves a central role of monocytes and, as shown recently, high levels of circulating microparticles. It was found that ABCA1 loss completely protects against CM and its associated mortality. This observation, together with the demonstration of quantitative and functional modifications of microparticles, suggests that microparticles may be involved in CM pathogenesis. The ABCA1 transporter thus appears to control susceptibility to CM, thereby providing new insights in its pathophysiological mechanisms and potential new therapeutic avenues.     

Specificity of ABCA7-mediated cell lipid efflux

Adenosine triphosphate-binding cassette transporter subfamily A member 7 (ABCA7) performs incompletely understood biochemical functions that affect pathogenesis of Alzheimer's disease. ABCA7 is most similar in primary structure to ABCA1, the protein that mediates cell lipid efflux and formation of high-density lipoprotein (HDL). Lipid metabolic labeling/tracer efflux assays were employed to investigate lipid efflux in BHK-ABCA7(low expression), BHK-ABCA7(high expression) and BHK-ABCA1 cells. Shotgun lipid mass spectrometry was used to determine lipid composition of HDL synthesized by BHK-ABCA7 and BHK-ABCA1 cells. BHK-ABCA7(low) cells exhibited significant efflux only of choline-phospholipid and phosphatidylinositol. BHK-ABCA7(high) cells had significant cholesterol and choline-phospholipid efflux to apolipoprotein (apo) A-I, apo E, the 18A peptide, HDL, plasma and cerebrospinal fluid and significant efflux of sphingosine-lipid, serine-lipid (which is composed of phosphatidylserine and phosphatidylethanolamine in BHK cells) and phosphatidylinositol to apo A-I. In efflux assays to apo A-I, after adjustment to choline-phospholipid, ABCA7-mediated efflux removed ~4 times more serine-lipid and phosphatidylinositol than ABCA1-mediated efflux, while ABCA1-mediated efflux removed ~3 times more cholesterol than ABCA7-mediated efflux. Shotgun lipidomic analysis revealed that ABCA7-HDL had ~20 mol% less phosphatidylcholine and 3–5 times more serine-lipid and phosphatidylinositol than ABCA1-HDL, while ABCA1-HDL contained only ~6 mol% (or ~1.1 times) more cholesterol than ABCA7-HDL. The discrepancy between the tracer efflux assays and shotgun lipidomics with respect to cholesterol may be explained by an underestimate of ABCA7-mediated cholesterol efflux in the former approach. Overall, these results suggest that ABCA7 lacks specificity for phosphatidylcholine and releases significantly but not dramatically less cholesterol in comparison with ABCA1.     

Lysophosphatidylcholine export by human ABCA7

The ATP-binding cassette transporter A7 (ABCA7), which is highly expressed in the brain, is associated with the pathogenesis of Alzheimer's disease (AD). However, the physiological function of ABCA7 and its transport substrates remain unclear. Immunohistochemical analyses of human brain sections from AD and non-AD subjects revealed that ABCA7 is expressed in neuron and microglia cells in the cerebral cortex. The transport substrates and acceptors were identified in BHK/ABCA7 cells and compared with those of ABCA1. Like ABCA1, ABCA7 exported choline phospholipids in the presence of apoA-I and apoE; however, unlike ABCA1, cholesterol efflux was marginal. Lipid efflux by ABCA7 was saturated by 5 μg/ml apoA-I and was not dependent on apoE isoforms, whereas efflux by ABCA1 was dependent on apoA-I up to 20 μg/ml and apoE isoforms. Liquid chromatography–tandem mass spectrometry analyses revealed that the two proteins had different preferences for phospholipid export: ABCA7 preferred phosphatidylcholine (PC) ≥ lysoPC > sphingomyelin (SM) = phosphatidylethanolamine (PE), whereas ABCA1 preferred PC > > SM > PE = lysoPC. The major difference in the pattern of lipid peaks between ABCA7 and ABCA1 was the high lysoPC/PC ratio of ABCA7. These results suggest that lysoPC is one of the major transport substrates for ABCA7 and that lysoPC export may be a physiologically important function of ABCA7 in the brain.